Host Matt Egan is joined this week by Henry Burrell, staff writer at Macworld UK to talk about the EU referendum website crashing Lewis Painter, staff writer at PC Advisor, jumps in to chat about Oculus Rift games and how immersive they are. Finally regular David Price comes on to talk about Apple's upcoming WWDC software developer conference and what we can (and cannot) expect.  

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In Matt Egan's absence first time host Henry Burrell heroically steps in to chat about Pokemon Go with producer Chris. Then fellow staff writer Chris Minasians discusses iOS 10 beta and all of the rumoured features for the latest Apple operating system, including dark mode (10:30). Finally, Lewis Painter, staff writer at PC Advisor, talks about Playstation VR being released for the PS4, pricing and games (23:30).  

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Macworld UK’s Acting Editor David Price takes the reins for this edition, and is joined by Online Editor of Computerworld UK Scott Carey to chat about billions and billions of pounds and the acquisition of ARM by SoftBank. Second up, Staff Writer at PC Advisor and Macworld UK Christopher Minasians plugs in to the haunting world of graphics cards and makes sense of it all for the rest of us. Digital Arts Staff Writer Mimi Launder then explains why Twitter has slapped a troll right in the face in order to stand up to Internet bullying.  

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Hosting duties fall to Henry Burrell this week as he discusses the deadline for the free Microsoft Windows 10 update with Chris Minasians, staff writer at PC Advisor. Scott Carey, online editor at Techworld.com jumps in to talk about why the Verizon deal for Yahoo is ridiculous and charts the missteps that got the company to this point (15:00). Finally, regular guest David Price discusses Apple's less than stellar financial results and if the iPhone is plateauing (26:00).  

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Host Matt Egan clips us round the ear and tells us to listen up as we chat yet more tech and then some other stuff about tech. Consumer tech editor at PC Advisor Chris Martin lays down his definitive opinion after he went hands on with the Nintendo Switch this week, and why the company really should have had their star plumber ready in time for launch. Tamlin Magee, Online Editor at Computerworld UK then takes us through the odd goings on at Davos, and whether or not the elite can identify with what tech actually means to real working people. To round us up, Acting Macworld UK Editor David Price explains why app prices are going up in the UK for iOS users, and why it might - might - not be UKIP's fault. Sort of.  

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The big story of the week is Wikileaks' CIA data dump, and we sandwich that topic between Apple's upcoming event and Nintendo Switch hardware issues. Lewis Painter kicks things off with Apple's (fingers crossed) late March event where we hope to see iPads, iMacs and get our six monthly fix of Jony Ive product videos. Then (13 minutes) Tamlin Magee tackles Vault 7 and the CIA's apparent ability to take control of iPhones and TVs, bypassing encryption. Will continuous leaks change anything? Finally (26 minutes) Dom Preston talks about his time over the last few weeks playing Zelda on the Nintendo Switch and why we should be a bit concerned about dodgy controllers.  

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David Price is back to host this week, with Christina Mercer breaking down what all the fuss about KRACK is and why you should take precautions with your Wi-Fi. Henry Burrell reviewed the Google Pixel 2 this week so gives us his thoughts on the device. You might want to avoid the larger XL though. We then discuss Facebook's idea to counterattack cyberbullying. Charlotte Jee recently spoke with the company to see what they are doing, but will it work?  

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We finally get the IoT acronym the pod has been waiting for as we discuss Bitcoin and 17th century Dutch finance. Charlotte Jee informs Henry Burrell, David Price and Scott Carey.

Scott then leads us down the winding road of the so-called platform wars, in a week when Google blocked the Amazon Echo Show from streaming YouTube. Is this a bump in the road or will services be cut from rival hardware?

And finally, led by David, the pod discusses its favourite tech of the year and what you might consider buying for friends and family for Christmas.

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This week it’s David Price in the hosting seat to dig into two meaty tech topics: how does Netflix buy and develop its massive content library, and why is Fortnite such a sensation?

Helping him dig into Netflix is Computerworld UK editor Scott Carey, with our new entertainment and lifestyle editor at Tech Advisor Dominic Preston joining in.

Then staff writer at Tech Advisor Sean Bradley is on hand to talk about how Fortnite has become such a sensation, and if it is built to last.

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This week our host Scott Carey is joined by Macworld UK editor Karen Khan to chat about Apple's latest blockbuster results.

Then group production editor Tamlin Magee jumps in to discuss Amazon's working practices following the collective action around Prime Day.

Finally, Scott chats through his experience at the Google Cloud Next conference in San Francisco last week to see how it is trying to compete with the big boys at Amazon and Microsoft.

