Member’s question time: Is Russia losing the South Caucasus? 22 October 2024 — 1:00PM TO 1:30PM Anonymous (not verified) 2 September 2024 Online

Join us and ask Chatham House Senior Research Fellow, Natalie Sabanadze anything about the situation in the Caucasus. Submit your questions in advance.

Whilst Russia focuses on its illegal invasion of Ukraine, the situation at its southern border is evolving. Relations between the three states in the South Caucasus and Moscow have never been easy as Russia tried to maintain its dominance by leveraging vulnerabilities, playing one side against another to keep conflicts simmering and even engaging in open military aggression.  Although the violence seen in the 1990s and early 2000s has abated, the war in Ukraine has had an indirect impact on the region, bringing a change to the status quo.

Russia abandoned its long-standing support for Armenia, allowing for the collapse of Nagorny-Karabakh and the restoration of the territorial integrity of Azerbaijan with the backing of Turkey. As a result, Azerbaijan has emerged as a dominant regional player with Baku recently declaring its interest to join BRICS.  Turkey’s influence has grown, while Armenia frustrated by Russia’s change of heart has been turning cautiously towards the EU and the US.

In Georgia, meanwhile, the ruling party has been consolidating its grasp on power, rolling back democratic reforms and pivoting away from the West. Georgia’s long-awaited European integration process has been suspended, following the adoption of the Russian-style foreign agents legislation.

Join us as our Senior Research Fellow in the Russia and Eurasia Programme answers your questions in this quick-fire session assessing the extent to which the Russian influence has changed since the start of the invasion of Ukraine and who is there to fill the vacuum; how geopolitical contestation in the region is going to impact aspirations of Armenia, Azerbaijan and Georgia; and much more.

Submit your questions to Natalie Sabanadze in advance of the event. Your questions will drive the conversation.

In their comment (1) on our publication (2), the authors make two points: (i) they raise concerns about the possible effect of residual NONOate in our study, and (ii) they promote nitrosylcobalamin (NOCbl) supplied by their own company. Both points lack merit for the following reasons. The authors make the astonishing claim that the spectra of nitric oxide (NO•) and cobalamins overlap. Unlike NO•, cobalamin absorbs in the visible region, permitting unequivocal spectral assignment of NOCbl as reported (3). We demonstrated that whereas NOCbl is highly unstable in solution, it is stabilized by the B12 trafficking protein CblC. So even if present, residual NONOate (which is unstable at neutral pH and is removed during the work-up (3)) could not account for the observed difference.The authors then misrepresent our synthetic method, claiming that anaerobic conditions were used to generate nitrocobalamin (NO2Cbl), which results in the transient formation of NOCbl. We synthesized NO2Cbl aerobically using nitrite as described (4); NOCbl is not an intermediate in this ligand exchange reaction. The aerobic instability of NOCbl has been rigorously described by inorganic chemists (3, 5) and raises obvious questions about its purported biological effects as exemplified by the authors' own 2003 JBC publication, which was later withdrawn.As to promoting NOCbl from their company, the authors refer to a synthetic route from a mixture of NO• gas and aquocobalamin. The authors' method (6) has been described as “dubious” by chemists (5). Whereas DEAE NONOate used in our method is widely known as an NO• donor,...

After a thorough read of this paper (1), we wish to clarify that the authors' anaerobic method of synthesis for the production of nitrocobalamin results in the transient formation of nitrosylcobalamin, an unstable intermediate upon exposure to air. We concur that the authors' method results in the production of nitrocobalamin based on the UV-visible data as shown. The authors' adapted anaerobic method consists of mixing hydroxocobalamin hydrochloride with diethylamine NONOate diethylammonium salt in aqueous solution. Of concern, the UV spectrum of nitric oxide overlaps that of all cobalamin species under anaerobic conditions, making any assignments of the binding of nitric oxide to hydroxocobalamin suspect (2). Additionally, the use of acetone to precipitate the authors' product causes precipitation of diethylamine NONOate, resulting in an impure product. As a result, its utility for drawing experimental conclusions is faulty.The product from the authors' anaerobic synthetic method has not been assessed for purity, and the synthetic method itself has not been validated using a stability-indicating method as required by the International Conference on Harmonization (ICH) (ICH Q2B, Validation of Analytical Procedures) methodology, which is a hallmark for analytical characterization. Our nitrosylcobalamin synthesis involves reacting nitric oxide gas with hydroxocobalamin acetate as a heterogeneous mixture in a non-electron-donating solvent followed by rotary evaporation. Our nitrosylcobalamin product is stable in air, releases nitric oxide gas in situ (3), and meets ICH stability guidelines (4). Additionally, our nitrosylcobalamin product demonstrates biological activity, which has not been observed for nitrocobalamin (3, 5).

