The Babraham Institute Publications database contains details of all publications resulting from our research groups and scientific services.

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Title / Authors / Details Open Access Download

Cyt-Geist: Current and Future Challenges in Cytometry: Reports of the CYTO 2019 Conference Workshops.
Czechowska K, Lannigan J, Aghaeepour N, Back JB, Begum J, Behbehani G, Bispo C, Bitoun D, Fernández AB, Boova ST, Brinkman RR, Ciccolella CO, Cotleur B, Davies D, Dela Cruz GV, Del Rio-Guerra R, Des Lauriers-Cox AM, Douagi I, Dumrese C, Bonilla Escobar DL, Estevam J, Ewald C, Fossum A, Gaudillière B, Green C, Groves C, Hall C, Haque Y, Hedrick MN, Hogg K, Hsieh EWY, Irish J, Lederer J, Leipold M, Lewis-Tuffin LJ, Litwin V, Lopez P, Nasdala I, Nedbal J, Ohlsson-Wilhelm BM, Price KM, Rahman AH, Rayanki R, Rieger AM, Robinson JP, Shapiro H, Sun YS, Tang VA, Tesfa L, Telford WG, Walker R, Welsh JA, Wheeler P, Tárnok A

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Cytometry. Part A : the journal of the International Society for Analytical Cytology , 95 , 12 ,

PMID: 31833655

Mammalian phospholipase D: Function, and therapeutics.
McDermott MI, Wang Y, Wakelam MJO, Bankaitis VA

Despite being discovered over 60 years ago, the precise role of Phospholipase D (PLD) is still being elucidated. PLD enzymes catalyze the hydrolysis of the phosphodiester bond of glycerophospholipids producing phosphatidic acid and the free headgroup. PLD family members are found in organisms ranging from viruses, and bacteria to plants, and mammals, and they display a range of substrate specificities, are regulated by a diverse range of molecules, and have been implicated in a broad range of cellular processes including receptor signaling, cytoskeletal regulation and membrane trafficking. Recent technological advances including: the development of PLD knockout mice, isoform-specific antibodies, and specific inhibitors are finally permitting a thorough analysis of the in vivo role of mammalian PLDs. These studies are facilitating increased recognition of PLD's role in disease states including cancers and Alzheimer's disease, offering potential as a target for therapeutic intervention.

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Progress in lipid research , 0 , 0 ,

PMID: 31830503

Multi-omics profiling of mouse gastrulation at single-cell resolution.
Argelaguet R, Clark SJ, Mohammed H, Stapel LC, Krueger C, Kapourani CA, Imaz-Rosshandler I, Lohoff T, Xiang Y, Hanna CW, Smallwood S, Ibarra-Soria X, Buettner F, Sanguinetti G, Xie W, Krueger F, Göttgens B, Rugg-Gunn PJ, Kelsey G, Dean W, Nichols J, Stegle O, Marioni JC, Reik W

Formation of the three primary germ layers during gastrulation is an essential step in the establishment of the vertebrate body plan and is associated with major transcriptional changes. Global epigenetic reprogramming accompanies these changes, but the role of the epigenome in regulating early cell-fate choice remains unresolved, and the coordination between different molecular layers is unclear. Here we describe a single-cell multi-omics map of chromatin accessibility, DNA methylation and RNA expression during the onset of gastrulation in mouse embryos. The initial exit from pluripotency coincides with the establishment of a global repressive epigenetic landscape, followed by the emergence of lineage-specific epigenetic patterns during gastrulation. Notably, cells committed to mesoderm and endoderm undergo widespread coordinated epigenetic rearrangements at enhancer marks, driven by ten-eleven translocation (TET)-mediated demethylation and a concomitant increase of accessibility. By contrast, the methylation and accessibility landscape of ectodermal cells is already established in the early epiblast. Hence, regulatory elements associated with each germ layer are either epigenetically primed or remodelled before cell-fate decisions, providing the molecular framework for a hierarchical emergence of the primary germ layers.