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Inhibition of acid sphingomyelinase (ASM), a lysosomal enzyme that catalyzes the hydrolysis of sphingomyelin into ceramide and phosphorylcholine, may serve as an investigational tool or a therapeutic intervention to control many diseases. Specific ASM inhibitors are currently not sufficiently characterized. Here, we found that 1-aminodecylidene bis-phosphonic acid (ARC39) specifically and efficiently (>90%) inhibits both lysosomal and secretory ASM in vitro. Results from investigating sphingomyelin phosphodiesterase 1 (SMPD1/Smpd1) mRNA and ASM protein levels suggested that ARC39 directly inhibits ASM’s catalytic activity in cultured cells, a mechanism which differs from that of functional inhibitors of ASM (FIASMAs). We further provide evidence that ARC39 dose- and time-dependently inhibits lysosomal ASM in intact cells, and we show that ARC39 also reduces platelet- and ASMpromoted adhesion of tumor cells. The observed toxicity of ARC39 is low at concentrations relevant for ASM inhibition in vitro, and it does not strongly alter the lysosomal compartment or induce phospholipidosis in vitro. When applied intraperitoneally in vivo, even subtoxic high doses administered short-term induced sphingomyelin accumulation only locally in the peritoneal lavage without significant accumulation in plasma, liver, spleen or brain. These findings require further investigation with other possible chemical modifications. In conclusion, our results indicate that ARC39 potently and selectively inhibits ASM in vitro and highlight the need for developing compounds that can reach tissue concentrations sufficient for ASM inhibition in vivo.

Excessive lipid deposition is a hallmark of nonalcoholic fatty liver disease (NAFLD). Although much has been learned about the enzymes and metabolites involved in NAFLD, few studies have focused on the role of long non-coding RNAs (lncRNAs) in hepatic lipid accumulation. Here, using in vitro and in vivo models of NAFLD, we found that the lncRNA Gm15622 is highly expressed in the liver of obese mice fed a high-fat diet (HFD) and in murine liver (AML-12) cells treated with free fatty acids. Investigating the molecular mechanism in the liver-enriched expression of Gm15622 and its effects on lipid accumulation in hepatocytes and on NAFLD pathogenesis, we found that Gm15622 acts as a sponge for the microRNA miR-742-3p. This sponging activity increased the expression of the transcriptional regulator sterol regulatory element–binding transcription factor 1c (SREBP-1c) and promoted lipid accumulation in the liver of the HFD mice and AML-12 cells. Moreover, further results indicated that metformin suppresses Gm15622 and alleviates NAFLD-associated lipid deposition in mice. In conclusion, we have identified an lncRNA Gm15622–miR-742-3p–SREBP-1c regulatory circuit associated with NAFLD in mice, a finding that significantly advances our insight into how lipid metabolism and accumulation are altered in this metabolic disorder. Our results also suggest that Gm15622 may be a potential therapeutic target for managing NAFLD.

Ether lipids (ELs) are lipids characterized by the presence of either an ether linkage (alkyl lipids) or a vinyl ether linkage (i.e. plasmalogens [Pls]) at the sn1 position of the glycerol backbone and they are enriched in PUFAs at the sn2 position. In this review, we highlight that ELs have various biological functions, act as a reservoir for second messengers (such as PUFAs), and have roles in many diseases. Some of the biological effects of ELs may be associated with their ability to regulate ion channels that control excitation-contraction/secretion/mobility coupling and therefore cell physiology. These channels are embedded in lipid membranes, and lipids can regulate their activities directly or indirectly as second messengers or by incorporating into membranes. Interestingly, ELs and EL-derived PUFAs have been reported to play a key role in several pathologies, including neurological disorders, cardiovascular diseases, and cancers. Investigations leading to a better understanding of their mechanisms of action in pathologies have opened a new field in cancer research. In summary, newly identified lipid regulators of ion channels, such as ELs and PUFAs, may represent valuable targets to improve disease diagnosis and advance the development of new therapeutic strategies for managing a range of diseases and conditions.

Oxidized low-density lipoprotein (oxLDL) is a known risk factor for atherogenesis. This study aimed to reveal structural features of oxLDL present in human circulation related to atherosclerosis. When LDL was fractionated on an anion-exchange column, in vivo-oxLDL, detected by the anti-oxidized phosphatidylcholine (oxPC) monoclonal antibody, was recovered in flow-through and electronegative LDL (LDL(-)) fractions. The amount of the electronegative in vivo-oxLDL, namely oxLDL in LDL(-) fraction, present in patients with acute myocardial infarction (AMI) was three-fold higher than that observed in healthy subjects. Surprisingly, LDL(-) fraction contained apoA1 in addition to apoB, and HDL-sized particles were observed with transmission electron microscopy. In LDL(-) fractions, acrolein adducts were identified at all lysine residues in apoA1, with only a small number of acrolein-modified residues were identified in apoB. The amount of oxPC adducts of apoB was higher in LDL(-) than in L1 fraction as determined using western blotting. The electronegative in vivo-oxLDL was immunologically purified from the LDL(-) fraction with an anti-oxPC monoclonal antibody. Majority of PC species was not oxidized, whereas oxPC and lysoPC did not accumulate. Here, we propose that there are two types of in vivo-oxLDL in human circulating plasma and the electronegative in vivo-oxLDL accompanies oxidized HDL.