Hdac3 is a lysine deacetylase that removes acetyl groups from histones and additional proteins. Although Hdac3 functions within mesenchymal lineage skeletal cells are defined, little is known about Hdac3 activities in bone-resorbing osteoclasts. In this study we conditionally deleted Hdac3 within Ctsk-expressing cells and examined the effects on bone modeling and osteoclast differentiation in mice. Hdac3 deficiency reduced femur and tibia periosteal circumference and increased cortical periosteal osteoclast number. Trabecular bone was likewise reduced and was accompanied by increased osteoclast number per trabecular bone surface. We previously showed that Hdac3 deacetylates the p65 subunit of the NF-κB transcriptional complex to decrease DNA-binding and transcriptional activity. Hdac3-deficient osteoclasts demonstrate increased K310 NF-κB acetylation and NF-κB transcriptional activity. Hdac3-deficient osteoclast lineage cells were hyper-responsive to RANKL and showed elevated ex vivo osteoclast number and size and enhanced bone resorption in pit formation assays. Osteoclast-directed Hdac3 deficiency decreased cortical and trabecular bone mass parameters, suggesting that Hdac3 regulates coupling of bone resorption and bone formation. We surveyed a panel of osteoclast-derived coupling factors and found that Hdac3 suppression diminished sphingosine-1-phosphate production. Osteoclast-derived sphingosine-1-phosphate acts in paracrine to promote bone mineralization. Mineralization of WT bone marrow stromal cells cultured with conditioned medium from Hdac3-deficient osteoclasts was markedly reduced. Expression of alkaline phosphatase, type 1a1 collagen, and osteocalcin was also suppressed, but no change in Runx2 expression was observed. Our results demonstrate that Hdac3 controls bone modeling by suppressing osteoclast lineage cell responsiveness to RANKL and coupling to bone formation.

Steven A. Carr
Mar 1, 2014; 13:907-917
Technological Innovation and Resources

Martin R. Larsen
Jul 1, 2005; 4:873-886
Technology

Philip L. Ross
Dec 1, 2004; 3:1154-1169
Research

Synapse loss is associated with motor and cognitive decline in multiple neurodegenerative disorders, and the cellular redistribution of tau is related to synaptic impairment in tauopathies, such as Alzheimer's disease and frontotemporal dementia. Here, we examined the cellular distribution of tau protein species in human tau overexpressing line 66 mice, a transgenic mouse model akin to genetic variants of frontotemporal dementia. Line 66 mice express intracellular tau aggregates in multiple brain regions and exhibit sensorimotor and motor learning deficiencies. Using a series of anti-tau antibodies, we observed, histologically, that nonphosphorylated transgenic human tau is enriched in synapses, whereas phosphorylated tau accumulates predominantly in cell bodies and axons. Subcellular fractionation confirmed that human tau is highly enriched in insoluble cytosolic and synaptosomal fractions, whereas endogenous mouse tau is virtually absent from synapses. Cytosolic tau was resistant to solubilization with urea and Triton X-100, indicating the formation of larger tau aggregates. By contrast, synaptic tau was partially soluble after Triton X-100 treatment and most likely represents aggregates of smaller size. MS corroborated that synaptosomal tau is nonphosphorylated. Tau enriched in the synapse of line 66 mice, therefore, appears to be in an oligomeric and nonphosphorylated state, and one that could have a direct impact on cognitive function.

How Beijing is closing surveillance gaps in the South China Sea The World Today iallan.drupal 15 October 2024

The discovery of a new radar system on China’s Triton Island military base shows that Beijing is rapidly developing its intelligence capacities in contested waters, write John Pollock and Damien Symon.

New satellite images reveal how the Chinese military is dramatically expanding its capabilities on Triton Island, which looks set to become a one of Beijing’s key signal intelligence bases in the South China Sea.

Once completed, the radar system would significantly increase China’s signals intercept and electronic warfare capabilities across the disputed Paracel Islands archipelago and add to a wider surveillance network spanning much of the South China Sea.

The enhanced facility on Triton is likely to offer a challenge to China’s competitors in the region and internationally.

