ot Exponentially Modified Protein Abundance Index (emPAI) for Estimation of Absolute Protein Amount in Proteomics by the Number of Sequenced Peptides per Protein By feedproxy.google.com Published On :: 2005-09-01 Yasushi IshihamaSep 1, 2005; 4:1265-1272Research Full Article
ot Phosphate-binding Tag, a New Tool to Visualize Phosphorylated Proteins By feedproxy.google.com Published On :: 2006-04-01 Eiji KinoshitaApr 1, 2006; 5:749-757Technology Full Article
ot Multiplexed Protein Quantitation in Saccharomyces cerevisiae Using Amine-reactive Isobaric Tagging Reagents By feedproxy.google.com Published On :: 2004-12-01 Philip L. RossDec 1, 2004; 3:1154-1169Research Full Article
ot Accurate Proteome-wide Label-free Quantification by Delayed Normalization and Maximal Peptide Ratio Extraction, Termed MaxLFQ By feedproxy.google.com Published On :: 2014-09-01 Jürgen CoxSep 1, 2014; 13:2513-2526Technological Innovation and Resources Full Article
ot Stable Isotope Labeling by Amino Acids in Cell Culture, SILAC, as a Simple and Accurate Approach to Expression Proteomics By feedproxy.google.com Published On :: 2002-05-01 Shao-En OngMay 1, 2002; 1:376-386Research Full Article
ot The hibernating 100S complex is a target of ribosome-recycling factor and elongation factor G in Staphylococcus aureus [Protein Synthesis and Degradation] By feedproxy.google.com Published On :: 2020-05-01T00:06:09-07:00 The formation of translationally inactive 70S dimers (called 100S ribosomes) by hibernation-promoting factor is a widespread survival strategy among bacteria. Ribosome dimerization is thought to be reversible, with the dissociation of the 100S complexes enabling ribosome recycling for participation in new rounds of translation. The precise pathway of 100S ribosome recycling has been unclear. We previously found that the heat-shock GTPase HflX in the human pathogen Staphylococcus aureus is a minor disassembly factor. Cells lacking hflX do not accumulate 100S ribosomes unless they are subjected to heat exposure, suggesting the existence of an alternative pathway during nonstressed conditions. Here, we provide biochemical and genetic evidence that two essential translation factors, ribosome-recycling factor (RRF) and GTPase elongation factor G (EF-G), synergistically split 100S ribosomes in a GTP-dependent but tRNA translocation-independent manner. We found that although HflX and the RRF/EF-G pair are functionally interchangeable, HflX is expressed at low levels and is dispensable under normal growth conditions. The bacterial RRF/EF-G pair was previously known to target only the post-termination 70S complexes; our results reveal a new role in the reversal of ribosome hibernation that is intimately linked to bacterial pathogenesis, persister formation, stress responses, and ribosome integrity. Full Article
ot SUMOylation of the transcription factor ZFHX3 at Lys-2806 requires SAE1, UBC9, and PIAS2 and enhances its stability and function in cell proliferation [Protein Synthesis and Degradation] By feedproxy.google.com Published On :: 2020-05-08T03:41:14-07:00 SUMOylation is a posttranslational modification (PTM) at a lysine residue and is crucial for the proper functions of many proteins, particularly of transcription factors, in various biological processes. Zinc finger homeobox 3 (ZFHX3), also known as AT motif-binding factor 1 (ATBF1), is a large transcription factor that is active in multiple pathological processes, including atrial fibrillation and carcinogenesis, and in circadian regulation and development. We have previously demonstrated that ZFHX3 is SUMOylated at three or more lysine residues. Here, we investigated which enzymes regulate ZFHX3 SUMOylation and whether SUMOylation modulates ZFHX3 stability and function. We found that SUMO1, SUMO2, and SUMO3 each are conjugated to ZFHX3. Multiple lysine residues in ZFHX3 were SUMOylated, but Lys-2806 was the major SUMOylation site, and we also found that it is highly conserved among ZFHX3 orthologs from different animal species. Using molecular analyses, we identified the enzymes that mediate ZFHX3 SUMOylation; these included SUMO1-activating enzyme subunit 1 (SAE1), an E1-activating enzyme; SUMO-conjugating enzyme UBC9 (UBC9), an E2-conjugating enzyme; and protein inhibitor of activated STAT2 (PIAS2), an E3 ligase. Multiple analyses established that both SUMO-specific peptidase 1 (SENP1) and SENP2 deSUMOylate ZFHX3. SUMOylation at Lys-2806 enhanced ZFHX3 stability by interfering with its ubiquitination and proteasomal degradation. Functionally, Lys-2806 SUMOylation enabled ZFHX3-mediated cell proliferation and xenograft tumor growth of the MDA-MB-231 breast cancer cell line. These findings reveal the enzymes involved in, and the functional consequences of, ZFHX3 SUMOylation, insights that may help shed light on ZFHX3's roles in various cellular and pathophysiological processes. Full Article
ot {gamma}-Hydroxybutyrate does not mediate glucose inhibition of glucagon secretion [Signal Transduction] By feedproxy.google.com Published On :: 2020-04-17T00:06:05-07:00 Hypersecretion of glucagon from pancreatic α-cells strongly contributes to diabetic hyperglycemia. Moreover, failure of α-cells to increase glucagon secretion in response to falling blood glucose concentrations compromises the defense against hypoglycemia, a common complication in diabetes therapy. However, the mechanisms underlying glucose regulation of glucagon secretion are poorly understood and likely involve both α-cell–intrinsic and intraislet paracrine signaling. Among paracrine factors, glucose-stimulated release of the GABA metabolite γ-hydroxybutyric acid (GHB) from pancreatic β-cells might mediate glucose suppression of glucagon release via GHB receptors on α-cells. However, the direct effects of GHB on α-cell signaling and glucagon release have not been investigated. Here, we found that GHB (4–10 μm) lacked effects on the cytoplasmic concentrations of the secretion-regulating messengers Ca2+ and cAMP in mouse α-cells. Glucagon secretion from perifused mouse islets was also unaffected by GHB at both 1 and 7 mm glucose. The GHB receptor agonist 3-chloropropanoic acid and the antagonist NCS-382 had no effects on glucagon secretion and did not affect stimulation of secretion induced by a drop in glucose from 7 to 1 mm. Inhibition of endogenous GHB formation with the GABA transaminase inhibitor vigabatrin also failed to influence glucagon secretion at 1 mm glucose and did not prevent the suppressive effect of 7 mm glucose. In human islets, GHB tended to stimulate glucagon secretion at 1 mm glucose, an effect mimicked by 3-chloropropanoic acid. We conclude that GHB does not mediate the inhibitory effect of glucose on glucagon secretion. Full Article
ot Heterotrimeric Gq proteins as therapeutic targets? [Molecular Bases of Disease] By feedproxy.google.com Published On :: 2020-04-17T00:06:05-07:00 Heterotrimeric G proteins are the core upstream elements that transduce and amplify the cellular signals from G protein–coupled receptors (GPCRs) to intracellular effectors. GPCRs are the largest family of membrane proteins encoded in the human genome and are the targets of about one-third of prescription medicines. However, to date, no single therapeutic agent exerts its effects via perturbing heterotrimeric G protein function, despite a plethora of evidence linking G protein malfunction to human disease. Several recent studies have brought to light that the Gq family–specific inhibitor FR900359 (FR) is unexpectedly efficacious in silencing the signaling of Gq oncoproteins, mutant Gq variants that mostly exist in the active state. These data not only raise the hope that researchers working in drug discovery may be able to potentially strike Gq oncoproteins from the list of undruggable targets, but also raise questions as to how FR achieves its therapeutic effect. Here, we place emphasis on these recent studies and explain why they expand our pharmacological armamentarium for targeting Gq protein oncogenes as well as broaden our mechanistic understanding of Gq protein oncogene function. We also highlight how this novel insight impacts the significance and utility of using G(q) proteins as targets in drug discovery efforts. Full Article
ot Reduction of protein phosphatase 2A (PP2A) complexity reveals cellular functions and dephosphorylation motifs of the PP2A/B'{delta} holoenzyme [Enzymology] By feedproxy.google.com Published On :: 2020-04-24T06:08:45-07:00 Protein phosphatase 2A (PP2A) is a large enzyme family responsible for most cellular Ser/Thr dephosphorylation events. PP2A substrate specificity, localization, and regulation by second messengers rely on more than a dozen regulatory subunits (including B/R2, B'/R5, and B″/R3), which form the PP2A heterotrimeric holoenzyme by associating with a dimer comprising scaffolding (A) and catalytic (C) subunits. Because of partial redundancy and high endogenous expression of PP2A holoenzymes, traditional approaches of overexpressing, knocking down, or knocking out PP2A regulatory subunits have yielded only limited insights into their biological roles and substrates. To this end, here we sought to reduce the complexity of cellular PP2A holoenzymes. We used tetracycline-inducible expression of pairs of scaffolding and regulatory subunits with complementary charge-reversal substitutions in their interaction interfaces. For each of the three regulatory subunit families, we engineered A/B charge–swap variants that could bind to one another, but not to endogenous A and B subunits. Because endogenous Aα was targeted by a co-induced shRNA, endogenous B subunits were rapidly degraded, resulting in expression of predominantly a single PP2A heterotrimer composed of the A/B charge–swap pair and the endogenous catalytic subunit. Using B'δ/PPP2R5D, we show that PP2A complexity reduction, but not PP2A overexpression, reveals a role of this holoenzyme in suppression of extracellular signal–regulated kinase signaling and protein kinase A substrate dephosphorylation. When combined with global phosphoproteomics, the PP2A/B'δ reduction approach identified consensus dephosphorylation motifs in its substrates and suggested that residues surrounding the phosphorylation site play roles in PP2A substrate specificity. Full Article
ot G{alpha}q splice variants mediate phototransduction, rhodopsin synthesis, and retinal integrity in Drosophila [Signal Transduction] By feedproxy.google.com Published On :: 2020-04-24T06:08:45-07:00 Heterotrimeric G proteins mediate a variety of signaling processes by coupling G protein–coupled receptors to intracellular effector molecules. In Drosophila, the Gαq gene encodes several Gαq splice variants, with the Gαq1 isoform protein playing a major role in fly phototransduction. However, Gαq1 null mutant flies still exhibit a residual light response, indicating that other Gαq splice variants or additional Gq α subunits are involved in phototransduction. Here, we isolated a mutant fly with no detectable light responses, decreased rhodopsin (Rh) levels, and rapid retinal degeneration. Using electrophysiological and genetic studies, biochemical assays, immunoblotting, real-time RT-PCR, and EM analysis, we found that mutations in the Gαq gene disrupt light responses and demonstrate that the Gαq3 isoform protein is responsible for the residual light response in Gαq1 null mutants. Moreover, we report that Gαq3 mediates rhodopsin synthesis. Depletion of all Gαq splice variants led to rapid light-dependent retinal degeneration, due to the formation stable Rh1-arrestin 2 (Arr2) complexes. Our findings clarify essential roles of several different Gαq splice variants in phototransduction and retinal integrity in Drosophila and reveal that Gαq3 functions in rhodopsin synthesis. Full Article
ot Mechanistic insights explain the transforming potential of the T507K substitution in the protein-tyrosine phosphatase SHP2 [Signal Transduction] By feedproxy.google.com Published On :: 2020-05-01T00:06:09-07:00 The protein-tyrosine phosphatase SHP2 is an allosteric enzyme critical for cellular events downstream of growth factor receptors. Mutations in the SHP2 gene have been linked to many different types of human diseases, including developmental disorders, leukemia, and solid tumors. Unlike most SHP2-activating mutations, the T507K substitution in SHP2 is unique in that it exhibits oncogenic Ras-like transforming activity. However, the biochemical basis of how the SHP2/T507K variant elicits transformation remains unclear. By combining kinetic and biophysical methods, X-ray crystallography, and molecular modeling, as well as using cell biology approaches, here we uncovered that the T507K substitution alters both SHP2 substrate specificity and its allosteric regulatory mechanism. We found that although SHP2/T507K exists in the closed, autoinhibited conformation similar to the WT enzyme, the interactions between its N-SH2 and protein-tyrosine phosphatase domains are weakened such that SHP2/T507K possesses a higher affinity for the scaffolding protein Grb2-associated binding protein 1 (Gab1). We also discovered that the T507K substitution alters the structure of the SHP2 active site, resulting in a change in SHP2 substrate preference for Sprouty1, a known negative regulator of Ras signaling and a potential tumor suppressor. Our results suggest that SHP2/T507K's shift in substrate specificity coupled with its preferential association of SHP2/T507K with Gab1 enable the mutant SHP2 to more efficiently dephosphorylate Sprouty1 at pTyr-53. This dephosphorylation hyperactivates Ras signaling, which is likely responsible for SHP2/T507K's Ras-like transforming activity. Full Article
ot DHHC7-mediated palmitoylation of the accessory protein barttin critically regulates the functions of ClC-K chloride channels [Cell Biology] By feedproxy.google.com Published On :: 2020-05-01T00:06:09-07:00 Barttin is the accessory subunit of the human ClC-K chloride channels, which are expressed in both the kidney and inner ear. Barttin promotes trafficking of the complex it forms with ClC-K to the plasma membrane and is involved in activating this channel. Barttin undergoes post-translational palmitoylation that is essential for its functions, but the enzyme(s) catalyzing this post-translational modification is unknown. Here, we identified zinc finger DHHC-type containing 7 (DHHC7) protein as an important barttin palmitoyl acyltransferase, whose depletion affected barttin palmitoylation and ClC-K-barttin channel activation. We investigated the functional role of barttin palmitoylation in vivo in Zdhhc7−/− mice. Although palmitoylation of barttin in kidneys of Zdhhc7−/− animals was significantly decreased, it did not pathologically alter kidney structure and functions under physiological conditions. However, when Zdhhc7−/− mice were fed a low-salt diet, they developed hyponatremia and mild metabolic alkalosis, symptoms characteristic of human Bartter syndrome (BS) type IV. Of note, we also observed decreased palmitoylation of the disease-causing R8L barttin variant associated with human BS type IV. Our results indicate that dysregulated DHHC7-mediated barttin palmitoylation appears to play an important role in chloride channel dysfunction in certain BS variants, suggesting that targeting DHHC7 activity may offer a potential therapeutic strategy for reducing hypertension. Full Article