Bis(monoacylglycero)phosphate (BMP), also known as lysobisphosphatidic acid (LBPA), is a phospholipid that promotes lipid sorting in late endosomes/lysosomes by activating lipid hydrolases and lipid transfer proteins. Changes in the cellular BMP content therefore reflect an altered metabolic activity of the endo-lysosomal system. Surprisingly, little is known about the physiological regulation of BMP. In this study, we investigated the effects of nutritional and metabolic factors on BMP profiles of whole tissues and  parenchymal and non-parenchymal cells. Tissue samples were obtained from fed, fasted, two-hours refed, and insulin-treated mice, as well as from mice housed at  5°C, 22°C, or 30°C. These tissues exhibited distinct BMP profiles, which were regulated by the nutritional state in a tissue-specific manner. Insulin treatment was not sufficient to mimic refeeding-induced changes in tissue BMP levels indicating that BMP metabolism is regulated by other hormonal or nutritional factors. Tissue fractionation experiments revealed that fasting drastically elevates BMP levels in hepatocytes and pancreatic cells. Furthermore, we observed that the BMP content in brown adipose tissue strongly depends on housing temperatures. In conclusion, our observations suggest that BMP concentrations adapt to the metabolic state in a tissue-and cell type-specific manner in mice. Drastic changes observed in hepatocytes, pancreatic cells, and brown adipocytes suggest that BMP possesses a role in the functional adaption to nutrient starvation and ambient temperature.

Niemann-Pick disease, type C1 (NPC1) is a lipid storage disorder in which cholesterol and glycosphingolipids accumulate in late endosomal/lysosomal compartments because of mutations in the NPC1 gene. A hallmark of NPC1 is progressive neurodegeneration of the cerebellum as well as visceral organ damage; however, the mechanisms driving this disease pathology are not fully understood. Phosphoinositides are phospholipids that play distinct roles in signal transduction and vesicle trafficking. Here, we utilized consensus spectra analysis of MS imaging datasets and orthogonal LC–MS analyses to evaluate the spatial distribution of phosphoinositides and quantify them in cerebellar tissue from Npc1-null mice. Our results suggest significant depletion of multiple phosphoinositide species, including phosphatidylinositol (PI), phosphatidylinositol monophosphate (PIP), and bisphosphate (PIP2), in the cerebellum of the Npc1-null mice in both whole-tissue lysates and myelin-enriched fractions. Additionally, we observed altered levels of the regulatory enzyme phosphatidylinositol 4-kinase type 2 α (PI4K2A) in Npc1-null mice. In contrast, the levels of related kinases, phosphatases, and transfer proteins were unaltered in the Npc1-null mouse model as observed by Western blot analysis. Our discovery of phosphoinositide lipid biomarkers for NPC1 opens new perspectives on the pathophysiology underlying this fatal neurodegenerative disease.

7 April 2020

15 April 2020

7 May 2020

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.

In quantitative proteomics work, the differences in expression of many separate proteins are routinely examined to test for significant differences between treatments. This leads to the multiple hypothesis testing problem: when many separate tests are performed many will be significant by chance and be false positive results. Statistical methods such as the false discovery rate (FDR) method that deal with this problem have been disseminated for more than one decade. However a survey of proteomics journals shows that such tests are not widely implemented in one commonly used technique, quantitative proteomics using two-dimensional electrophoresis (2-DE). We outline a selection of multiple hypothesis testing methods, including some that are well known and some lesser known, and present a simple strategy for their use by the experimental scientist in quantitative proteomics work generally. The strategy focuses on the desirability of simultaneous use of several different methods, the choice and emphasis dependent on research priorities and the results in hand. This approach is demonstrated using case scenarios with experimental and simulated model data.