China seized control of the Paracels from Vietnam in a 1974 naval battle, and competition for access to it waters has intensified since the recent discovery of oil and gas reserves. Chinese and Vietnamese maritime militia clashed off the coast of Triton in 2014.

In addition, American, British and Australian naval forces have for the past decade patrolled the waters to collectively challenge China’s contested ‘nine-dash-line’ claim to large stretches of the South China Sea. 

Overlapping anti-stealth network

A year after work was first identified on Triton, satellite images from Maxar have helped build a clearer picture of Beijing’s efforts to defend this strategic waterway.

The most striking development is the construction of a new radar system, known as SIAR – synthetic impulse and aperture radar – which purportedly detects stealth aircraft. The counter-stealth radar on Triton is characterized by its distinctive octagonal structure, which resembles another SIAR system built by China on Subi Reef in the Spratly Islands, south of the Paracels, in 2017. A half-completed tower also sits near the SIAR radar on Triton, which is likely to be the operations centre.

Once completed, the radar on Triton will form what is believed to be a wider network of at least three overlapping counter-stealth radars built across Chinese bases in the South China Sea over the past decade, including on Hainan Island, home to several Chinese naval bases. 

The positioning of the radar on Triton, 320km south of Hainan, is telling, says J. Michael Dahm, Senior Resident Fellow for Aerospace and China Studies at the Mitchell Institute. 

‘SIAR radars cannot see over the curve of the Earth, which means there is a gap in China’s air surveillance coverage between Subi Reef and Hainan Island. The Triton Island site will help close that gap’, said Dahm.

The aim, he suggests, is to give China contiguous counter-stealth radar coverage of the South China Sea.

More construction underway

The satellite images reveal other building projects on Triton. One is a large pad at the end of the road network which will probably be used as a launching point for a mobile anti-ship missile battery. The building at the northeast end of the road is probably a storage building for missile transport vehicles. 

Diminishing Vietnam’s options

The development of a new counter-stealth radar system and other suspected signals intercept structures on Triton represents a notable increase in China’s intelligence capabilities in the Paracels. The Chinese Communist Party has not disclosed the purpose of the building work on Triton, but its effects on regional and global competitors are likely to be wide ranging.

For Vietnam, which is rapidly expanding its own bases in the South China Sea, the intelligence structures on Triton would significantly diminish its capacity to operate undetected in the area. Alongside existing radar on Triton which can detect sea-going vessels, Beijing now has the potential to track Vietnamese air movements and gain forewarning of Hanoi’s manoeuvres in the area, including efforts to access oil and gas deposits.

The desire to strengthen control over these resources may explain why China is fortifying Triton Island, says Bill Hayton, Associate Fellow in the Asia-Pacific Programme, Chatham House. ‘The developments might be a warning that China is planning to mount another drilling expedition’, he suggests.

Addressing Russia’s use of forced displacement in Ukraine 7 November 2024 — 12:30PM TO 2:00PM Anonymous (not verified) 17 October 2024 Chatham House and Online

Experts consider the role international law could play in responding to Russia’s forcible movement of people during its war against Ukraine.

In the two and a half years since its full-scale invasion of Ukraine, evidence has emerged of Russia’s use of forced deportation and forcible transfer. Russia has also employed arbitrary detention as a tool of war and occupation.

Much attention has been on the International Criminal Court’s arrest warrants against Russian President Vladimir Putin and his children’s commissioner Maria Alekseyevna Lvova-Belova. These warrants were issued in relation to the alleged war crimes concerning the unlawful deportation of children from Ukraine to Russia and the unlawful transfer of thousands of children from occupied areas of Ukraine.

Meanwhile, Ukrainian citizens are being arrested and sent to Russia to serve prison sentences. They are often detained without charge and conviction.

This panel discussion explores:

• What evidence is emerging of Russia using unlawful deportation and transfer of children, and the arbitrary detention of civilians?
• What is the role and significance of international law on these issues?
• What challenges might these practices create for later peace negotiations, as well as the securing of justice?
• What is the process of releasing illegally detained Ukrainians, and Ukrainian children in particular, and reuniting them with their families? How do Russian volunteers inside Russia cooperate with Ukrainian NGOs to facilitate family reunification?

The event includes a screening of part of the documentary After the Rain: Putin’s Stolen Children. 

The institute occupies a position of respect and trust, and is committed to fostering inclusive dialogue at all events. Event attendees are expected to uphold this by adhering to our code of conduct.