ot The focal adhesion protein kindlin-2 controls mitotic spindle assembly by inhibiting histone deacetylase 6 and maintaining {alpha}-tubulin acetylation [Signal Transduction] By feedproxy.google.com Published On :: 2020-05-01T00:06:09-07:00 Kindlins are focal adhesion proteins that regulate integrin activation and outside-in signaling. The kindlin family consists of three members, kindlin-1, -2, and -3. Kindlin-2 is widely expressed in multiple cell types, except those from the hematopoietic lineage. A previous study has reported that the Drosophila Fit1 protein (an ortholog of kindlin-2) prevents abnormal spindle assembly; however, the mechanism remains unknown. Here, we show that kindlin-2 maintains spindle integrity in mitotic human cells. The human neuroblastoma SH-SY5Y cell line expresses only kindlin-2, and we found that when SH-SY5Y cells are depleted of kindlin-2, they exhibit pronounced spindle abnormalities and delayed mitosis. Of note, acetylation of α-tubulin, which maintains microtubule flexibility and stability, was diminished in the kindlin-2–depleted cells. Mechanistically, we found that kindlin-2 maintains α-tubulin acetylation by inhibiting the microtubule-associated deacetylase histone deacetylase 6 (HDAC6) via a signaling pathway involving AKT Ser/Thr kinase (AKT)/glycogen synthase kinase 3β (GSK3β) or paxillin. We also provide evidence that prolonged hypoxia down-regulates kindlin-2 expression, leading to spindle abnormalities not only in the SH-SY5Y cell line, but also cell lines derived from colon and breast tissues. The findings of our study highlight that kindlin-2 regulates mitotic spindle assembly and that this process is perturbed in cancer cells in a hypoxic environment. Full Article
ot Polarization of protease-activated receptor 2 (PAR-2) signaling is altered during airway epithelial remodeling and deciliation [Immunology] By feedproxy.google.com Published On :: 2020-05-08T03:41:14-07:00 Protease-activated receptor 2 (PAR-2) is activated by secreted proteases from immune cells or fungi. PAR-2 is normally expressed basolaterally in differentiated nasal ciliated cells. We hypothesized that epithelial remodeling during diseases characterized by cilial loss and squamous metaplasia may alter PAR-2 polarization. Here, using a fluorescent arrestin assay, we confirmed that the common fungal airway pathogen Aspergillus fumigatus activates heterologously-expressed PAR-2. Endogenous PAR-2 activation in submerged airway RPMI 2650 or NCI–H520 squamous cells increased intracellular calcium levels and granulocyte macrophage–colony-stimulating factor, tumor necrosis factor α, and interleukin (IL)-6 secretion. RPMI 2650 cells cultured at an air–liquid interface (ALI) responded to apically or basolaterally applied PAR-2 agonists. However, well-differentiated primary nasal epithelial ALIs responded only to basolateral PAR-2 stimulation, indicated by calcium elevation, increased cilia beat frequency, and increased fluid and cytokine secretion. We exposed primary cells to disease-related modifiers that alter epithelial morphology, including IL-13, cigarette smoke condensate, and retinoic acid deficiency, at concentrations and times that altered epithelial morphology without causing breakdown of the epithelial barrier to model early disease states. These altered primary cultures responded to both apical and basolateral PAR-2 stimulation. Imaging nasal polyps and control middle turbinate explants, we found that nasal polyps, but not turbinates, exhibit apical calcium responses to PAR-2 stimulation. However, isolated ciliated cells from both polyps and turbinates maintained basolateral PAR-2 polarization, suggesting that the calcium responses originated from nonciliated cells. Altered PAR-2 polarization in disease-remodeled epithelia may enhance apical responses and increase sensitivity to inhaled proteases. Full Article
ot Phosphoproteomic characterization of the signaling network resulting from activation of the chemokine receptor CCR2 [Genomics and Proteomics] By feedproxy.google.com Published On :: 2020-05-08T03:41:14-07:00 Leukocyte recruitment is a universal feature of tissue inflammation and regulated by the interactions of chemokines with their G protein–coupled receptors. Activation of CC chemokine receptor 2 (CCR2) by its cognate chemokine ligands, including CC chemokine ligand 2 (CCL2), plays a central role in recruitment of monocytes in several inflammatory diseases. In this study, we used phosphoproteomics to conduct an unbiased characterization of the signaling network resulting from CCL2 activation of CCR2. Using data-independent acquisition MS analysis, we quantified both the proteome and phosphoproteome in FlpIn-HEK293T cells stably expressing CCR2 at six time points after activation with CCL2. Differential expression analysis identified 699 significantly regulated phosphorylation sites on 441 proteins. As expected, many of these proteins are known to participate in canonical signal transduction pathways and in the regulation of actin cytoskeleton dynamics, including numerous guanine nucleotide exchange factors and GTPase-activating proteins. Moreover, we identified regulated phosphorylation sites in numerous proteins that function in the nucleus, including several constituents of the nuclear pore complex. The results of this study provide an unprecedented level of detail of CCR2 signaling and identify potential targets for regulation of CCR2 function. Full Article
ot What Boris Johnson’s Big Win Means for Brexit and Scotland By feedproxy.google.com Published On :: Fri, 13 Dec 2019 14:25:22 +0000 13 December 2019 Thomas Raines Director, Europe Programme @TomHRaines Google Scholar Jason Naselli Senior Digital Editor LinkedIn Thomas Raines tells Jason Naselli about the impact the large Conservative majority will have on the next phase of Brexit negotiations and Scotland’s place in the United Kingdom. 2019-12-13-Johnson.jpg Boris Johnson speaks after the Conservatives secured a majority in the UK general election. Photo: Getty Images. What does the UK election result mean for Brexit and forthcoming trade negotiations with the EU?The most important thing is that it means Brexit will definitely happen. Since the referendum, we’ve had three-and-a-half years of continued uncertainty where all outcomes were still possible. We now know that Brexit will become irreversible from 31 January.That’s the biggest thing, because I think that will have a big psychological impact on politics, both in the UK and also on the EU side. The EU has been working with a partner that has been unsure about its direction, and perhaps some had still hoped that the process might still be reversed, but that direction is now completely clear.Obviously the first order of business is to pass the withdrawal agreement, which should be pretty straightforward given the majority that the Conservatives have. That’s a formality now.Then, the question becomes about the level of ambition for the next year. It is an exceptionally ambitious timetable to negotiate, ratify and implement a new relationship before the end of the transition period in December 2020.What is achievable by the end of next year? I think there are three possible outcomes here. One: that timetable doesn’t work and Boris Johnson follows through on his pledge to leave the transition period anyway, leading to a ‘no trade deal’ outcome.Two: the negotiations are able to deliver something by the end of 2020, either because the depth and ambition of any agreement is relatively low level (what Michel Barnier has called a ‘vital minimum’) and/or because they come up with some type of compromise on the process which is not called an extension, but something else: a type of temporary agreement or a new implementation period.This is a situation where you might have a bare-bones agreement for the end of the transition period, but with an extended period of negotiation for different unresolved issues. The EU will probably insist upon level playing field guarantees and fishing access rights as a component of any such agreement.Three: Boris Johnson breaks his manifesto pledge not to extend the transition. Now, he has stared down the barrel of leaving with no deal before and he made a political judgment that it was better to make significant compromises on his negotiating position than to follow through with ‘no deal’. I suspect he might make that same judgment again.No option is ideal. The first is the most economically disruptive, the second means the EU will be in an even stronger position to dictate terms and the third means breaking a manifesto pledge.How important is that end of transition deadline now? It was an important issue for the Brexit Party and hardliners in the European Research Group of Conservative MPs. But given the size of a majority, he may need to worry less about them. Is the transition deadline that important to people who voted Conservative, especially if he can show that he has taken the UK out of the EU by the end of January?I think there may indeed be some political space for Johnson here, given the size of his majority and given that the first phase of Brexit will have been done, along with the debate about withdrawal.There will be a lot of difficult, technical negotiations in all sorts of areas, some of which I think will become quite rancorous, but won’t necessarily become front page news in the way some of the first phase of negotiations has, not least because you won’t have the theatre of a hung parliament.Hopefully, there will be more focus on the substance of the agreement, and the debate will be about the consequences of divergence versus staying more aligned with the EU, which is basically the central question now about the future relationship.I still think for UK prime ministers to pick arbitrary dates, and then to make domestic political promises based around them, actually undermines the UK’s negotiating position. It would be in Britain’s interest to have more flexibility rather than a ticking clock.Moving to the other big story from the night, the SNP won 48 of 59 seats in Scotland. How does the debate over Scotland’s future in the United Kingdom play out from here?The SNP has really strengthened its position, more than many expected. This is now set up for a huge constitutional struggle over the future of the United Kingdom.I think there is a key dilemma for Scottish independence supporters, which is that on the one hand Brexit greatly strengthens the political case for independence. The difference between the political preferences in Scotland and the rest of the UK, particularly in England, is a perfect demonstration of that.At the same time, once the UK has left the EU, independence becomes much more difficult technically and economically. There will be many of the same difficulties that there have been in discussing Northern Ireland’s relationship with the Republic of Ireland. There will be a difficult debate over the currency. There are all sorts of challenges to creating a trade or regulatory border between England and Scotland. This is particularly true if there is a harder Brexit outcome, where Britain leaves the EU without a large amount of regulatory alignment.On demands for a second independence referendum, I think in the first instance Boris Johnson will simply refuse to hold one. It’s probably not in his short-term interest to do anything else. Theresa May played it this way in 2017, repeatedly saying ‘now is not the time’.In a similar way I think Johnson will just try to ride the pressure out, to the point where the SNP will need to face the challenges of advocating independence with the UK outside the EU. The next flashpoint will be the elections to the Scottish Parliament in 2021.Ultimately, though, it will become a democratically unsustainable position if Scotland continues to vote for the SNP, and refusing to sanction a second independence referendum might only reinforce that sentiment.Follow Chatham House Twitter for more election coverage Full Article
ot Nkirote Laiboni By feedproxy.google.com Published On :: Wed, 15 Jan 2020 16:12:35 +0000 Robert Bosch Stiftung Academy Fellow (Central & Eastern Europe and Africa), Europe Programme Biography Nkirote Laiboni is a Kenyan international development and human rights practitioner with over 10 years of experience in managing humanitarian, human rights and international development programmes and coordinating research projects in Sub-Saharan Africa.She has worked with international nongovernmental organisations on a wide range of sectors and issues including migration, humanitarian support, media development, democracy and governance, education and health.Having previously worked on projects benefiting refugees, internally displaced persons, migrant workers, and trafficked persons in Kenya, Uganda, South Sudan and other African countries, Nkirote’s professional interest in migration has grown in recent years. Her research at Chatham House will explore and compare labour migration trends and patterns in Eastern Africa and Central and Eastern Europe.She holds an MA in International Law and International Human Rights from the University of Peace, Costa Rica. Areas of expertise Non-profit managementProgramme and project managementParticipatory action researchMigration and human rightsInternational developmentHumanitarian support Email LinkedIn Full Article