Oxidative stress has been implicated in aging and many human diseases, notably neurodegenerative disorders and various cancers. The reactive oxygen species that are generated by aerobic metabolism and environmental stressors can chemically modify proteins and alter their biological functions. Cells possess protein repair pathways to rescue oxidized proteins and restore their functions. If these repair processes fail, oxidized proteins may become cytotoxic. Cell homeostasis and viability are therefore dependent on the removal of oxidatively damaged proteins. Numerous studies have demonstrated that the proteasome plays a pivotal role in the selective recognition and degradation of oxidized proteins. Despite extensive research, oxidative stress-triggered regulation of proteasome complexes remains poorly defined. Better understanding of molecular mechanisms underlying proteasome function in response to oxidative stress will provide a basis for developing new strategies aimed at improving cell viability and recovery as well as attenuating oxidation-induced cytotoxicity associated with aging and disease. Here we highlight recent advances in the understanding of proteasome structure and function during oxidative stress and describe how cells cope with oxidative stress through proteasome-dependent degradation pathways.

Protein tyrosine kinases (PTKs) play key roles in cellular signal transduction, cell cycle regulation, cell division, and cell differentiation. Dysregulation of PTK-activated pathways, often by receptor overexpression, gene amplification, or genetic mutation, is a causal factor underlying numerous cancers. In this study, we have developed a parallel reaction monitoring (PRM)-based assay for quantitative profiling of 83 PTKs. The assay detects 308 proteotypic peptides from 54 receptor tyrosine kinases and 29 nonreceptor tyrosine kinases in a single run. Quantitative comparisons were based on the labeled reference peptide method. We implemented the assay in four cell models: 1) a comparison of proliferating versus epidermal growth factor (EGF)-stimulated A431 cells, 2) a comparison of SW480Null (mutant APC) and SW480APC (APC restored) colon tumor cell lines, and 3) a comparison of 10 colorectal cancer cell lines with different genomic abnormalities, and 4) lung cancer cell lines with either susceptibility (11-18) or acquired resistance (11-18R) to the epidermal growth factor receptor tyrosine kinase inhibitor erlotinib. We observed distinct PTK expression changes that were induced by stimuli, genomic features or drug resistance, which were consistent with previous reports. However, most of the measured expression differences were novel observations. For example, acquired resistance to erlotinib in the 11-18 cell model was associated not only with previously reported upregulation of MET, but also with upregulation of FLK2 and downregulation of LYN and PTK7. Immunoblot analyses and shotgun proteomics data were highly consistent with PRM data. Multiplexed PRM assays provide a targeted, systems-level profiling approach to evaluate cancer-related proteotypes and adaptations. Data are available through Proteome eXchange Accession PXD002706.

Stimulating brown adipose tissue (BAT) activity represents a promising therapy for overcoming metabolic diseases. mTORC2 is important for regulating BAT metabolism, but its downstream targets have not been fully characterized. In this study, we apply proteomics and phosphoproteomics to investigate the downstream effectors of mTORC2 in brown adipocytes. We compare wild-type controls to isogenic cells with an induced knockout of the mTORC2 subunit RICTOR (Rictor-iKO) by stimulating each with insulin for a 30-minute time course. In Rictor-iKO cells, we identify decreases to the abundance of glycolytic and de novo lipogenesis enzymes, and increases to mitochondrial proteins as well as a set of proteins known to increase upon interferon stimulation. We also observe significant differences to basal phosphorylation due to chronic RICTOR loss including decreased phosphorylation of the lipid droplet protein perilipin-1 in Rictor-iKO cells, suggesting that RICTOR could be involved with regulating basal lipolysis or droplet dynamics. Finally, we observe mild dampening of acute insulin signaling response in Rictor-iKO cells, and a subset of AKT substrates exhibiting statistically significant dependence on RICTOR.

The human leucocyte antigen (HLA)-B*27:05 allele and the endoplasmic reticulum-resident aminopeptidases are strongly associated with ankylosing spondylitis (AS), a chronic inflammatory spondyloarthropathy. This study examined the effect of endoplasmic reticulum aminopeptidase 2 (ERAP2) in the generation of the natural HLA-B*27:05 ligandome in live cells. Complexes of HLA-B*27:05-bound peptide pools were isolated from human ERAP2-edited cell clones and the peptides were identified using high throughput mass spectrometry analyses. The relative abundance of thousand ligands was established by quantitative tandem mass spectrometry and bioinformatics analysis. The residue frequencies at different peptide position, identified in presence or absence of ERAP2, determined structural features of ligands and their interactions with specific pockets of antigen binding site of HLA-B*27:05 molecule. Sequence alignment of ligands identified with species of bacteria associated with HLA-B*27-dependent reactive arthritis was performed. In the absence of ERAP2, peptides with N-terminal basic residues, and minority canonical P2 residues are enriched in the natural ligandome. Further, alterations of residue frequencies and hydrophobicity profile at P3, P7, and P positions were detected. In addition, several ERAP2-dependent cellular peptides were highly similar to protein sequences of arthritogenic bacteria, including one human HLA-B*27:05 ligand fully conserved in a protein from Campylobacter jejuni. These findings highlight the pathogenic role of this aminopeptidase in the triggering of AS autoimmune disease.