The MENA uprisings: Five years on, what role is civil society playing? 31 October 2024 — 2:00PM TO 3:00PM Anonymous (not verified) 23 October 2024 Online

Panellists examine the protest movements’ legacies in different context and how civil society continues to work towards positive change.

Five years after nation-wide protests in Algeria, Iraq, Lebanon, and Sudan demanded systemic changes and political reforms, the situation remains largely unchanged. In some cases, it has deteriorated. Sudan is facing a humanitarian catastrophe amidst an ongoing civil war. Lebanon is dealing with Israeli aggression amidst a severe economic crisis with little response from the caretaker government. In Iraq and Algeria, relative stability masks the reality of increased suppression of dissent. This preservation of the status quo supports the entrenched political structures that strive to uphold it.

The current absence of large-scale street protests in these countries should not be taken as an indication that populations are content with the status quo. The issues that ignited the initial uprisings remain and in many cases have worsened. Despite enormous challenges, activists continue to navigate their systems to survive and instigate change. In the face of increasing difficulties, they are raising awareness of their countries’ predicaments and are finding alternative economic solutions. Additionally they are mobilizing community support, and pushing to voice their disillusionment. All these efforts aim at actively participating in shaping decisions that determines their future.

This webinar explores:

• What has been the impact of the uprisings in Algeria, Iraq, Lebanon, and Sudan?
• How are civil society and activists contributing to change within their communities?
• What is the current landscape for civic engagement within the politics, society and economy in these countries?
• What prospects are there for solidarity and cooperation among civil society actors across these regions and beyond?

Assessing the trajectory of the Middle East conflict 4 November 2024 — 4:00PM TO 5:00PM Anonymous (not verified) 29 October 2024 Online

Experts examine how the conflict may develop and what we can expect from regional and international actors.

A year on, the war in Gaza has spilled beyond Israel and Palestine with escalation across the region intensifying.

Recent weeks have seen Israel deepening its military offensive on Lebanon and keeping the north of the Gaza strip under siege, while leaders of Hezbollah and Hamas have been successfully targeted by its forces. Israel also launched an unprecedented assault against Iran in response to Tehran’s missile attacks on Israeli territory earlier in October.

Against this backdrop, regional states, particularly in the Gulf, in line with their overall approach to the conflict, are prioritizing diplomacy over escalation. They maintain their neutrality on the hostility between Israel and Iran and its aligned groups from the axis of resistance.

The strength of old alliances is being tested while new alignments are uncovered that may reshape the geopolitical landscape of the region, particularly following the US presidential election.

In this webinar, experts will examine:

• What are Israel’s calculations at this stage and how have the domestic political dynamics changed over recent weeks?
• What are the impacts of the war on Iran and its aligned actors and what can we expect from Tehran and groups from the axis of resistance?
• How are the wars in Gaza and Lebanon connected and would ending one stop the other?
• What is the response from regional states, particularly in the Gulf, and what role can they play?
• What are the possible scenarios for a post-election US policy on Israel and the Middle East?

Addressing illegal gold mining: International policy priorities 18 November 2024 — 11:30AM TO 1:00PM Anonymous (not verified) 6 November 2024 Chatham House and Online

Held in partnership with the World Gold Council, this panel of experts examines the global Artisanal and Small-Scale Gold Mining (ASGM) landscape and propose policy priorities essential for fostering sector formalization.

In this panel discussion, held in partnership with the World Gold Council, experts will examine the global Artisanal and Small-Scale Gold Mining (ASGM) landscape and propose policy priorities essential for fostering sector formalization.

An estimated twenty million people worldwide are involved in the Artisanal and Small-Scale Gold Mining (ASGM) sector, which now accounts for around 20 percent of global gold output. However, 85 percent of this production occurs outside formal legal frameworks. While many in ASGM operate within informal economies or seek pathways to formalization, a significant portion is also vulnerable to criminal exploitation, involving organized crime and armed groups.

In Ethiopia, the sector’s informality is tied to the nation’s volatile security dynamics, with illicit gold mining proliferating as non-state actors compete for control in conflict-prone regions such as Tigray. Additionally, gold is increasingly trafficked through transnational illicit networks in the Sahel and Sudan fuelling instability. South America is also impacted, for example in Peru where the government’s response to illegal ASGM initially saw success in 2019 but has faced sustainability challenges, and environmental impact.

ASGM growth is driven by the rising value of gold, youth unemployment, weak law enforcement, climate impacts, and conflict. While formalization offers significant economic potential, robust international cooperation and industry commitment is required to address the human toll, environmental impact and support sustainable practices.