ot In Next Round of EU Negotiations, Britain Faces Familiar Pitfalls By feedproxy.google.com Published On :: Fri, 31 Jan 2020 14:33:27 +0000 31 January 2020 Thomas Raines Director, Europe Programme @TomHRaines Google Scholar Professor Richard G Whitman Associate Fellow, Europe Programme @RGWhitman Google Scholar Despite being free of the constraints and the theatre of a hung parliament, there is a risk that over the coming year the British government repeats too many of the mistakes of the withdrawal negotiations. 2020-01-31-Ldn.jpg The Elizabeth Tower remains under renovation on 31 January 2020. Photo: Getty Images. Whether feared or longed for, Brexit day has arrived. It is positive for all sides that the process is thus far managed and ordered, with debts paid, rights guaranteed and borders still invisible on the island of Ireland. But in a difficult new phase of negotiations, as the UK and EU try to hammer out the terms of their relationship after 2020, Britain is at risk of repeating many of its mistakes from the withdrawal talks.First, the government, through the negotiation timeline, has reduced its own room for manoeuvre. The failure of the initial withdrawal agreement and subsequent turbulent politics have reduced a planned 21-month transition to an 11-month one. Even though half the original negotiation time has been lost, 31 December 2020 remains in place and indeed has been written into UK law as the date the transition arrangement ends. Boris Johnson has followed Theresa May in investing symbolism and significance in an arbitrary date.By promising not to extend negotiations, the UK is boxing itself in, creating domestic political barriers where it may well need flexibility. The familiar face of Michel Barnier, who proved adept in leading the withdrawal negotiations for the EU side, will be back in phase two to tell Britain the clock is ticking. This new timeline is intended to focus minds but more likely it will limit ambitions. Second, this government has continued the pattern of its predecessor in making no effort to manage public expectations about the consequences of Brexit. It is naïve to have followed the last years of British politics and expect an outbreak of sobriety and levelheadedness. The entrenched positions of each side have offered little political space or electoral incentive for realism.During the 2020 transition period, the UK will lose the political rights of EU membership but it will retain the benefits and obligations. Most citizens and business will not be able to tell the difference. But a reckoning is inevitable. There will come a moment when the effects of this slow-motion political revolution – particularly in the hard form envisioned by Boris Johnson – become real, when the trade-offs and compromises, especially for business and the economy, will bite. The public deserve some realism about the price of sovereignty.Third, there is a risk that government remains underprepared. While its headline goals are clear – at least in terms of what it does not want – the UK government will need thorough, realistic and coherent proposals on what it wants in every area of negotiations, and crucially develop a process by which to make political trade-offs between the demands of different sectors and issues. The government must also then prepare for their implementation in every area. This would be a huge challenge even if the final destination was already known, which it is not. Fourth, the continued uncertainty in the process means businesses and civil servants will again be left with little time to adapt to what will face them in January 2021 and must prepare for multiple outcomes.‘Transition’ has always been a misleading term, since it implies clarity about the destination to which the UK–EU relationship will be transitioning. The government’s red lines for that future relationship provide a sketch: outside of the single market and the jurisdiction of the European Court of Justice, with an independent trade policy and free movement ended.But businesses and civil servants are not likely to know until very late in the process if the basis for future trade with the EU will be in the form of a free trade agreement, to be negotiated and implemented by the end of the year, or no trade deal at all. This last outcome is a realistic prospect. Michel Barnier speaks in the European Parliament on 29 January. Photo: Getty Images. During withdrawal negotiations, the extensions were both unlimited in number and required decisions only at the last moment. In this phase, the talks may only be extended once, and that decision must be taken six months from the final deadline. It is difficult to see circumstances in which Boris Johnson agrees to break a political promise and manifesto pledge when he still has six more months to achieve his desired outcome.The UK, it is often noted, is already fully compliant with EU law and this shared starting point is often cited as a reason this negotiation will be simple, since the parties begin in alignment. But this novel negotiation will create new trade barriers in goods and services rather than remove them. Trade deals are often politically difficult since they create winners and losers. The Brexit negotiations, in terms of UK–EU trade at least, will generally create only different levels of losers, on both sides of the Channel.That means difficult politics, challenging negotiations and hard compromises, another reason to expect some ugly politics along the way and accept that failure is a plausible outcome.We do not yet know how Brexit will change Britain in the long term, whether a settled majority will ever come to view it as political folly or liberation, choice or inevitability. If its politically fragile union can withstand the pressures of the next few years, the UK may yet find a new stable position on the EU’s periphery and, after a period of economic adjustment, begin to address the many pressing domestic challenges which have suffered from neglect amid the all-consuming Brexit saga.But whatever happens in this next chapter, the EU can no longer be an excuse for national problems. As the UK takes back control it also returns accountability. In the future, there will be no one else to credit or to blame. Full Article
ot The EU Cannot Build a Foreign Policy on Regulatory Power Alone By feedproxy.google.com Published On :: Tue, 11 Feb 2020 16:33:26 +0000 11 February 2020 Alan Beattie Associate Fellow, Global Economy and Finance Programme and Europe Programme @alanbeattie LinkedIn Brussels will find its much-vaunted heft in setting standards cannot help it advance its geopolitical interests. 2020-02-11-Leyen.jpg EU Commission President Ursula von der Leyen speaks at the European Parliament in Strasbourg in February. Photo: Getty Images. There are two well-established ideas in trade. Individually, they are correct. Combined, they can lead to a conclusion that is unfortunately wrong.The first idea is that, across a range of economic sectors, the EU and the US have been engaged in a battle to have their model of regulation accepted as the global one, and that the EU is generally winning.The second is that governments can use their regulatory power to extend strategic and foreign policy influence.The conclusion would seem to be that the EU, which has for decades tried to develop a foreign policy, should be able to use its superpower status in regulation and trade to project its interests and its values abroad.That’s the theory. It’s a proposition much welcomed by EU policymakers, who know they are highly unlikely any time soon to acquire any of the tools usually required to run an effective foreign policy.The EU doesn’t have an army it can send into a shooting war, enough military or political aid to prop up or dispense of governments abroad, or a centralized intelligence service. Commission President Ursula von der Leyen has declared her outfit to be a ‘geopolitical commission’, and is casting about for any means of making that real.Through the ‘Brussels effect’ whereby European rules and standards are exported via both companies and governments, the EU has indeed won many regulatory battles with the US.Its cars, chemicals and product safety regulations are more widely adopted round the world than their American counterparts. In the absence of any coherent US offering, bar some varied state-level systems, the General Data Protection Regulation (GDPR) is the closest thing the world has to a single model for data privacy, and variants of it are being adopted by dozens of countries.The problem is this. Those parts of global economic governance where the US is dominant – particularly the dollar payments system – are highly conducive to projecting US power abroad. The extraterritorial reach of secondary sanctions, plus the widespread reliance of banks and companies worldwide on dollar funding – and hence the American financial system – means that the US can precisely target its influence.The EU can enforce trade sanctions, but not in such a powerful and discriminatory way, and it will always be outgunned by the US. Donald Trump could in effect force European companies to join in his sanctions on Iran when he pulled out of the nuclear deal, despite EU legislation designed to prevent their businesses being bullied. He can go after the chief financial officer of Huawei for allegedly breaching those sanctions.By contrast, the widespread adoption of GDPR or data protection regimes inspired by it may give the EU a warm glow of satisfaction, but it cannot be turned into a geopolitical tool in the same way.Nor, necessarily, does it particularly benefit the EU economy. Europe’s undersized tech sector seems unlikely to unduly benefit from the fact that data protection rules were written in the EU. Indeed, one common criticism of the regulations is that they entrench the power of incumbent tech giants like Google.There is a similar pattern at work in the adoption of new technologies such as artificial intelligence and the Internet of Things. In that field, the EU and its member states are also facing determined competition from China, which has been pushing its technologies and standards through forums such as the International Telecommunication Union.The EU has been attempting to write international rules for the use of AI which it hopes to be widely adopted. But again, these are a constraint on the use of new technologies largely developed by others, not the control of innovation.By contrast, China has created a vast domestic market in technologies like facial recognition and unleashed its own companies on it. The resulting surveillance kit can then be marketed to emerging market governments as part of China’s enduring foreign policy campaign to build up supporters in the developing world.If it genuinely wants to turn its economic power into geopolitical influence – and it’s not entirely clear what it would do with it if it did – the EU needs to recognize that not all forms of regulatory and trading dominance are the same.Providing public goods to the world economy is all very well. But unless they are so particular in nature that they project uniquely European values and interests, that makes the EU a supplier of useful plumbing but not a global architect of power.On the other hand, it could content itself with its position for the moment. It could recognize that not until enough hard power – guns, intelligence, money – is transferred from the member states to the centre, or until the member states start acting collectively, will the EU genuinely become a geopolitical force. Speaking loudly and carrying a stick of foam rubber is rarely a way to gain credibility in international relations.This article is part of a series of publications and roundtable discussions in the Chatham House Global Trade Policy Forum. Full Article
ot Another CDU Leadership Race Begins in Merkel’s Shadow By feedproxy.google.com Published On :: Fri, 28 Feb 2020 13:59:32 +0000 28 February 2020 Quentin Peel Associate Fellow, Europe Programme @QuentinPeel The election of a new leader of the chancellor’s party will be another contest over her legacy. 2020-02-28-Merkel.jpg German Chancellor Angela Merkel is depicted on a float in the Rosenmontag parade in Mainz on 24 February. Photo: Getty Images. Perhaps it will be second time lucky. At the end of April, Germany’s Christian Democratic Union (CDU) will elect a new party leader to follow in the footsteps of Angela Merkel. An emergency party congress has been summoned to do that after the surprise resignation of Annegret Kramp-Karrenbauer, Merkel’s chosen successor.The plan is to leave the decision on who will be the CDU candidate for chancellor at the next election until after Germany’s EU presidency concludes in December. So Merkel will keep her job until 2021, and the new leader will have to learn to live with her.The three leading candidates are Armin Laschet, Friedrich Merz and Norbert Röttgen, all from the state of North Rhine-Westphalia. Two of the three – Merz and Röttgen – were sacked by Merkel from their former jobs. They have not forgotten. Only Armin Laschet, currently CDU leader in North Rhine-Westphalia and state premier, can be described as a Merkel loyalist, true to her centrist mantra.He is the man to beat, having teamed up with Jens Spahn, the 39-year-old health minister, who is popular with party conservatives. Spahn will run as his deputy, so the team straddles the left-right divide in the party. But the contest still seems set to be a bitter battle between pro- and anti-Merkel factions that could leave the party badly split.After nearly 15 years as chancellor, and 18 years as CDU leader, Merkel remains the most popular politician in Germany. In spite of criticism that she lacks vision, her caution and predictability appear to be just what most German voters like.But her term in office has also seen the steady shrinking of the centre ground in German politics, with the rise of the environmentalist Green party and the far-right Alternative for Germany (AfD) at the expense of the centre-right CDU and the centre-left Social Democratic Party (SPD).The battle for the soul of the CDU is between those who think Merkel has been too left-wing, and want a more conservative leader to win back AfD voters, and those who believe that the CDU must stay in the centre, and prepare for a future coalition with the Greens. Merz is seen as the former, Laschet and Röttgen the latter.Unless Laschet emerges as the clear winner in April, the leadership contest is likely to leave Germany sorely distracted by domestic politics just as it takes over the EU presidency in the second half of the year. Instead of Merkel having a triumphant international swansong on the EU stage, she could be battling to protect her inheritance at home.The one area on which all three leadership candidates seem to agree is foreign policy: they all want Germany to take more leadership and responsibility, and for the European Union to play a bigger role in security, defence and international affairs. They are all Atlanticists, but critical of Donald Trump’s ‘America First’ stance. All are on the record criticizing the chancellor – at least tacitly – for not having a more vigorous foreign policy.There the similarity ends.On the right, the 64-year-old Merz is both the most conservative and the most popular with the party grassroots. He fell out with the chancellor when she took over his job as CDU leader in parliament in 2002. He quit politics to become a corporate lawyer (and a millionaire), but never lost his political ambition. He is an economic liberal but socially conservative, a strong critic of Merkel’s migration policy and her lack of clear leadership. Critics say he is a man of the past, and not a team player.On the EU, he believes Germany is ‘leaving too much to the French’. If France and Germany cannot agree on financial matters, he said at the London School of Economics in February, they should instead forge a stronger EU industrial policy focused on creating more ‘European champions’.Laschet, the Merkel loyalist, is four years younger, and from the left of the party. Like Merz, he is a former member of the European parliament. In 2015, he defended Merkel’s open border policy to accept refugees stranded in the Balkans. On Russia, however, he is more critical, calling for a new effort to re-engage with Vladimir Putin. Most recently, at the Munich Security Conference, he called for stronger Franco-German relations, and more support for the eurozone reforms proposed by Emmanuel Macron.As CDU leader in North Rhine-Westphalia, Laschet has the strongest power base. He earned his political spurs there by winning the last state election in 2017, in contrast to Röttgen, who lost to the SPD and Greens five years earlier.Röttgen, chairman of the Bundestag foreign affairs committee, is the surprise candidate. Once a Merkel favourite, they fell out when she sacked him as environment minister after he lost the North Rhine-Westphalia election. By throwing his hat in the ring, he has forced it to become an open contest. He is independent-minded and outspoken, but not as bitterly hostile to the chancellor as Merz, so he could be a compromise candidate.Laschet is clearly the man Merkel would find it easiest to live with. The decision will be taken by a party congress, not a grassroots ballot, which gives him a better chance. But Merz is the most eloquent orator and seen as the best campaigner. The challenge for party members is whether they believe it is better to swing right and squeeze the AfD, or stick to the centre to hold onto voters tempted by the Greens, who have replaced the SPD as the second-most popular party in Germany.The race is wide open. So is the future of the CDU. The only prediction one can make with much certainty is that as long as Merkel remains chancellor, any successor will struggle to get out of her shadow. Full Article