Accumulation and propagation of hyperphosphorylated tau (p-tau) is a common neuropathological hallmark associated with neurodegeneration of Alzheimer's disease (AD), frontotemporal dementia and parkinsonism linked to chromosome 17 (FTDP-17), and related tauopathies. Extracellular vesicles, specifically exosomes, have recently been demonstrated to participate in mediating tau propagation in brain. Exosomes produced by human induced pluripotent stem cell (iPSC)-derived neurons expressing mutant Tau (mTau), containing the P301L and V337M Tau mutations of FTDP-17, possess the ability to propagate p-tau pathology after injection into mouse brain.  To gain an understanding of the mTau exosome cargo involved in tau pathogenesis, these pathogenic exosomes were analyzed by proteomics and bioinformatics. The data showed that mTau expression dysregulates the exosome proteome to result in (1) proteins uniquely present only in mTau, and not control exosomes, (2) the absence of proteins in mTau exosomes, uniquely present in control exosomes, and (3) shared proteins which were significantly up-regulated or down-regulated in mTau compared to control exosomes. Notably, mTau exosomes (not control exosomes) contain ANP32A (also known as I1PP2A), an endogenous inhibitor of the PP2A phosphatase which regulates the phosphorylation state of p-tau.  Several of the mTau exosome-specific proteins have been shown to participate in AD mechanisms involving lysosomes, inflammation, secretases, and related processes.  Furthermore, the mTau exosomes lacked a substantial portion of proteins present in control exosomes involved in pathways of localization, vesicle transport, and protein binding functions. The shared proteins present in both mTau and control exosomes represented exosome functions of vesicle-mediated transport, exocytosis, and secretion processes. These data illustrate mTau as a dynamic regulator of the biogenesis of exosomes to result in acquisition, deletion, and up- or down-regulation of protein cargo to result in pathogenic mTau exosomes capable of in vivo propagation of p-tau neuropathology in mouse brain. 

Assembly factors play a critical role in the biogenesis of mitochondrial respiratory chain complexes I-IV where they assist in the membrane insertion of subunits, attachment of co-factors, and stabilization of assembly intermediates. The major fraction of complexes I, III and IV are present together in large molecular structures known as respiratory chain supercomplexes. A number of assembly factors have been proposed as required for supercomplex assembly, including the hypoxia inducible gene 1 domain family member HIGD2A. Using gene-edited human cell lines and extensive steady state, translation and affinity enrichment proteomics techniques we show that loss of HIGD2A leads to defects in the de novo biogenesis of mtDNA-encoded COX3, subsequent accumulation of complex IV intermediates and turnover of COX3 partner proteins. Deletion of HIGD2A also leads to defective complex IV activity. The impact of HIGD2A loss on complex IV was not altered by growth under hypoxic conditions, consistent with its role being in basal complex IV assembly. While in the absence of HIGD2A we show that mitochondria do contain an altered supercomplex assembly, we demonstrate it to harbor a crippled complex IV lacking COX3. Our results redefine HIGD2A as a classical assembly factor required for building the COX3 module of complex IV.

Perlecan is a critical proteoglycan found in the extracellular matrix (ECM) of cartilage. In healthy cartilage, perlecan regulates cartilage biomechanics and we previously demonstrated perlecan deficiency leads to reduced cellular and ECM stiffness in vivo. This change in mechanics may lead to the early onset osteoarthritis seen in disorders resulting from perlecan knockdown such as Schwartz-Jampel syndrome (SJS). To identify how perlecan knockdown affects the material properties of developing cartilage, we used imaging and liquid chromatography–tandem mass spectrometry (LC-MS/MS) to study the ECM in a murine model of SJS, Hspg2C1532Y-Neo. Perlecan knockdown led to defective pericellular matrix formation, whereas the abundance of bulk ECM proteins, including many collagens, increased. Post-translational modifications and ultrastructure of collagens were not significantly different; however, LC-MS/MS analysis showed more protein was secreted by Hspg2C1532Y-Neo cartilage in vitro, suggesting that the incorporation of newly synthesized ECM was impaired. In addition, glycosaminoglycan deposition was atypical, which may explain the previously observed decrease in mechanics. Overall, these findings provide insight into the influence of perlecan on functional cartilage assembly and the progression of osteoarthritis in SJS.

1 April 2020

Considerations of justice and social equity are as important for the circular economy transition as they are in the contexts of low-carbon transitions and digitalization of the economy. This paper sets out the just transition approach, and its relevance in climate change and energy transition debates.