In this panel discussion, experts will focus on addressing the expansion of illicit control and exploitation within ASGM, highlighting the pathways for governments and large-scale mining companies to drive meaningful change.

This event is hosted in partnership with the World Gold Council. There will be a reception with light refreshments hosted at Chatham House following the event.

This event will be livestreamed via the Africa Programme Facebook page.

The institute occupies a position of respect and trust, and is committed to fostering inclusive dialogue at all events. Event attendees are expected to uphold this by adhering to our code of conduct.

M el-Saadani
Apr 1, 1989; 30:627-630
Articles

Expert

In animals, the response to chronic hypoxia is mediated by prolyl hydroxylases (PHDs) that regulate the levels of hypoxia-inducible transcription factor α (HIFα). PHD homologues exist in other types of eukaryotes and prokaryotes where they act on non HIF substrates. To gain insight into the factors underlying different PHD substrates and properties, we carried out biochemical and biophysical studies on PHD homologues from the cellular slime mold, Dictyostelium discoideum, and the protozoan parasite, Toxoplasma gondii, both lacking HIF. The respective prolyl-hydroxylases (DdPhyA and TgPhyA) catalyze prolyl-hydroxylation of S-phase kinase-associated protein 1 (Skp1), a reaction enabling adaptation to different dioxygen availability. Assays with full-length Skp1 substrates reveal substantial differences in the kinetic properties of DdPhyA and TgPhyA, both with respect to each other and compared with human PHD2; consistent with cellular studies, TgPhyA is more active at low dioxygen concentrations than DdPhyA. TgSkp1 is a DdPhyA substrate and DdSkp1 is a TgPhyA substrate. No cross-reactivity was detected between DdPhyA/TgPhyA substrates and human PHD2. The human Skp1 E147P variant is a DdPhyA and TgPhyA substrate, suggesting some retention of ancestral interactions. Crystallographic analysis of DdPhyA enables comparisons with homologues from humans, Trichoplax adhaerens, and prokaryotes, informing on differences in mobile elements involved in substrate binding and catalysis. In DdPhyA, two mobile loops that enclose substrates in the PHDs are conserved, but the C-terminal helix of the PHDs is strikingly absent. The combined results support the proposal that PHD homologues have evolved kinetic and structural features suited to their specific sensing roles.

Stacey Finley from University of Southern California discusses how mathematical models support the research of cancer biology. Cancer research is a crucial job, but a difficult one. Tumors growing inside the human body are affected by all kinds of factors. These conditions are difficult (if not impossible) to recreate in the lab, and using real patients as subjects can be painful and invasive. Mathematical models give cancer researchers the ability to run experiments virtually, testing the effects of any number of factors on tumor growth and other processes — all with far less money and time than an experiment on human subjects or in the lab would use.

Benjamin Jaye and Manasa N. Vempati
Proc. Amer. Math. Soc. 152 (), 5105-5116.
Abstract, references and article information

R. Gibara and D. Kinzebulatov
St. Petersburg Math. J. 35 (), 445-460.
Abstract, references and article information

Dynamic singer Tosh Alexander has been lighting up the music scene with her latest track, ' My Ting Different', a thrilling collaboration with American rapper and songwriter Lady London. The song fuses R...

Nigy Boy's management team was forced to put out a scam alert on Thursday as news surfaced that a promoter in the Turks and Caicos Islands, inadvertently wired thousands of US dollars to who he believed was the artiste's booking team as a deposit...

When a scuba diver comes up from the deep too quickly, the rapid decrease in pressure can give them a case of "the bends" or decompression sickness. It is caused by bubbles of gas building up in the body, causing pain. It can also be fatal....

If you connect to a FTP/TLS server that is configured to use implicit FTP/TLS, FILENAME FTP/TLS might fail with the following error:

CD81 plays a central role in a variety of physiological and pathological processes. Recent structural analysis of CD81 indicates that it contains an intramembrane cholesterol-binding pocket and that interaction with cholesterol may regulate a conformational switch in the large extracellular domain of CD81. Therefore, CD81 possesses a potential cholesterol-sensing mechanism; however, its relevance for protein function is thus far unknown. In this study we investigate CD81 cholesterol sensing in the context of its activity as a receptor for hepatitis C virus (HCV). Structure-led mutagenesis of the cholesterol-binding pocket reduced CD81–cholesterol association but had disparate effects on HCV entry, both reducing and enhancing CD81 receptor activity. We reasoned that this could be explained by alterations in the consequences of cholesterol binding. To investigate this further we performed molecular dynamic simulations of CD81 with and without cholesterol; this identified a potential allosteric mechanism by which cholesterol binding regulates the conformation of CD81. To test this, we designed further mutations to force CD81 into either the open (cholesterol-unbound) or closed (cholesterol-bound) conformation. The open mutant of CD81 exhibited reduced HCV receptor activity, whereas the closed mutant enhanced activity. These data are consistent with cholesterol sensing switching CD81 between a receptor active and inactive state. CD81 interactome analysis also suggests that conformational switching may modulate the assembly of CD81–partner protein networks. This work furthers our understanding of the molecular mechanism of CD81 cholesterol sensing, how this relates to HCV entry, and CD81's function as a molecular scaffold; these insights are relevant to CD81's varied roles in both health and disease.

Replication protein A (RPA) is a eukaryotic ssDNA-binding protein and contains three subunits: RPA70, RPA32, and RPA14. Phosphorylation of the N-terminal region of the RPA32 subunit plays an essential role in DNA metabolism in processes such as replication and damage response. Phosphorylated RPA32 (pRPA32) binds to RPA70 and possibly regulates the transient RPA70-Bloom syndrome helicase (BLM) interaction to inhibit DNA resection. However, the structural details and determinants of the phosphorylated RPA32–RPA70 interaction are still unknown. In this study, we provide molecular details of the interaction between RPA70 and a mimic of phosphorylated RPA32 (pmRPA32) using fluorescence polarization and NMR analysis. We show that the N-terminal domain of RPA70 (RPA70N) specifically participates in pmRPA32 binding, whereas the unphosphorylated RPA32 does not bind to RPA70N. Our NMR data revealed that RPA70N binds pmRPA32 using a basic cleft region. We also show that at least 6 negatively charged residues of pmRPA32 are required for RPA70N binding. By introducing alanine mutations into hydrophobic positions of pmRPA32, we found potential points of contact between RPA70N and the N-terminal half of pmRPA32. We used this information to guide docking simulations that suggest the orientation of pmRPA32 in complex with RPA70N. Our study demonstrates detailed features of the domain-domain interaction between RPA70 and RPA32 upon phosphorylation. This result provides insight into how phosphorylation tunes transient bindings between RPA and its partners in DNA resection.

8 July 2020

17 July 2020

12 August 2020

Cellular energy demands are met by uptake and metabolism of nutrients like glucose. The principal transcriptional regulator for adapting glycolytic flux and downstream pathways like de novo lipogenesis to glucose availability in many cell types is carbohydrate response element–binding protein (ChREBP). ChREBP is activated by glucose metabolites and post-translational modifications, inducing nuclear accumulation and regulation of target genes. Here we report that ChREBP is modified by proline hydroxylation at several residues. Proline hydroxylation targets both ectopically expressed ChREBP in cells and endogenous ChREBP in mouse liver. Functionally, we found that specific hydroxylated prolines were dispensable for protein stability but required for the adequate activation of ChREBP upon exposure to high glucose. Accordingly, ChREBP target gene expression was rescued by re-expressing WT but not ChREBP that lacks hydroxylated prolines in ChREBP-deleted hepatocytes. Thus, proline hydroxylation of ChREBP is a novel post-translational modification that may allow for therapeutic interference in metabolic diseases.

Visual Abstract

Visual Abstract

Xiao-Ting Wen
Dec 17, 2020; 0:RA120.002119v1-mcp.RA120.002119
Research

Alison M. Kurimchak
Dec 1, 2020; 19:2068-2089
Research

When you publish a model to SAS Metadata Repository by using SAS Model Manager, the publishing process fails and the following error is generated: "The model model-name has a function of ';Transformation';, which is not supported for

Lipoprotein (a) [Lp(a)] is a risk factor for CVD and a target of therapy, but Lp(a) measurements are not globally standardized. Commercially available assays generally use polyclonal antibodies that detect multiple sites within the kringle (K)IV₂ repeat region of Lp(a) and may lead to inaccurate assessments of plasma levels. With increasing awareness of Lp(a) as a cardiovascular risk factor and the active clinical development of new potential therapeutic approaches, the broad availability of reagents capable of providing isoform independence of Lp(a) measurements is paramount. To address this issue, we generated a murine monoclonal antibody that binds to only one site on apo(a). A BALB/C mouse was immunized with a truncated version of apo(a) that contained eight total KIV repeats, including only one copy of KIV₂. We generated hybridomas, screened them, and successfully produced a KIV₂-independent monoclonal antibody, named LPA-KIV9. Using a variety of truncated apo(a) constructs to map its binding site, we found that LPA-KIV9 binds to KIV₉ without binding to plasminogen. Fine peptide mapping revealed that LPA-KIV9 bound to the sequence ⁴⁰⁷⁶LETPTVV⁴⁰⁸² on KIV₉. In conclusion, the generation of monoclonal antibody LPA-KIV9 may be a useful reagent in basic research studies and in the clinical application of Lp(a) measurements.