ot Evolution, expression, and substrate specificities of aldehyde oxidase enzymes in eukaryotes [Enzymology] By feedproxy.google.com Published On :: 2020-04-17T00:06:05-07:00 Aldehyde oxidases (AOXs) are a small group of enzymes belonging to the larger family of molybdo-flavoenzymes, along with the well-characterized xanthine oxidoreductase. The two major types of reactions that are catalyzed by AOXs are the hydroxylation of heterocycles and the oxidation of aldehydes to their corresponding carboxylic acids. Different animal species have different complements of AOX genes. The two extremes are represented in humans and rodents; whereas the human genome contains a single active gene (AOX1), those of rodents, such as mice, are endowed with four genes (Aox1-4), clustering on the same chromosome, each encoding a functionally distinct AOX enzyme. It still remains enigmatic why some species have numerous AOX enzymes, whereas others harbor only one functional enzyme. At present, little is known about the physiological relevance of AOX enzymes in humans and their additional forms in other mammals. These enzymes are expressed in the liver and play an important role in the metabolisms of drugs and other xenobiotics. In this review, we discuss the expression, tissue-specific roles, and substrate specificities of the different mammalian AOX enzymes and highlight insights into their physiological roles. Full Article
ot The heme-regulatory motifs of heme oxygenase-2 contribute to the transfer of heme to the catalytic site for degradation [Protein Structure and Folding] By feedproxy.google.com Published On :: 2020-04-17T00:06:05-07:00 Heme-regulatory motifs (HRMs) are present in many proteins that are involved in diverse biological functions. The C-terminal tail region of human heme oxygenase-2 (HO2) contains two HRMs whose cysteine residues form a disulfide bond; when reduced, these cysteines are available to bind Fe3+-heme. Heme binding to the HRMs occurs independently of the HO2 catalytic active site in the core of the protein, where heme binds with high affinity and is degraded to biliverdin. Here, we describe the reversible, protein-mediated transfer of heme between the HRMs and the HO2 core. Using hydrogen-deuterium exchange (HDX)-MS to monitor the dynamics of HO2 with and without Fe3+-heme bound to the HRMs and to the core, we detected conformational changes in the catalytic core only in one state of the catalytic cycle—when Fe3+-heme is bound to the HRMs and the core is in the apo state. These conformational changes were consistent with transfer of heme between binding sites. Indeed, we observed that HRM-bound Fe3+-heme is transferred to the apo-core either upon independent expression of the core and of a construct spanning the HRM-containing tail or after a single turnover of heme at the core. Moreover, we observed transfer of heme from the core to the HRMs and equilibration of heme between the core and HRMs. We therefore propose an Fe3+-heme transfer model in which HRM-bound heme is readily transferred to the catalytic site for degradation to facilitate turnover but can also equilibrate between the sites to maintain heme homeostasis. Full Article
ot An enzyme-based protocol for cell-free synthesis of nature-identical capsular oligosaccharides from Actinobacillus pleuropneumoniae serotype 1 [Enzymology] By feedproxy.google.com Published On :: 2020-04-24T06:08:45-07:00 Actinobacillus pleuropneumoniae (App) is the etiological agent of acute porcine pneumonia and responsible for severe economic losses worldwide. The capsule polymer of App serotype 1 (App1) consists of [4)-GlcNAc-β(1,6)-Gal-α-1-(PO4-] repeating units that are O-acetylated at O-6 of the GlcNAc. It is a major virulence factor and was used in previous studies in the successful generation of an experimental glycoconjugate vaccine. However, the application of glycoconjugate vaccines in the animal health sector is limited, presumably because of the high costs associated with harvesting the polymer from pathogen culture. Consequently, here we exploited the capsule polymerase Cps1B of App1 as an in vitro synthesis tool and an alternative for capsule polymer provision. Cps1B consists of two catalytic domains, as well as a domain rich in tetratricopeptide repeats (TPRs). We compared the elongation mechanism of Cps1B with that of a ΔTPR truncation (Cps1B-ΔTPR). Interestingly, the product profiles displayed by Cps1B suggested processive elongation of the nascent polymer, whereas Cps1B-ΔTPR appeared to work in a more distributive manner. The dispersity of the synthesized products could be reduced by generating single-action transferases and immobilizing them on individual columns, separating the two catalytic activities. Furthermore, we identified the O-acetyltransferase Cps1D of App1 and used it to modify the polymers produced by Cps1B. Two-dimensional NMR analyses of the products revealed O-acetylation levels identical to those of polymer harvested from App1 culture supernatants. In conclusion, we have established a protocol for the pathogen-free in vitro synthesis of tailored, nature-identical App1 capsule polymers. Full Article
ot Detailed analyses of the crucial functions of Zn transporter proteins in alkaline phosphatase activation [Enzymology] By feedproxy.google.com Published On :: 2020-04-24T06:08:45-07:00 Numerous zinc ectoenzymes are metalated by zinc and activated in the compartments of the early secretory pathway before reaching their destination. Zn transporter (ZNT) proteins located in these compartments are essential for ectoenzyme activation. We have previously reported that ZNT proteins, specifically ZNT5–ZNT6 heterodimers and ZNT7 homodimers, play critical roles in the activation of zinc ectoenzymes, such as alkaline phosphatases (ALPs), by mobilizing cytosolic zinc into these compartments. However, this process remains incompletely understood. Here, using genetically-engineered chicken DT40 cells, we first determined that Zrt/Irt-like protein (ZIP) transporters that are localized to the compartments of the early secretory pathway play only a minor role in the ALP activation process. These transporters included ZIP7, ZIP9, and ZIP13, performing pivotal functions in maintaining cellular homeostasis by effluxing zinc out of the compartments. Next, using purified ALP proteins, we showed that zinc metalation on ALP produced in DT40 cells lacking ZNT5–ZNT6 heterodimers and ZNT7 homodimers is impaired. Finally, by genetically disrupting both ZNT5 and ZNT7 in human HAP1 cells, we directly demonstrated that the tissue-nonspecific ALP-activating functions of both ZNT complexes are conserved in human cells. Furthermore, using mutant HAP1 cells, we uncovered a previously-unrecognized and unique spatial regulation of ZNT5–ZNT6 heterodimer formation, wherein ZNT5 recruits ZNT6 to the Golgi apparatus to form the heterodimeric complex. These findings fill in major gaps in our understanding of the molecular mechanisms underlying zinc ectoenzyme activation in the compartments of the early secretory pathway. Full Article
ot Reduction of protein phosphatase 2A (PP2A) complexity reveals cellular functions and dephosphorylation motifs of the PP2A/B'{delta} holoenzyme [Enzymology] By feedproxy.google.com Published On :: 2020-04-24T06:08:45-07:00 Protein phosphatase 2A (PP2A) is a large enzyme family responsible for most cellular Ser/Thr dephosphorylation events. PP2A substrate specificity, localization, and regulation by second messengers rely on more than a dozen regulatory subunits (including B/R2, B'/R5, and B″/R3), which form the PP2A heterotrimeric holoenzyme by associating with a dimer comprising scaffolding (A) and catalytic (C) subunits. Because of partial redundancy and high endogenous expression of PP2A holoenzymes, traditional approaches of overexpressing, knocking down, or knocking out PP2A regulatory subunits have yielded only limited insights into their biological roles and substrates. To this end, here we sought to reduce the complexity of cellular PP2A holoenzymes. We used tetracycline-inducible expression of pairs of scaffolding and regulatory subunits with complementary charge-reversal substitutions in their interaction interfaces. For each of the three regulatory subunit families, we engineered A/B charge–swap variants that could bind to one another, but not to endogenous A and B subunits. Because endogenous Aα was targeted by a co-induced shRNA, endogenous B subunits were rapidly degraded, resulting in expression of predominantly a single PP2A heterotrimer composed of the A/B charge–swap pair and the endogenous catalytic subunit. Using B'δ/PPP2R5D, we show that PP2A complexity reduction, but not PP2A overexpression, reveals a role of this holoenzyme in suppression of extracellular signal–regulated kinase signaling and protein kinase A substrate dephosphorylation. When combined with global phosphoproteomics, the PP2A/B'δ reduction approach identified consensus dephosphorylation motifs in its substrates and suggested that residues surrounding the phosphorylation site play roles in PP2A substrate specificity. Full Article
ot Specificity and affinity of the N-terminal residues in staphylocoagulase in binding to prothrombin [Computational Biology] By feedproxy.google.com Published On :: 2020-04-24T06:08:45-07:00 In Staphylococcus aureus–caused endocarditis, the pathogen secretes staphylocoagulase (SC), thereby activating human prothrombin (ProT) and evading immune clearance. A previous structural comparison of the SC(1–325) fragment bound to thrombin and its inactive precursor prethrombin 2 has indicated that SC activates ProT by inserting its N-terminal dipeptide Ile1-Val2 into the ProT Ile16 pocket, forming a salt bridge with ProT's Asp194, thereby stabilizing the active conformation. We hypothesized that these N-terminal SC residues modulate ProT binding and activation. Here, we generated labeled SC(1–246) as a probe for competitively defining the affinities of N-terminal SC(1–246) variants preselected by modeling. Using ProT(R155Q,R271Q,R284Q) (ProTQQQ), a variant refractory to prothrombinase- or thrombin-mediated cleavage, we observed variant affinities between ∼1 and 650 nm and activation potencies ranging from 1.8-fold that of WT SC(1–246) to complete loss of function. Substrate binding to ProTQQQ caused allosteric tightening of the affinity of most SC(1–246) variants, consistent with zymogen activation through occupation of the specificity pocket. Conservative changes at positions 1 and 2 were well-tolerated, with Val1-Val2, Ile1-Ala2, and Leu1-Val2 variants exhibiting ProTQQQ affinity and activation potency comparable with WT SC(1–246). Weaker binding variants typically had reduced activation rates, although at near-saturating ProTQQQ levels, several variants exhibited limiting rates similar to or higher than that of WT SC(1–246). The Ile16 pocket in ProTQQQ appears to favor nonpolar, nonaromatic residues at SC positions 1 and 2. Our results suggest that SC variants other than WT Ile1-Val2-Thr3 might emerge with similar ProT-activating efficiency. Full Article
ot Impact of 1,N6-ethenoadenosine, a damaged ribonucleotide in DNA, on translesion synthesis and repair [Enzymology] By feedproxy.google.com Published On :: 2020-05-01T00:06:09-07:00 Incorporation of ribonucleotides into DNA can severely diminish genome integrity. However, how ribonucleotides instigate DNA damage is poorly understood. In DNA, they can promote replication stress and genomic instability and have been implicated in several diseases. We report here the impact of the ribonucleotide rATP and of its naturally occurring damaged analog 1,N6-ethenoadenosine (1,N6-ϵrA) on translesion synthesis (TLS), mediated by human DNA polymerase η (hpol η), and on RNase H2–mediated incision. Mass spectral analysis revealed that 1,N6-ϵrA in DNA generates extensive frameshifts during TLS, which can lead to genomic instability. Moreover, steady-state kinetic analysis of the TLS process indicated that deoxypurines (i.e. dATP and dGTP) are inserted predominantly opposite 1,N6-ϵrA. We also show that hpol η acts as a reverse transcriptase in the presence of damaged ribonucleotide 1,N6-ϵrA but has poor RNA primer extension activities. Steady-state kinetic analysis of reverse transcription and RNA primer extension showed that hpol η favors the addition of dATP and dGTP opposite 1,N6-ϵrA. We also found that RNase H2 recognizes 1,N6-ϵrA but has limited incision activity across from this lesion, which can lead to the persistence of this detrimental DNA adduct. We conclude that the damaged and unrepaired ribonucleotide 1,N6-ϵrA in DNA exhibits mutagenic potential and can also alter the reading frame in an mRNA transcript because 1,N6-ϵrA is incompletely incised by RNase H2. Full Article