1 May 2008 , Number 3

Rowdy sessions of the Iraq High Tribunal attracted sensational daily news coverage while Saddam Hussein was being tried. However, following his grim execution in December 2006, coverage all but evaporated. The foreign press and most western monitors packed their bags and left, and television reporting in Iraq dwindled. Now even the United States Department of Justice, which initially provided key financial and political support, is quietly withdrawing its advisers. So what is happening at the Tribunal and why are the Americans pulling out?

1 October 2008 , Number 4

Russia’s military action in Georgia was driven by Moscow’s domestic agenda. Foreign policy has become a tool of collective control back home. While the ‘ruling tandem’ has gained popularity, the new consensus does not provide for modernisation. Will the predatory state reform or collapse?

20 April 2020

There is and will continue to be an edge of rivalry in Egypt’s relations with the dominant Gulf Arab powers. This paper will focus primarily on the Egypt–Gulf relationship during the Sisi era.

1 March 2007 , Number 1

The Iranian nuclear site at Natanz is already one of the most protected places on earth, and the apparently never-ending diplomatic minuet over Iran’s nuclear activities may be no more than Tehran stringing out negotiations until its nuclear protection programme is complete.

7 May 2020 , Volume 96, Number 3

7 May 2020 , Volume 96, Number 3

7 May 2020 , Volume 96, Number 3

4 February 2020

14 February 2020

Invitation Only Research Event

Event participants

Ryhor Astapenia, Robert Bosch Stiftung Academy Fellow, Russia and Eurasia Programme, Chatham House
Chair: James Nixey, Programme Director, Russia and Eurasia, Chatham House

Nonalcoholic fatty liver disease (NAFLD) and nonalcoholic steatohepatitis (NASH) are emerging as leading causes of liver disease worldwide and have been recognized as one of the major unmet medical needs of the 21st century. Our recent translational studies in mouse models, human cell lines, and well-characterized patient cohorts have identified serine/threonine kinase (STK)25 as a protein that coats intrahepatocellular lipid droplets (LDs) and critically regulates liver lipid homeostasis and progression of NAFLD/NASH. Here, we studied the mechanism-of-action of STK25 in steatotic liver by relative quantification of the hepatic LD-associated phosphoproteome from high-fat diet-fed Stk25 knockout mice compared with their wild-type littermates. We observed a total of 131 proteins and 60 phosphoproteins that were differentially represented in STK25-deficient livers. Most notably, a number of proteins involved in peroxisomal function, ubiquitination-mediated proteolysis, and antioxidant defense were coordinately regulated in Stk25^–/– versus wild-type livers. We confirmed attenuated peroxisomal biogenesis and protection against oxidative and ER stress in STK25-deficient human liver cells, demonstrating the hepatocyte-autonomous manner of STK25’s action. In summary, our results suggest that regulation of peroxisomal function and metabolic stress response may be important molecular mechanisms by which STK25 controls the development and progression of NAFLD/NASH.

GPR120 is implicated as a lipid receptor in the oro-sensory detection of dietary fatty acids. However, the effects of GPR120 activation on dietary fat intake or obesity are not clearly understood. We investigated to determine whether the binding of TUG891, a novel GPR120 agonist, to lingual GPR120 modulates fat preference in mice. We explored the effects of TUG891 on obesity-related hormones and conducted behavioral choice tests on mice to better understand the physiologic relevance of the action of TUG891. In cultured mouse and human taste bud cells (TBCs), TUG891 induced a rapid increase in Ca²⁺ by acting on GPR120. A long-chain dietary fatty acid, linoleic acid (LA), also recruited Ca²⁺ via GPR120 in human and mouse TBCs. Both TUG891 and LA induced ERK1/2 phosphorylation and enhanced in vitro release of glucagon-like peptide-1 from cultured human and mouse TBCs. In situ application of TUG891 onto the tongue of anesthetized mice triggered the secretion of pancreatobiliary juice, probably via the tongue-brain-gut axis. Furthermore, lingual application of TUG891 altered circulating concentrations of cholecystokinin and adipokines, associated with decreased circulating LDL, in conscious mice. In behavioral tests, mice exhibited a spontaneous preference for solutions containing either TUG891 or LA instead of a control. However, addition of TUG891 to a solution containing LA significantly curtailed fatty acid preference. Our study demonstrates that TUG891 binds to lingual GPR120 receptors, activates the tongue-brain-gut axis, and modulates fat preference. These findings may support the development of new fat taste analogs that can change the approach to obesity prevention and treatment.