Atherosclerosis is characterized by the pathological accumulation of cholesterol-laden macrophages in the arterial wall. Atherosclerosis is also the main underlying cause of CVDs, and its development is largely driven by elevated plasma cholesterol. Strong epidemiological data find an inverse association between plasma β-carotene with atherosclerosis, and we recently showed that β-carotene oxygenase 1 (BCO1) activity, responsible for β-carotene cleavage to vitamin A, is associated with reduced plasma cholesterol in humans and mice. In this study, we explore whether intact β-carotene or vitamin A affects atherosclerosis progression in the atheroprone LDLR-deficient mice. Compared with control-fed Ldlr^–/– mice, β-carotene-supplemented mice showed reduced atherosclerotic lesion size at the level of the aortic root and reduced plasma cholesterol levels. These changes were absent in Ldlr^–/–/Bco1^–/– mice despite accumulating β-carotene in plasma and atherosclerotic lesions. We discarded the implication of myeloid BCO1 in the development of atherosclerosis by performing bone marrow transplant experiments. Lipid production assays found that retinoic acid, the active form of vitamin A, reduced the secretion of newly synthetized triglyceride and cholesteryl ester in cell culture and mice. Overall, our findings provide insights into the role of BCO1 activity and vitamin A in atherosclerosis progression through the regulation of hepatic lipid metabolism.

Fabry disease is caused by deficient activity of α-galactosidase A, an enzyme that hydrolyzes the terminal α-galactosyl moieties from glycolipids and glycoproteins, and subsequent accumulation of glycosphingolipids, mainly globotriaosylceramide (Gb₃), globotriaosylsphingosine (lyso-Gb₃), and galabiosylceramide. However, there is no known link between these compounds and disease severity. In this study, we compared Gb₃ isoforms (various fatty acids) and lyso-Gb₃ analogs (various sphingosine modifications) in two strains of Fabry disease mouse models: a pure C57BL/6 (B6) background or a B6/129 mixed background, with the latter exhibiting more prominent cardiac and renal hypertrophy and thermosensation deficits. Total Gb₃ and lyso-Gb₃ levels in the heart, kidney, and dorsal root ganglion (DRG) were similar in the two strains. However, levels of the C20-fatty acid isoform of Gb₃ and particular lyso-Gb₃ analogs (+18, +34) were significantly higher in Fabry-B6/129 heart tissue when compared with Fabry-B6. By contrast, there was no difference in Gb₃ and lyso-Gb₃ isoforms/analogs in the kidneys and DRG between the two strains. Furthermore, using immunohistochemistry, we found that Gb₃ massively accumulated in DRG mechanoreceptors, a sensory neuron subpopulation with preserved function in Fabry disease. However, Gb₃ accumulation was not observed in nonpeptidergic nociceptors, the disease-relevant subpopulation that has remarkably increased isolectin-B4 (the marker of nonpeptidergic nociceptors) binding and enlarged cell size. These findings suggest that specific species of Gb₃ or lyso-Gb₃ may play major roles in the pathogenesis of Fabry disease, and that Gb₃ and lyso-Gb₃ are not responsible for the pathology in all tissues or cell types.