ot Spectral and photochemical diversity of tandem cysteine cyanobacterial phytochromes [Plant Biology] By feedproxy.google.com Published On :: 2020-05-08T03:41:14-07:00 The atypical trichromatic cyanobacterial phytochrome NpTP1 from Nostoc punctiforme ATCC 29133 is a linear tetrapyrrole (bilin)-binding photoreceptor protein that possesses tandem-cysteine residues responsible for shifting its light-sensing maximum to the violet spectral region. Using bioinformatics and phylogenetic analyses, here we established that tandem-cysteine cyanobacterial phytochromes (TCCPs) compose a well-supported monophyletic phytochrome lineage distinct from prototypical red/far-red cyanobacterial phytochromes. To investigate the light-sensing diversity of this family, we compared the spectroscopic properties of NpTP1 (here renamed NpTCCP) with those of three phylogenetically diverged TCCPs identified in the draft genomes of Tolypothrix sp. PCC7910, Scytonema sp. PCC10023, and Gloeocapsa sp. PCC7513. Recombinant photosensory core modules of ToTCCP, ScTCCP, and GlTCCP exhibited violet-blue–absorbing dark-states consistent with dual thioether-linked phycocyanobilin (PCB) chromophores. Photoexcitation generated singly-linked photoproduct mixtures with variable ratios of yellow-orange and red-absorbing species. The photoproduct ratio was strongly influenced by pH and by mutagenesis of TCCP- and phytochrome-specific signature residues. Our experiments support the conclusion that both photoproduct species possess protonated 15E bilin chromophores, but differ in the ionization state of the noncanonical “second” cysteine sulfhydryl group. We found that the ionization state of this and other residues influences subsequent conformational change and downstream signal transmission. We also show that tandem-cysteine phytochromes present in eukaryotes possess similar amino acid substitutions within their chromophore-binding pocket, which tune their spectral properties in an analogous fashion. Taken together, our findings provide a roadmap for tailoring the wavelength specificity of plant phytochromes to optimize plant performance in diverse natural and artificial light environments. Full Article
ot Quantification of the affinities of CRISPR-Cas9 nucleases for cognate protospacer adȷacent motif (PAM) sequences [Molecular Biophysics] By feedproxy.google.com Published On :: 2020-05-08T03:41:14-07:00 The CRISPR/Cas9 nucleases have been widely applied for genome editing in various organisms. Cas9 nucleases complexed with a guide RNA (Cas9–gRNA) find their targets by scanning and interrogating the genomic DNA for sequences complementary to the gRNA. Recognition of the DNA target sequence requires a short protospacer adjacent motif (PAM) located outside this sequence. Given that the efficiency of target location may depend on the strength of interactions that promote target recognition, here we sought to compare affinities of different Cas9 nucleases for their cognate PAM sequences. To this end, we measured affinities of Cas9 nucleases from Streptococcus pyogenes, Staphylococcus aureus, and Francisella novicida complexed with guide RNAs (gRNAs) (SpCas9–gRNA, SaCas9–gRNA, and FnCas9–gRNA, respectively) and of three engineered SpCas9–gRNA variants with altered PAM specificities for short, PAM-containing DNA probes. We used a “beacon” assay that measures the relative affinities of DNA probes by determining their ability to competitively affect the rate of Cas9–gRNA binding to fluorescently labeled target DNA derivatives called “Cas9 beacons.” We observed significant differences in the affinities for cognate PAM sequences among the studied Cas9 enzymes. The relative affinities of SpCas9–gRNA and its engineered variants for canonical and suboptimal PAMs correlated with previous findings on the efficiency of these PAM sequences in genome editing. These findings suggest that high affinity of a Cas9 nuclease for its cognate PAM promotes higher genome-editing efficiency. Full Article
ot Roles of active-site residues in catalysis, substrate binding, cooperativity, and the reaction mechanism of the quinoprotein glycine oxidase [Enzymology] By feedproxy.google.com Published On :: 2020-05-08T03:41:14-07:00 The quinoprotein glycine oxidase from the marine bacterium Pseudoalteromonas luteoviolacea (PlGoxA) uses a protein-derived cysteine tryptophylquinone (CTQ) cofactor to catalyze conversion of glycine to glyoxylate and ammonia. This homotetrameric enzyme exhibits strong cooperativity toward glycine binding. It is a good model for studying enzyme kinetics and cooperativity, specifically for being able to separate those aspects of protein function through directed mutagenesis. Variant proteins were generated with mutations in four active-site residues, Phe-316, His-583, Tyr-766, and His-767. Structures for glycine-soaked crystals were obtained for each. Different mutations had differential effects on kcat and K0.5 for catalysis, K0.5 for substrate binding, and the Hill coefficients describing the steady-state kinetics or substrate binding. Phe-316 and Tyr-766 variants retained catalytic activity, albeit with altered kinetics and cooperativity. Substitutions of His-583 revealed that it is essential for glycine binding, and the structure of H583C PlGoxA had no active-site glycine present in glycine-soaked crystals. The structure of H767A PlGoxA revealed a previously undetected reaction intermediate, a carbinolamine product-reduced CTQ adduct, and exhibited only negligible activity. The results of these experiments, as well as those with the native enzyme and previous variants, enabled construction of a detailed mechanism for the reductive half-reaction of glycine oxidation. This proposed mechanism includes three discrete reaction intermediates that are covalently bound to CTQ during the reaction, two of which have now been structurally characterized by X-ray crystallography. Full Article
ot 20 Years Promoting Women in Peace and Security By feedproxy.google.com Published On :: Thu, 05 Mar 2020 15:46:54 +0000 6 March 2020 Gitika Bhardwaj Editor, Communications & Publishing, Chatham House @GitikaBhardwaj LinkedIn Dr Joan Johnson-Freese Professor and Charles F. Bolden Jr. Chair in Science, Space & Technology, US Naval War College In a series exploring women in international affairs, Dr Joan Johnson-Freese speaks to Gitika Bhardwaj about the women in peace and security agenda, 20 years since its adoption, and how far women’s inclusion in space security is being considered, 50 years since women helped men take their first steps on the moon. GettyImages-73154695 (Cropped).jpg The UN's first all-female peacekeeping force of more than 100 Indian women stand in Monrovia, Liberia. Photo: Getty Images. This year marks the 20th anniversary of the adoption of UN Security Council Resolution 1325 on women, peace and security – the first to recognize the important role of women in peacebuilding. How did the resolution come into being and how significant was its adoption in 2000? Well it has quite an amazing history that goes back to other UN resolutions, such as the International Covenant on Civil and Political Rights and the International Covenant on Economic and Social Rights, which were passed in the 1960s and came into force in the 1970s. These were some of the biggest covenants on human and civil rights at the time but it was only later that people realised, that those who passed them, did not assume that they applied to women.There was an attempt, subsequently, to pass the UN Convention on the Elimination of All Forms of Discrimination Against Women – commonly called the CEDAW Convention – which was widely adopted in 1979, when 187 out of 194 UN members signed, although the United States was not one of them, and in fact, the US has still not ratified the treaty. But the understanding that women’s rights were not necessarily assumed in human and civil rights action was beginning to gain recognition.Then, in 1995, Hillary Clinton spoke in Beijing and really put it forward that women’s rights were human rights and also civil rights and we all have to address them as such. So, ‘95 really brought together all the different social groups – women’s groups, human’s rights groups, civil rights groups, and more, who pushed for the women, peace and security agenda to be passed in 2000 at the United Nations. The resolution on women, peace and security was a significant moment because it recognized gender equality issues were national security issues – not just social justice issues – and was soon followed by a number of other resolutions which make up the women, peace and security agenda today. Conflict has a disproportionate effect on women and girls, with global security threats, such as climate change, reported to impact women more than men. In light of this, growing numbers of women are now serving on the frontlines of conflicts, in comparison to 1957-1989, when only 20 women served as UN peacekeepers. In your view, what have been the successes of the women, peace and security agenda so far? I think some of the successes specific to peacekeeping have been, as you mentioned, that women are increasingly part of peacekeeping forces being deployed to conflict and post-conflict situations.Importantly, the nature of war is changing – we are no longer primarily engaged in interstate work in some places – it’s mostly intrastate work where there are often ethnic or religious overtones.In this landscape, women are often caught up in the battle lines. They often become the heads of their households when the men are gone or injured or killed. There have also been instances of rape being used as a weapon of war and other forms of sexual violence being committed in conflict and post-conflict situations even by peacekeepers.So, having more women as peacekeepers is important because, number one, when women see women peacekeepers, they are much less likely to fear them, and therefore, feel less threatened speaking to them. Number two, women are a less threatening presence so civil society begins to build again. Number three, women peacekeepers give women in the local area a role model of strength showing them that they can play an active role in their own security. Finally, I would say that women peacekeepers are all impressively trained to guard those under their protection.What other successes have there been more broadly outside of peacekeeping? Well, I think one that is often cited is that there’s a 20 per cent increase in the probability of a peace agreement lasting at least two years and a 35 per cent increase in the probability of that peace agreement lasting 15 years if women are at the negotiating table. The reason being that women bring things to the table – for example focussing on the root causes of conflict – that men neglect either because they’re not aware of them or it’s not considered an issue of importance to them. We have a pretty abysmal history of peace agreements holding so including more women in peace negotiations, given these increases in the probability of agreements holding, seems to me the only logical thing to do.You mentioned the inclusion of women in peace processes increases the likelihood of agreements succeeding, however, women continue to be underrepresented, comprising under 10 per cent of peace negotiators and under 4 per cent of signatories to peace agreements. Do you think there are any shortcomings with focusing on increasing the presence of women over the positions they hold and how their positions are used to further gender equality? I think there are a couple of aspects to this. Importantly, women have been extremely effective in leadership positions, for example, in Liberia. But it is true having women at the table does not necessarily further gender equality in the long-term and I think this was the case in Northern Ireland where the women who were at the table did not include provisions for women. But, I think, we’ve learned since then.The most important case to cite right now, in my view, is that of women in Afghanistan. They came out of their homes, they went to school, they identified themselves as proponents of gender equality, yet now, with the US-Taliban deal, there were almost no women at the table and not a single provision in the peace agreement that deals with women. So what’s going to happen to all of these women?Since 2000, the number of agreements referring to women has grown to 28 per cent – more than double the number between 1990-2000. However, some critics have pointed to the gap between theory and practice since many peace agreements still omit a gender perspective on peacekeeping operations. What, in your opinion, have been the failings of realising the women, peace and security agenda so far? You know, it’s not just critics who point this out, it’s advocates as well, that there is a big gap between rhetoric and implementation. I think the reason for this, in most cases, is political will. It’s the idea that gender equality is an optional luxury – we’ll get to it when we can – and that we have more important issues to work out. Well the agency of 50 per cent of the population should not be seen as an optional luxury.In all cases, however, it’s a question of power. There are only so many seats at the table where power is doled out and nobody wants to give up theirs to let somebody else sit down. So, I think, there is active resistance to implementing gender equality in the peace and security arena because it would mean sharing power. There are also some other reasons. The first is something we call the ‘blind fish’ – people who are simply unaware of gender equality issues – the second, which is interestingly by adamant supporters of the women in peace and security agenda, and that’s they are not given the budget and the authority to carry out the agenda as it should be. A female Italian soldier from the United Nations Interim Force in Lebanon (UNIFIL) talks with a Lebanese woman in Tibnine, south Lebanon. Photo: Getty Images. The concept of a feminist foreign policy, which places gender equality and women’s rights at the centre of its foreign policy goals, has been gaining momentum globally. Sweden became the first country in the world to adopt a feminist foreign policy in 2015, and since then, 82 countries have adopted national action plans to raise the role of women in peace processes. How effective have these measures been to furthering women in peace and security? That, I think, is one of the issues that women in peace and security advocates, such as myself, are currently looking at because the word ‘feminist’, for better or for worse, is a trigger in many countries.In many countries, it’s a negative trigger, which is kind of ironic because, if you ask people, ‘Do you believe in gender equality?’, a lot of people will say, ‘Yes absolutely.’ But if you ask people, ‘Are you a feminist?’, a lot of people will say, ‘No not me.’ So, the question is, do we aim for a broad goal like a feminist foreign policy which would look at defining peace as, not just the absence of war, but a lot more then than that, such as creating the conditions needed for gender equality and aiming for peace and stability among other broader goals.Or do we aim to work on a more incremental basis by trying to get more women into peacekeeping, trying to get more women into leadership positions, trying to move gender equality up the agenda as the more effective path forward?I think the answer to the question is that it depends. If you’re Sweden, Canada or Mexico, a feminist foreign policy might be acceptable. But, if you’re the United States, it’s nowhere near acceptable. Even getting the US Defence Department to take the incremental steps of the women, peace and security agenda has been challenging.Why do you think that is the case in the United States?I think a lot of it has to do with power as I mentioned earlier. It has to do with an assumption that women aren’t assertive and don’t see security under the same lens as men, which is true, but which is why they are needed in this space.I think it also, again, goes back to the point that some see it as an optional luxury rather than an absolute necessity and everybody is too busy – or simply unwilling – to change the status quo.You mentioned the cases of Liberia and Northern Ireland, but another example that struck me was Rwanda, where women make up 62 per cent of the national legislature, far more proportionally than any other country, following provisions included in its constitution in the aftermath of the genocide of 1994.Though equal representation between men and women is still far off for most of the world, what does the case of Rwanda and other post-conflict countries demonstrate about how to go about including more women in peacebuilding?Well, I think the number one way to get more women into political leadership roles, where the women, peace and security agenda could then be implemented, is quotas. Many countries use quotas to increase more women in political participation, which Rwanda certainly does, although there are different types. There are quotas that say each political party must have X number of women as candidates and then there are quotas that say the overall number of women in the parliament must reach a certain level. So there are different varieties of quotas but they are all used as, kind of, affirmative action methods to at least temporarily bring the numbers up to where women’s voices are inclusive not token. Research has shown that until you have at least 33 per cent of a minority in an overall group – so if it’s all men then 33 per cent of this group as women – then you won’t see any change because having one or two will likely be drowned out. But, at about 33 per cent, they’re able to have political power which then means their views and their agendas are seriously considered. So, in Rwanda, that has certainly been the case and that’s been one of the big lessons learned. As I mentioned earlier, in Northern Ireland, the lesson learned was that it’s not enough to just have women in on peace agreements. There need to be implementation assurances written into the peace agreement that says it must be taken forward. So, in this vein, women have been learning over the years how to make a difference in male-dominated spaces. 50 years ago, humans landed on the moon, becoming one of the most significant moments in human history. The stories of women, from Margaret Hamilton, to Katherine Johnson, to JoAnn Morgan, who all helped men take their first steps on the moon at the height of the space race, have since come into the spotlight. How will the inclusion of women need to be considered more in space security as it becomes increasingly important in international relations?Right now, space security is at a very critical point. We have moved from a situation where there was both co-operation and competition during the space race to a situation of great power competition in space where the United States, China and Russia are, for the first time, overtly weaponizing space. When I say overtly, much of space technology is dual-use, meaning it could be used as a weapon or it could be used as something for non-military purposes.In the past, the United States and other countries have been very careful, kind of, not to cross the Rubicon into the overt weaponization of space but that’s now ending which I think puts us in a very precarious situation. What seems to be missing from considerations of space security at the moment is the most threatening issue – space debris – which can only be dealt with on a multinational basis meaning it inherently requires co-operation. So, what I think more women in space security positions would bring, would be that inclusion and the insistence on inclusion as a pillar of space diplomacy.If there was just a fraction of the money, and manpower, spent on space diplomacy as there is on planning for space warfare, I think we’d all be a lot better off. NASA engineer, JoAnn Morgan, watches the blast off of Apollo 11 at the Kennedy Space Center on 16 July 1969. She was at the time NASA's only female engineer. Photo: NASA via Getty Images. How far do current discussions about women in peace and security factor in space security?Not at all.Do you see the role of women in space security progressing in the future in spite of this and also despite what some have described as a broader backlash against women around the world whether in the political or in the security space?I think there have been events over the past five or so years that have made women around the world, if anything, more acutely insistent on their participation than ever before.We saw the marches in 2017, in the United States and worldwide, in response to what women felt was a rise in authoritarian and misogynistic governments and we have seen the rollbacks in gender equality rights in areas like reproduction too but I don’t think they’re going to take it lying down. The backlash, if anything, is going to spur women to be more, not less, active in all spaces. Some have argued for the need of a men, peace and security agenda, to compliment the work on women, with proponents arguing that men are needed to realise gender equality worldwide. How far are men needed as allies to realise the women, peace and security agenda? Well, I think, though the women, peace and security agenda has women in the title, it argues for gendered perspectives, that policies affect men and women differently. So, I think it is very important that it not be seen as dealing only with women’s issues – it deals with gendered perspectives. In that regard, it is very much needed to have a broadening of all of those involved. I mentioned earlier it was all women’s groups that got the women, peace and security agenda passed and now we need to include men. In fact, I would point out, NATO is a great example of an organization that has recognized the importance of looking at how policies affect men and women, girls and boys. So bringing more men in to support gendered perspectives is absolutely essential and looking at gendered perspectives in things like leadership roles is critical as well as gendered perspectives in everything from space policy to nuclear policy to human security issues too.In your view, what are the greatest challenges to the uptake of gendered perspectives across the board and what, if anything, needs to change in order to realise the goals set out by the UNSC 20 years ago?On a macro level, we need accountability. We have lots of policies, laws, national action plans and strategies of all kinds but we need accountability.In the United States, in particular, I very much hope that accountability comes from Congress. In 2017, Congress passed the Women, Peace and Security Act on a bipartisan basis but I think it’s now up to Congress to hold organizations responsible for its implementation. On an organizational level, we need to get, as you said, more men involved. But, interestingly, not all women agree, so we need to have more talks among women too, be they liberal, conservative, working, non-working, mothers, not mothers etc. We may have different views but where we’re trying to go is the same and we need to work together better. I think among the advocates of women, peace and security, there are still issues that are up for debate like do we go for big feminist foreign policies or do we go for incremental change? In addition, are there lynchpin issues such as reproductive rights, women’s healthcare, gender equality or budget? You know, in the United States, I wrote in an article that although the US Defence Department gave $4 million for women, peace and security in 2018, which they were patting themselves on the back for, the Military Times pointed out that they are spending $84 million a year on Viagra.But this is not just in the United States. 140 countries stood up at the UN to advocate for women, peace and security in 2000 but only 25 per cent of those have national action plans and any budget connected to those plans. Everyone everywhere needs to put their money where their mouths are if we are to realise the goals set out by the UNSC 20 years ago. Full Article
ot Legal Provision for Crisis Preparedness: Foresight not Hindsight By feedproxy.google.com Published On :: Tue, 21 Apr 2020 17:03:31 +0000 21 April 2020 Dr Patricia Lewis Research Director, Conflict, Science & Transformation; Director, International Security Programme @PatriciaMary COVID-19 is proving to be a grave threat to humanity. But this is not a one-off, there will be future crises, and we can be better prepared to mitigate them. 2020-04-21-Nurse-COVID-Test Examining a patient while testing for COVID-19 at the Velocity Urgent Care in Woodbridge, Virginia. Photo by Chip Somodevilla/Getty Images. A controversial debate during COVID-19 is the state of readiness within governments and health systems for a pandemic, with lines of the debate drawn on the issues of testing provision, personal protective equipment (PPE), and the speed of decision-making.President Macron in a speech to the nation admitted French medical workers did not have enough PPE and that mistakes had been made: ‘Were we prepared for this crisis? We have to say that no, we weren’t, but we have to admit our errors … and we will learn from this’.In reality few governments were fully prepared. In years to come, all will ask: ‘how could we have been better prepared, what did we do wrong, and what can we learn?’. But after every crisis, governments ask these same questions.Most countries have put in place national risk assessments and established processes and systems to monitor and stress-test crisis-preparedness. So why have some countries been seemingly better prepared?Comparing different approachesSome have had more time and been able to watch the spread of the disease and learn from those countries that had it first. Others have taken their own routes, and there will be much to learn from comparing these different approaches in the longer run.Governments in Asia have been strongly influenced by the experience of the SARS epidemic in 2002-3 and - South Korea in particular - the MERS-CoV outbreak in 2015 which was the largest outside the Middle East. Several carried out preparatory work in terms of risk assessment, preparedness measures and resilience planning for a wide range of threats.Case Study of Preparedness: South KoreaBy 2007, South Korea had established the Division of Public Health Crisis Response in Korea Centers for Disease Control and Prevention (KCDC) and, in 2016, the KCDC Center for Public Health Emergency Preparedness and Response had established a round-the-clock Emergency Operations Center with rapid response teams.KCDC is responsible for the distribution of antiviral stockpiles to 16 cities and provinces that are required by law to hold and manage antiviral stockpiles.And, at the international level, there are frameworks for preparedness for pandemics. The International Health Regulations (IHR) - adopted at the 2005 World Health Assembly and binding on member states - require countries to report certain disease outbreaks and public health events to the World Health Organization (WHO) and ‘prevent, protect against, control and provide a public health response to the international spread of disease in ways that are commensurate with and restricted to public health risks, and which avoid unnecessary interference with international traffic and trade’.Under IHR, governments committed to a programme of building core capacities including coordination, surveillance, response and preparedness. The UN Sendai Framework for Disaster Risk highlights disaster preparedness for effective response as one of its main purposes and has already incorporated these measures into the Sustainable Development Goals (SDGs) and other Agenda 2030 initiatives. UN Secretary-General António Guterres has said COVID-19 ‘poses a significant threat to the maintenance of international peace and security’ and that ‘a signal of unity and resolve from the Council would count for a lot at this anxious time’.Case Study of Preparedness: United StatesThe National Institutes of Health (NIH) and the Center for Disease Control (CDC) established PERRC – the Preparedness for Emergency Response Research Centers - as a requirement of the 2006 Pandemic and All-Hazards Preparedness Act, which required research to ‘improve federal, state, local, and tribal public health preparedness and response systems’.The 2006 Act has since been supplanted by the 2019 Pandemic and All-Hazards Preparedness and Advancing Innovation Act. This created the post of Assistant Secretary for Preparedness and Response (ASPR) in the Department for Health and Human Services (HHS) and authorised the development and acquisitions of medical countermeasures and a quadrennial National Health Security Strategy.The 2019 Act also set in place a number of measures including the requirement for the US government to re-evaluate several important metrics of the Public Health Emergency Preparedness cooperative agreement and the Hospital Preparedness Program, and a requirement for a report on the states of preparedness and response in US healthcare facilities.This pandemic looks set to continue to be a grave threat to humanity. But there will also be future pandemics – whether another type of coronavirus or a new influenza virus – and our species will be threatened again, we just don’t know when.Other disasters too will befall us – we already see the impacts of climate change arriving on our doorsteps characterised by increased numbers and intensity of floods, hurricanes, fires, crop failure and other manifestations of a warming, increasingly turbulent atmosphere and we will continue to suffer major volcanic eruptions, earthquakes and tsunamis. All high impact, unknown probability events.Preparedness for an unknown future is expensive and requires a great deal of effort for events that may not happen within the preparers’ lifetimes. It is hard to imagine now, but people will forget this crisis, and revert to their imagined projections