Plasma lipoprotein (a) [Lp(a)] levels are largely determined by variation in the LPA gene, which codes for apo(a). Genome-wide association studies (GWASs) have identified nonsynonymous variants in LPA that associate with low Lp(a) levels, although their effect on apo(a) function is unknown. We investigated two such variants, R990Q and R1771C, which were present in four null Lp(a) individuals, for structural and functional effects. Sequence alignments showed the R990 and R1771 residues to be highly conserved and homologous to each other and to residues associated with plasminogen deficiency. Structural modeling showed both residues to make several polar contacts with neighboring residues that would be ablated on substitution. Recombinant expression of the WT and R1771C apo(a) in liver and kidney cells showed an abundance of an immature form for both apo(a) proteins. A mature form of apo(a) was only seen with the WT protein. Imaging of the recombinant apo(a) proteins in conjunction with markers of the secretory pathway indicated a poor transit of R1771C into the Golgi. Furthermore, the R1771C mutant displayed a glycosylation pattern consistent with ER, but not Golgi, glycosylation. We conclude that R1771 and the equivalent R990 residue facilitate correct folding of the apo(a) kringle structure and mutations at these positions prevent the proper folding required for full maturation and secretion. To our knowledge, this is the first example of nonsynonymous variants in LPA being causative of a null Lp(a) phenotype.

Cholesteryl ester transfer protein (CETP) exists as full-length (FL) and exon 9 (E9)-deleted isoforms. The function of E9-deleted CETP is poorly understood. Here, we investigated the role of E9-deleted CETP in regulating the secretion of FL-CETP by cells and explored its possible role in intracellular lipid metabolism. CETP overexpression in cells that naturally express CETP confirmed that E9-deleted CETP is not secreted, and showed that cellular FL- and E9-deleted CETP form an isolatable complex. Coexpression of CETP isoforms lowered cellular levels of both proteins and impaired FL-CETP secretion. These effects were due to reduced synthesis of both isoforms; however, the predominate consequence of FL- and E9-deleted CETP coexpression is impaired FL-CETP synthesis. We reported previously that reducing both CETP isoforms or overexpressing FL-CETP impairs cellular triglyceride (TG) storage. To investigate this further, E9-deleted CETP was expressed in SW872 cells that naturally synthesize CETP and in mouse 3T3-L1 cells that do not. E9-deleted CETP overexpression stimulated SW872 triglyceride synthesis and increased stored TG 2-fold. Expression of E9-deleted CETP in mouse 3T3-L1 cells produced a similar lipid phenotype. In vitro, FL-CETP promotes the transfer of TG from ER-enriched membranes to lipid droplets. E9-deleted CETP also promoted this transfer, although less effectively, and it inhibited the transfer driven by FL-CETP. We conclude that FL- and E9-deleted CETP isoforms interact to mutually decrease their intracellular levels and impair FL-CETP secretion by reducing CETP biosynthesis. E9-deleted CETP, like FL-CETP, alters cellular TG metabolism and storage but in a contrary manner.

Zinc metallopeptidase STE24 (ZMPSTE24) is essential for the conversion of farnesyl–prelamin A to mature lamin A, a key component of the nuclear lamina. In the absence of ZMPSTE24, farnesyl–prelamin A accumulates in the nucleus and exerts toxicity, causing a variety of disease phenotypes. By ~4 months of age, both male and female Zmpste24^–/– mice manifest a near-complete loss of adipose tissue, but it has never been clear whether this phenotype is a direct consequence of farnesyl–prelamin A toxicity in adipocytes. To address this question, we generated a conditional knockout Zmpste24 allele and used it to create adipocyte-specific Zmpste24–knockout mice. To boost farnesyl–prelamin A levels, we bred in the "prelamin A–only" Lmna allele. Gene expression, immunoblotting, and immunohistochemistry experiments revealed that adipose tissue in these mice had decreased Zmpste24 expression along with strikingly increased accumulation of prelamin A. In male mice, Zmpste24 deficiency in adipocytes was accompanied by modest changes in adipose stores (an 11% decrease in body weight, a 23% decrease in body fat mass, and significantly smaller gonadal and inguinal white adipose depots). No changes in adipose stores were detected in female mice, likely because prelamin A expression in adipose tissue is lower in female mice. Zmpste24 deficiency in adipocytes did not alter the number of macrophages in adipose tissue, nor did it alter plasma levels of glucose, triglycerides, or fatty acids. We conclude that ZMPSTE24 deficiency in adipocytes, and the accompanying accumulation of farnesyl–prelamin A, reduces adipose tissue stores, but only modestly and only in male mice.