The neuronal basis of complex social behavior is still poorly understood. In honeybees, reproductive investment decisions are made at the colony-level. Queens develop from female-destined larvae that receive alloparental care from nurse bees in the form of ad-libitum royal jelly (RJ) secretions. Typically, the number of raised new queens is limited but genetic breeding of "royal jelly bees" (RJBs) for enhanced RJ production over decades has led to a dramatic increase of reproductive investment in queens. Here, we compare RJBs to unselected Italian bees (ITBs) to investigate how their cognitive processing of larval signals in the mushroom bodies (MBs) and antennal lobes (ALs) may contribute to their behavioral differences. A cross-fostering experiment confirms that the RJB syndrome is mainly due to a shift in nurse bee alloparental care behavior. Using olfactory conditioning of the proboscis extension reflex, we show that the RJB nurses spontaneously respond more often to larval odors compared with ITB nurses but their subsequent learning occurs at similar rates. These phenotypic findings are corroborated by our demonstration that the proteome of the brain, particularly of the ALs differs between RJBs and ITBs. Notably, in the ALs of RJB newly emerged bees and nurses compared with ITBs, processes of energy and nutrient metabolism, signal transduction are up-regulated, priming the ALs for receiving and processing the brood signals from the antennae. Moreover, highly abundant major royal jelly proteins and hexamerins in RJBs compared with ITBs during early life when the nervous system still develops suggest crucial new neurobiological roles for these well-characterized proteins. Altogether, our findings reveal that RJBs have evolved a strong olfactory response to larvae, enabled by numerous neurophysiological adaptations that increase the nurse bees' alloparental care behavior.

MS-based proteome profiling has become increasingly comprehensive and quantitative, yet a persistent shortcoming has been the relatively large samples required to achieve an in-depth measurement. Such bulk samples, typically comprising thousands of cells or more, provide a population average and obscure important cellular heterogeneity. Single-cell proteomics capabilities have the potential to transform biomedical research and enable understanding of biological systems with a new level of granularity. Recent advances in sample processing, separations and MS instrumentation now make it possible to quantify >1000 proteins from individual mammalian cells, a level of coverage that required an input of thousands of cells just a few years ago. This review discusses important factors and parameters that should be optimized across the workflow for single-cell and other low-input measurements. It also highlights recent developments that have advanced the field and opportunities for further development.

Endometrial carcinoma (EC) is the most common gynecologic malignancy in the United States, with limited effective targeted therapies. Endometrial tumors exhibit frequent alterations in protein kinases, yet only a small fraction of the kinome has been therapeutically explored. To identify kinase therapeutic avenues for EC, we profiled the kinome of endometrial tumors and normal endometrial tissues using Multiplexed Inhibitor Beads and Mass Spectrometry (MIB-MS). Our proteomics analysis identified a network of kinases overexpressed in tumors, including Serine/Arginine-Rich Splicing Factor Kinase 1 (SRPK1). Immunohistochemical (IHC) analysis of endometrial tumors confirmed MIB-MS findings and showed SRPK1 protein levels were highly expressed in endometrioid and uterine serous cancer (USC) histological subtypes. Moreover, querying large-scale genomics studies of EC tumors revealed high expression of SRPK1 correlated with poor survival. Loss-of-function studies targeting SRPK1 in an established USC cell line demonstrated SRPK1 was integral for RNA splicing, as well as cell cycle progression and survival under nutrient deficient conditions. Profiling of USC cells identified a compensatory response to SRPK1 inhibition that involved EGFR and the up-regulation of IGF1R and downstream AKT signaling. Co-targeting SRPK1 and EGFR or IGF1R synergistically enhanced growth inhibition in serous and endometrioid cell lines, representing a promising combination therapy for EC.

Ankylosing Spondylitis (AS) is a common, inflammatory rheumatic disease, which primarily affects the axial skeleton and is associated with sacroiliitis, uveitis and enthesitis. Unlike other autoimmune rheumatic diseases, such as rheumatoid arthritis or systemic lupus erythematosus, autoantibodies have not yet been reported to be a feature of AS. We therefore wished to determine if plasma from patients with AS contained autoantibodies and if so, characterize and quantify this response in comparison to patients with Rheumatoid Arthritis (RA) and healthy controls. Two high-density nucleic acid programmable protein arrays expressing a total of 3498 proteins were screened with plasma from 25 patients with AS, 17 with RA and 25 healthy controls. Autoantigens identified were subjected to Ingenuity Pathway Analysis in order to determine patterns of signalling cascades or tissue origin. 44% of patients with Ankylosing Spondylitis demonstrated a broad autoantibody response, as compared to 33% of patients with RA and only 8% of healthy controls. Individuals with AS demonstrated autoantibody responses to shared autoantigens, and 60% of autoantigens identified in the AS cohort were restricted to that group. The AS patients autoantibody responses were targeted towards connective, skeletal and muscular tissue, unlike those of RA patients or healthy controls. Thus, patients with AS show evidence of systemic humoral autoimmunity and multispecific autoantibody production. Nucleic Acid Programmable Protein Arrays constitute a powerful tool to study autoimmune diseases.