of the future where crises don’t occur, and progress follows progress. But history shows us otherwise.Preparations for future crises always fall prey to financial cuts and austerity measures in lean times unless there is a mechanism to prevent that. Cost-benefit analyses will understandably tend to prioritise the urgent over the long-term. So governments should put in place legislation – or strengthen existing legislation – now to ensure their countries are as prepared as possible for whatever crisis is coming.Such a legal requirement would require governments to report back to parliament every year on the state of their national preparations detailing such measures as:The exact levels of stocks of essential materials (including medical equipment)The ability of hospitals to cope with large influx of patientsHow many drills, exercises and simulations had been organised – and their findingsWhat was being done to implement lessons learned & improve preparednessIn addition, further actions should be taken:Parliamentary committees such as the UK Joint Committee on the National Security Strategy should scrutinise the government’s readiness for the potential threats outlined in the National Risk register for Civil Emergencies in-depth on an annual basis.Parliamentarians, including ministers, with responsibility for national security and resilience should participate in drills, table-top exercises and simulations to see for themselves the problems inherent with dealing with crises.All governments should have a minister (or equivalent) with the sole responsibility for national crisis preparedness and resilience. The Minister would be empowered to liaise internationally and coordinate local responses such as local resilience groups.There should be ring-fenced budget lines in annual budgets specifically for preparedness and resilience measures, annually reported on and assessed by parliaments as part of the due diligence process.And at the international level:The UN Security Council should establish a Crisis Preparedness Committee to bolster the ability of United Nations Member States to respond to international crisis such as pandemics, within their borders and across regions. The Committee would function in a similar fashion as the Counter Terrorism Committee that was established following the 9/11 terrorist attacks in the United States.States should present reports on their level of preparedness to the UN Security Council. The Crisis Preparedness Committee could establish a group of experts who would conduct expert assessments of each member state’s risks and preparedness and facilitate technical assistance as required.Regional bodies such as the OSCE, ASEAN and ARF, the AU, the OAS, the PIF etc could also request national reports on crisis preparedness for discussion and cooperation at the regional level.COVID-19 has been referred to as the 9/11 of crisis preparedness and response. Just as that shocking terrorist attack shifted the world and created a series of measures to address terrorism, we now recognise our security frameworks need far more emphasis on being prepared and being resilient. Whatever has been done in the past, it is clear that was nowhere near enough and that has to change.Case Study of Preparedness: The UKThe National Risk Register was first published in 2008 as part of the undertakings laid out in the National Security Strategy (the UK also published the Biological Security Strategy in July 2018). Now entitled the National Risk Register for Civil Emergencies it has been updated regularly to analyse the risks of major emergencies that could affect the UK in the next five years and provide resilience advice and guidance.The latest edition - produced in 2017 when the UK had a Minister for Government Resilience and Efficiency - placed the risk of a pandemic influenza in the ‘highly likely and most severe’ category. It stood out from all the other identified risks, whereas an emerging disease (such as COVID-19) was identified as ‘highly likely but with moderate impact’.However, much preparatory work for an influenza pandemic is the same as for COVID-19, particularly in prepositioning large stocks of PPE, readiness within large hospitals, and the creation of new hospitals and facilities.One key issue is that the 2017 NHS Operating Framework for Managing the Response to Pandemic Influenza was dependent on pre-positioned ’just in case’ stockpiles of PPE. But as it became clear the PPE stocks were not adequate for the pandemic, it was reported that recommendations about the stockpile by NERVTAG (the New and Emerging Respiratory Virus Threats Advisory Group which advises the government on the threat posed by new and emerging respiratory viruses) had been subjected to an ‘economic assessment’ and decisions reversed on, for example, eye protection.The UK chief medical officer Dame Sally Davies, when speaking at the World Health Organization about Operation Cygnus – a 2016 three-day exercise on a flu pandemic in the UK – reportedly said the UK was not ready for a severe flu attack and ‘a lot of things need improving’.Aware of the significance of the situation, the UK Parliamentary Joint Committee on the National Security Strategy launched an inquiry in 2019 on ‘Biosecurity and human health: preparing for emerging infectious diseases and bioweapons’ which intended to coordinate a cross-government approach to biosecurity threats. But the inquiry had to postpone its oral hearings scheduled for late October 2019 and, because of the general election in December 2019, the committee was obliged to close the inquiry. Full Article
ot Nuclear Tensions Must Not Be Sidelined During Coronavirus By feedproxy.google.com Published On :: Fri, 01 May 2020 11:18:35 +0000 1 May 2020 Ana Alecsandru Research Assistant, International Security Programme @anaalecs LinkedIn Although the pandemic means the Nuclear Non-Proliferation Treaty (NPT) Review Conference (RevCon) is postponed, the delay could be an opportunity to better the health of the NPT regime. 2020-05-01-Iran-Peace-Nuclear Painted stairs in Tehran, Iran symbolizing hope. Photo by Fatemeh Bahrami/Anadolu Agency/Getty Images. Despite face-to-face diplomatic meetings being increasingly rare during the current disruption, COVID-19 will ultimately force a redefinition of national security and defence spending priorities, and this could provide the possibility of an improved political climate at RevCon when it happens in 2021.With US presidential elections due in November and a gradual engagement growing between the EU and Iran, there could be a new context for more cooperation between states by 2021. Two key areas of focus over the coming months will be the arms control talks between the United States and Russia, and Iran’s compliance with the 2015 Joint Comprehensive Plan of Action (JCPOA), also known as the Iran Nuclear Deal.It is too early to discern the medium- and longer-term consequences of COVID-19 for defence ministries, but a greater focus on societal resilience and reinvigorating economic productivity will likely undercut the rationale for expensive nuclear modernization.Therefore, extending the current New START (Strategic Arms Reduction Treaty) would be the best, most practical option to give both Russia and the United States time to explore more ambitious multilateral arms control measures, while allowing their current focus to remain on the pandemic and economic relief.Continuing distrustBut with the current treaty — which limits nuclear warheads, missiles, bombers, and launchers — due to expire in February 2021, the continuing distrust between the United States and Russia makes this extension hard to achieve, and a follow-on treaty even less likely.Prospects for future bilateral negotiations are hindered by President Donald Trump’s vision for a trilateral arms control initiative involving both China and Russia. But China opposes this on the grounds that its nuclear arsenal is far smaller than that of the two others.While there appears to be agreement that the nuclear arsenals of China, France, and the UK (the NPT nuclear-weapons states) and those of the states outside the treaty (India, Pakistan, North Korea, and Israel) will all have to be taken into account going forward, a practical mechanism for doing so proves elusive.If Joe Biden wins the US presidency he seems likely to pursue an extension of the New START treaty and could also prevent a withdrawal from the Open Skies treaty, the latest arms control agreement targeted by the Trump administration.Under a Biden administration, the United States would also probably re-join the JCPOA, provided Tehran returned to strict compliance with the deal. Biden could even use the team that negotiated the Iran deal to advance the goal of denuclearization of the Korean peninsula.For an NPT regime already confronted by a series of longstanding divergences, it is essential that Iran remains a signatory especially as tensions between Iran and the United States have escalated recently — due to the Qassim Suleimani assassination and the recent claim by Iran’s Revolutionary Guard Corps to have successfully placed the country’s first military satellite into orbit.This announcement raised red flags among experts about whether Iran is developing intercontinental ballistic missiles due to the dual-use nature of space technology. The satellite launch — deeply troubling for Iran’s neighbours and the EU countries — may strengthen the US argument that it is a cover for the development of ballistic missiles capable of delivering nuclear weapons.However, as with many other countries, Iran is struggling with a severe coronavirus crisis and will be pouring its scientific expertise and funds into that rather than other efforts — including the nuclear programme.Those European countries supporting the trading mechanism INSTEX (Instrument in Support of Trade Exchanges) for sending humanitarian goods into Iran could use this crisis to encourage Iran to remain in compliance with the JCPOA and its NPT obligations.France, Germany and the UK (the E3) have already successfully concluded the first transaction, which was to facilitate the export of medical goods from Europe to Iran. But the recent Iranian escalatory steps will most certainly place a strain on the preservation of this arrangement.COVID-19 might have delayed Iran’s next breach of the 2015 nuclear agreement but Tehran will inevitably seek to strengthen its hand before any potential negotiations with the United States after the presidential elections.As frosty US-Iranian relations — exacerbated by the coronavirus pandemic — prevent diplomatic negotiations, this constructive engagement between the E3 and Iran might prove instrumental in reviving the JCPOA and ensuring Iran stays committed to both nuclear non-proliferation and disarmament.While countries focus their efforts on tackling the coronavirus pandemic, it is understandable resources may be limited for other global challenges, such as the increasing risk of nuclear weapons use across several regions.But the potential ramifications of the COVID-19 crisis for the NPT regime are profound. Ongoing tensions between the nuclear-armed states must not be ignored while the world’s focus is elsewhere, and the nuclear community should continue to work together to progress nuclear non-proliferation and disarmament, building bridges of cooperation and trust that can long outlast the pandemic. Full Article
ot Demystifying the media caricatures of Pussy Riot By feedproxy.google.com Published On :: Wed, 05 Feb 2014 12:13:19 +0000 6 February 2014 , Volume 70, Number 1 Masha Gessen, Words will Break Cement: The Passion of Pussy Riot, Granta, £8.70 Sean Guillory, author of seansrussiablog.org Guillory.jpg Maria Alyokhina and Nadazhda Tolokonnikova, two members of Pussy Riot, speak with their lawyer from a glass-walled cage in a court in Moscow. Photo: AFP/Getty Images Full Article
ot Alain de Botton on how the news should aim to improve our lives By feedproxy.google.com Published On :: Wed, 05 Feb 2014 12:47:15 +0000 6 February 2014 , Volume 70, Number 1 The author of books on love, religion and Proust, explains why the news agenda should be more positive Download PDF Agnes Frimston deBotton.jpg Photo: Getty Images Is the news our society’s religion today?In the developed economies, the news now occupies a position of power at least equal to that formerly enjoyed by the faiths. Matins has been transubstantiated into the breakfast bulletin, vespers into the evening report. It also demands that we approach it with some of the same deferential expectations we would once have harboured for the faiths. Here, too, we hope to receive revelations, learn who is good and bad, fathom suffering and understand the unfolding logic of existence. And here too, if we refuse to take part in the rituals, there could be imputations of heresy.What would you like the news to be?It feels like there is always an infinite amount of news, so much is happening in the world every day. Yet after a while it becomes clear that the same kinds of event are recurring again and again. The details change, but all circle round the same archetypal story. Men in highly responsible positions are coming unstuck because their desires lead them to do things that, when made public, are shameful.Identifying the underlying theme is more important in the long run than going through the details of every case. The news that matters is not so much that this MP or banker did what they did. What we need to address is why such things happen.How do you keep the public interested in the news that does matter?We cannot be collectively dragged into being more responsible through guilt. The Arctic ice is melting and this is going to have major, lasting implications for sea levels and weather around the world. A few people care a lot but, strangely, Taylor Swift’s legs are far more captivating. The starting point has to be indulgence towards the way our minds work. We are interested in Swift’s legs not because we are evil – but because we are wired in unhelpful ways. If we are going to be interested en masse in the defrosting poles, we need to take our fragilities on board and therefore get serious about trying to make important news not just ‘important’, but also beguiling. Then things stand a chance of changing.You argue that we should set aside ‘neutral reporting’. Are you asking for more bias?Many people imagine that what makes news organizations serious is their ability to provide us with information that is ‘unbiased’. But facts can only become meaningful and relevant to us when they slot into some picture of important or trivial, right or wrong. News organizations that vaunt their neutrality forget that neutrality is simply impossible. There is no risk-free, all-knowing sober set of answers to cling to. At heart, the word ‘bias’ simply alludes to the business of having a ‘take’ on existence. One may