Obesity, dyslipidemia, and insulin resistance, the increasingly common metabolic syndrome, are risk factors for CVD and type 2 diabetes that warrant novel therapeutic interventions. The flavonoid nobiletin displays potent lipid-lowering and insulin-sensitizing properties in mice with metabolic dysfunction. However, the mechanisms by which nobiletin mediates metabolic protection are not clearly established. The central role of AMP-activated protein kinase (AMPK) as an energy sensor suggests that AMPK is a target of nobiletin. We tested the hypothesis that metabolic protection by nobiletin required phosphorylation of AMPK and acetyl-CoA carboxylase (ACC) in mouse hepatocytes, in mice deficient in hepatic AMPK (Ampkβ1^–/–), in mice incapable of inhibitory phosphorylation of ACC (AccDKI), and in mice with adipocyte-specific AMPK deficiency (iβ1β2AKO). We fed mice a high-fat/high-cholesterol diet with or without nobiletin. Nobiletin increased phosphorylation of AMPK and ACC in primary mouse hepatocytes, which was associated with increased FA oxidation and attenuated FA synthesis. Despite loss of ACC phosphorylation in Ampkβ1^–/– hepatocytes, nobiletin suppressed FA synthesis and enhanced FA oxidation. Acute injection of nobiletin into mice did not increase phosphorylation of either AMPK or ACC in liver. In mice fed a high-fat diet, nobiletin robustly prevented obesity, hepatic steatosis, dyslipidemia, and insulin resistance, and it improved energy expenditure in Ampkβ1^–/–, AccDKI, and iβ1β2AKO mice to the same extent as in WT controls. Thus, the beneficial metabolic effects of nobiletin in vivo are conferred independently of hepatic or adipocyte AMPK activation. These studies further underscore the therapeutic potential of nobiletin and begin to clarify possible mechanisms.

The composition-function relationship of HDL particles and its effects on the mechanisms driving coronary heart disease (CHD) is poorly understood. We tested the hypothesis that the functionality of HDL particles is significantly influenced by their lipid composition. Using a novel 3D-separation method, we isolated five different-sized HDL subpopulations from CHD patients who had low preβ-1 functionality (low-F) (ABCA1-dependent cholesterol-efflux normalized for preβ-1 concentration) and controls who had either low-F or high preβ-1 functionality (high-F). Molecular numbers of apoA-I, apoA-II, and eight major lipid classes were determined in each subpopulation by LC-MS. The average number of lipid molecules decreased from 422 in the large spherical α-1 particles to 57 in the small discoid preβ-1 particles. With decreasing particle size, the relative concentration of free cholesterol (FC) decreased in α-mobility but not in preβ-1 particles. Preβ-1 particles contained more lipids than predicted; 30% of which were neutral lipids (cholesteryl ester and triglyceride), indicating that these particles were mainly remodeled from larger particles not newly synthesized. There were significant correlations between HDL-particle functionality and the concentrations of several lipids. Unexpectedly, the phospholipid:FC ratio was significantly correlated with large-HDL-particle functionality but not with preβ-1 functionality. There was significant positive correlation between particle functionality and total lipids in high-F controls, indicating that the lipid-binding capacity of apoA-I plays a major role in the cholesterol efflux capacity of HDL particles. Functionality and lipid composition of HDL particles are significantly correlated and probably both are influenced by the lipid-binding capacity of apoA-I.

Bile acids (BAs) facilitate intestinal absorption of lipid-soluble nutrients and modulate various metabolic pathways through the farnesoid X receptor (FXR) and Takeda G-protein-coupled receptor 5. These receptors are targets for therapy in cholestatic and metabolic diseases. However, dissimilarities in BA metabolism between humans and mice complicate translation of preclinical data. Cytochrome P450 family 2 subfamily c polypeptide 70 (CYP2C70) was recently proposed to catalyze the formation of rodent-specific muricholic acids (MCAs). With CRISPR/Cas9-mediated somatic genome editing, we generated an acute hepatic Cyp2c70 knockout mouse model (Cyp2c70^ako) to clarify the role of CYP2C70 in BA metabolism in vivo and evaluate whether its activity modulates effects of pharmacologic FXR activation on cholesterol homeostasis. In Cyp2c70^ako mice, chenodeoxycholic acid (CDCA) increased at the expense of βMCA, resulting in a more hydrophobic human-like BA pool. Tracer studies demonstrated that, in vivo, CYP2C70 catalyzes the formation of βMCA primarily by sequential 6β-hydroxylation and C7-epimerization of CDCA, generating αMCA as an intermediate metabolite. Physiologically, the humanized BA composition in Cyp2c70^ako mice blunted the stimulation of fecal cholesterol disposal in response to FXR activation compared with WT mice, predominantly due to reduced stimulation of transintestinal cholesterol excretion. Thus, deletion of hepatic Cyp2c70 in adult mice translates into a human-like BA pool composition and impacts the response to pharmacologic FXR activation. This Cyp2c70^ako mouse model may be a useful tool for future studies of BA signaling and metabolism that informs human disease development and treatment.