have a better or worse take, but one can’t make any sense of the flotsam of daily events in the news without having one. All of the figures we revere in history have been highly biased: each of them had a strong sense of what mattered and why, and their judgments were anything but perfectly balanced. They were just flavoured in the right way. We don’t need news stripped of bias, we need news presented to us with the best kinds of bias.What is the purpose of foreign reporting?Foreign reporting implicitly defers to the priorities of the state and of business, occupying itself almost exclusively with whom and where we should fight, trade or sympathize. But it should instead offer us a means by which to humanize the Other who instinctively repels, bores or frightens us and with whom we can’t, without help, imagine having anything in common.Foreign countries also furnish a scale against which our own nation and ways of living can be assessed; they may help us to see our national oddities, blind spots and strengths. Stories from them may lead us to a fresh appreciation of the imperfect freedoms and comparative abundance of our homelands, which otherwise would be treated only as matters for grumbling or blame. Alternatively, problems with which we are all too familiar may be revealed to have been solved better elsewhere.You mention the importance of historical perspective in reporting, so that we can respond to issues with context.Contrary to what the news usually suggests, hardly anything is ever totally new, few things are truly amazing and very little is absolutely terrible. The economic indices are grim, but we have weathered comparable drops many times over the past century and even the worst scenarios only predict that we will return to a standard of living we had a few decades ago. A bad avian flu may disrupt international travel and defeat known drugs for a while, but research will eventually understand and contain it. The floods look dramatic, but in the end, they will affect merely a fraction of the population and recede soon.How can photography change the way we report news?There are now more images than ever before in the coverage we consume, but the problem lies in the lack of ambition behind their production and display. We might usefully divide news photographs into two genres. The first are images of corroboration, which do little other than confirm something we have learnt about a person or an event through an accompanying article. The second is a rarer kind of image, the photograph of revelation, whose ambition is not simply to back up what the text tells us but to advance our level of knowledge to a new point. It sets out to challenge cliché. We have lost any sense of photography’s potential as an information-bearing medium, as a force to properly introduce us to a planet that we keep conceitedly assuming that we know rather well already.Do you feel oppressed by the news?The pressure of not missing out makes one feel one has to care about a given topic, even when one doesn’t want to. Take Mandela’s funeral. One was supposed to care a lot, and yet, you don’t. You know the reasons why it is important, but they don’t grip you because you are focused elsewhere on subjects that, while tiny in the grand scheme of things, matter a lot within your context. It would be dangerous if hardly anyone paid attention to what the Government was doing, or what was happening to the environment. But it is not right to go from this to the demand that everyone should be interested in every item whenever the news machine calls. We badly need people whose attention is not caught up in the trends of the moment and who are not looking in the same direction as everyone else. We need people scanning the less familiar parts of the horizon.Do you think we get the news we deserve?Much of what we now take for granted as news has its origins in the information needed by those people taking major decisions or who are at the centre of national affairs. Ease of communication and a generous democratic impulse means that selections from the knowledge base, originally designed for decision-makers, now gets routinely sent via the media to very large numbers of people. It is as if a dossier which might properly arrive upon the desk of a Minister has accidentally been delivered to the wrong address and ends up on the breakfast table of an electrician in Pitlochry. Every day the news gives us stuff that is both interesting for some people and irrelevant to you. No wonder we’re sometimes a bit bored. It’s not our fault. Full Article
ot Lesotho’s Domestic Priorities and Global Interests: Small Country Levers for International Influence By feedproxy.google.com Published On :: Tue, 01 Apr 2014 14:57:42 +0000 Research Event 7 April 2014 - 3:00pm to 4:00pm Chatham House, London Meeting Summarypdf | 59.43 KB Event participants HE Dr Motsoahae Thomas Thabane, Prime Minister of LesothoChair: Ian Lucas MP, Shadow Minister for Africa and the Middle East The landlocked southern African nation of Lesotho faces a number of domestic challenges in 2014. Reducing reliance on the agricultural sector, containing the prevalence of HIV and improving service delivery are all important issues that must be addressed to ensure sustainable growth. However, social achievements including literacy rates and levels of gender parity among the highest in Africa, the country’s potential for electricity exports, and the booming relationship between its textile industry and the United States, could mean Lesotho is well-placed for future growth and development. HE Prime Minister Tom Thabane will discuss how his government seeks to address the Lesotho’s domestic issues and how the country seeks to exert its influence in the regional Southern African Development Community. Full Article
ot Scotland's Place in the World By feedproxy.google.com Published On :: Tue, 22 Jul 2014 11:15:01 +0000 Members Event 2 September 2014 - 6:00pm to 7:00pm Chatham House, London Transcriptpdf | 129.63 KB Transcript Q&Apdf | 102.4 KB Event participants Rt Hon Danny Alexander MP, Chief Secretary to the Treasury, UKChair: Faisal Islam, Political Editor, Sky News Ahead of the referendum on Scottish independence, the speaker will set out his vision for Scotland to remain as an integral and full member of the United Kingdom. He will explain that Scotland’s global influence and international reach – in terms of diplomacy, peacekeeping and aid relief – is significantly enhanced by being part of a bigger, stronger and more powerful state such as the UK.LIVE STREAM: This event will be live streamed. The live stream will be made available here at 18:00 BST on Tuesday 02 September.ASK A QUESTION: Send questions for the speaker by email to questions@chathamhouse.org or using #askCH on Twitter. A selection will be put to him during the event. Event attributes Livestream Members Events Team Email Full Article
ot Labour Cannot Be Complacent About UKIP’s Advance By feedproxy.google.com Published On :: Tue, 02 Sep 2014 08:38:31 +0000 2 September 2014 Professor Matthew Goodwin Visiting Senior Fellow, Europe Programme @GoodwinMJ LinkedIn Google Scholar Support for UKIP is growing among the groups in which Labour is struggling. 20140902Farage.jpg Nigel Farage speaks to voters in Clacton-on-Sea the day after Douglas Carswell MP announced he is switching allegiance from the Conservative party to UKIP. Photo by Oli Scarff/Getty Images. The UK Independence Party (UKIP) turns 21 years old this month and has cause to celebrate. The insurgent party won a national election in May and last week enjoyed the most significant coup in its history when Conservative MP Douglas Carswell defected to UKIP and announced a forthcoming by-election. Given that his coastal seat of Clacton is UKIP’s most demographically favourable seat in the country, Carswell has almost certainly handed the party its first elected member of parliament.The events in Clacton will be seen by many as validating one of the oldest myths about UKIP; that it is nothing more than a second home for disgruntled Conservatives. Carswell’s defection will be especially welcomed on the left, where many argue UKIP is dividing the right and clearing the path for Labour’s return to power in 2015. This is dangerously misguided.To understand why, we can start with Clacton. The seat has the largest concentration of the 'left behind' demographic; older, white, blue-collar voters who lack qualifications, felt excluded from Britain’s economic transformation long before the crisis, are cut adrift from politics, and are intensely anxious over the cultural as well as economic effects of migration.But as a Conservative-held seat, Clacton is also an outlier. For our book, Revolt on the Right, Robert Ford and I ranked all seats according to their demographic receptiveness to UKIP. Contrary to the prevailing wisdom, most are in Labour territory where MPs are battling with the same cocktail as Carswell: economic stagnation, unease over migration, an entrenched anti-politics consensus and anxieties over rapid social change. Of the 20 seats most demographically receptive to UKIP, 18 have Labour incumbents. Of the top 50, Labour holds 42. Of course, demography alone is not necessarily destiny. Aside from a receptive local population, UKIP also needs a favourable political context. Unlike Conservative-held seats that are at genuine risk from UKIP, Labour’s heartland seats are currently protected by large majorities.But a cursory glance at the recent European parliament results reveals the direction of travel. UKIP comfortably won the popular vote in a swath of Labour territory, and talks ambitiously of becoming the main rival to Labour in northern England. It appears to be succeeding. Across 39 local authorities in the northwest, UKIP won more votes than Labour in 13 and finished as its main rival in 23. It is similarly bleak in the northeast; across 12 authorities UKIP won the popular vote in five and finished second to Labour in seven.Since 2010, UKIP has grown fastest among the groups in which Labour is struggling most: the over-65s, the working-class and those who left education early. UKIP is tearing off this section of the electorate, creating a fundamental divide in British politics between those with the skills, education and resources to adapt, and those who have little and feel intensely angry. This is why some Ukippers talk of a '2020 strategy' and plot further advances under an Ed Miliband-led post-2015 government.Those who compare the party to earlier attempts to redraw the political map, such as the Social Democratic Party in the 1980s, or populist crusaders like the French Poujadists in the 1950s, miss a crucial point. UKIP is anchored in modern Britain’s most socially distinctive support base; it is the most working-class movement since Michael Foot’s Labour Party. Labelling UKIP as 'populist' implies that it appeals across society. It does not. Its strength is concentrated in the 'left behind', who cluster in specific geographical areas. Crucially, this is essential for success under first-past-the-post.Labour should be under no illusion. UKIP is attracting the Carswells of this world but it is also emerging as the main opposition in many northern heartlands, where it benefits from the toxicity of the Tories, moribund Labour machines that have not had to compete for decades, and the short-sightedness of some close to Ed Miliband who think only of the impact UKIP might have in 2015, and not beyond. Should UKIP’s insurgency continue, not only will it cause a rupture on the centre right, but also bring back into play Labour constituencies that have not been competitive for generations.This article was originally published in the Financial TimesTo comment on this article, please contact Chatham House Feedback Full Article
ot Promoting a Culture of Development and Investment: Lessons from the Post-War Era By feedproxy.google.com Published On :: Fri, 17 Oct 2014 10:30:01 +0000 Research Event 5 December 2014 - 1:00pm to 2:00pm Chatham House, London Event participants Giovanni Farese, Assistant Professor of Economic History, European University of RomeChair: Dr Paola Subacchi, Research Director, International Economics, Chatham House This event will discuss the rise of the culture of world development. It will examine the post-war reconstruction and development projects of the 1940s through to the 1960s, including those devised at Chatham House. The speaker will argue that these projects hold valuable lessons that still apply to the current economic environment. The speaker will also discuss the key role played by Eugene R Black (1898-1992), the third president of the International Bank for Reconstruction and Development (the World Bank), who was one of the main architects of post-war reconstruction and development projects and a promoter of a ‘culture of development’. Department/project Global Economy and Finance Programme Effie Theodoridou +44 (0)20 7314 2760 Email Full Article
ot Apolipoproteins of HDL can directly mediate binding to the scavenger receptor SR-BI, an HDL receptor that mediates selective lipid uptake By feedproxy.google.com Published On :: 1997-07-01 S XuJul 1, 1997; 38:1289-1298Articles Full Article
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ot Disruption of endoplasmic reticulum structure and integrity in lipotoxic cell death By feedproxy.google.com Published On :: 2006-12-01 Nica M. BorradaileDec 1, 2006; 47:2726-2737Research Articles Full Article
ot The role of short-chain fatty acids in the interplay between diet, gut microbiota, and host energy metabolism By feedproxy.google.com Published On :: 2013-09-01 Gijs den BestenSep 1, 2013; 54:2325-2340Reviews Full Article
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ot Procedure for determination of free and total cholesterol in micro- or nanogram amounts suitable for studies with cultured cells By feedproxy.google.com Published On :: 1978-11-01 W GambleNov 1, 1978; 19:1068-1070Articles Full Article
ot A spectrophotometric assay for lipid peroxides in serum lipoproteins using a commercially available reagent By feedproxy.google.com Published On :: 1989-04-01 M el-SaadaniApr 1, 1989; 30:627-630Articles Full Article
ot Thematic review series: The Pathogenesis of Atherosclerosis The oxidation hypothesis of atherogenesis: the role of oxidized phospholipids and HDL By feedproxy.google.com Published On :: 2004-06-01 Mohamad NavabJun 1, 2004; 45:993-1007Thematic Reviews Full Article
ot Fish oils and plasma lipid and lipoprotein metabolism in humans: a critical review By feedproxy.google.com Published On :: 1989-06-01 WS HarrisJun 1, 1989; 30:785-807Reviews Full Article
ot Normal high density lipoprotein inhibits three steps in the formation of mildly oxidized low density lipoprotein: steps 2 and 3 By feedproxy.google.com Published On :: 2000-09-01 Mohamad NavabSep 1, 2000; 41:1495-1508Articles Full Article