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Nature , 576 , 7787 ,

PMID: 31827285

Open Access

Azathioprine Has a Deleterious Effect on the Bone Health of Mice with DSS-Induced Inflammatory Bowel Disease.
Morgan S, Hooper KM, Milne EM, Farquharson C, Stevens C, Staines KA

Patients with inflammatory bowel disease (IBD) often present poor bone health and are 40% more at risk of bone fracture. Studies have implicated autophagy in IBD pathology and drugs used to treat IBD stimulate autophagy in varying degrees, however, their effect on the skeleton is currently unknown. Here, we have utilised the dextran sulphate sodium (DSS) model of colitis in mice to examine the effects of the thiopurine drug azathioprine on the skeleton. Ten-week-old male mice ( = 6/group) received 3.0% DSS in their drinking water for four days, followed by a 14-day recovery period. Mice were treated with 10 mg/kg/day azathioprine or vehicle control. Histopathological analysis of the colon from DSS mice revealed significant increases in scores for inflammation severity, extent, and crypt damage ( < 0.05). Azathioprine provided partial protection to the colon, as reflected by a lack of significant difference in crypt damage and tissue regeneration with DSS treatment. MicroCT of vehicle-treated DSS mice revealed azathioprine treatment had a significant detrimental effect on the trabecular bone microarchitecture, independent of DSS treatment. Specifically, significant decreases were observed in bone volume/tissue volume ( < 0.01), and trabecular number ( < 0.05), with a concurrent significant increase in trabecular pattern factor ( < 0.01). Immunohistochemical labelling for LC3 revealed azathioprine to induce autophagy in the bone marrow. Together these data suggest that azathioprine treatment may have a deleterious effect on IBD patients who may already be at increased risk of osteoporotic bone fractures and thus will inform on future treatment strategies for patient stratification.

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International journal of molecular sciences , 20 , 23 ,

PMID: 31816823

Open Access

Transcription-induced formation of extrachromosomal DNA during yeast ageing.
Hull RM, King M, Pizza G, Krueger F, Vergara X, Houseley J

Extrachromosomal circular DNA (eccDNA) facilitates adaptive evolution by allowing rapid and extensive gene copy number variation and is implicated in the pathology of cancer and ageing. Here, we demonstrate that yeast aged under environmental copper accumulate high levels of eccDNA containing the copper-resistance gene CUP1. Transcription of the tandemly repeated CUP1 gene causes CUP1 eccDNA accumulation, which occurs in the absence of phenotypic selection. We have developed a sensitive and quantitative eccDNA sequencing pipeline that reveals CUP1 eccDNA accumulation on copper exposure to be exquisitely site specific, with no other detectable changes across the eccDNA complement. eccDNA forms de novo from the CUP1 locus through processing of DNA double-strand breaks (DSBs) by Sae2, Mre11 and Mus81, and genome-wide analyses show that other protein coding eccDNA species in aged yeast share a similar biogenesis pathway. Although abundant, we find that CUP1 eccDNA does not replicate efficiently, and high-copy numbers in aged cells arise through frequent formation events combined with asymmetric DNA segregation. The transcriptional stimulation of CUP1 eccDNA formation shows that age-linked genetic change varies with transcription pattern, resulting in gene copy number profiles tailored by environment.

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PLoS biology , 17 , 12 ,

PMID: 31794573

Open Access

The application of cell surface markers to demarcate distinct human pluripotent states.
Goodwin J, Laslett AL, Rugg-Gunn PJ

Recent advances in human pluripotent stem cell (hPSC) research have uncovered different subpopulations within stem cell cultures and have captured a range of pluripotent states that hold distinct molecular and functional properties. At the two ends of the pluripotency spectrum are naïve and primed hPSC, whereby naïve hPSC grown in stringent conditions recapitulate features of the preimplantation human embryo, and the conventionally grown primed hPSC align closer to the early postimplantation embryo. Investigating these cell types will help to define the mechanisms that control early development and should provide new insights into stem cell properties such as cell identity, differentiation and reprogramming. Monitoring cell surface marker expression provides a valuable approach to resolve complex cell populations, to directly compare between cell types, and to isolate viable cells for functional experiments. This review discusses the discovery and applications of cell surface markers to study human pluripotent cell types with a particular focus on the transitions between naïve and primed states. Highlighted areas for future study include the potential functions for the identified cell surface proteins in pluripotency, the production of new high-quality monoclonal antibodies to naïve-specific protein epitopes and the use of cell surface markers to characterise subpopulations within pluripotent states.

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Experimental cell research , 0 , 0 ,

PMID: 31790696

Open Access

The role and mechanisms of DNA methylation in the oocyte.
Sendžikaitė G, Kelsey G

Epigenetic information in the mammalian oocyte has the potential to be transmitted to the next generation and influence gene expression; this occurs naturally in the case of imprinted genes. Therefore, it is important to understand how epigenetic information is patterned during oocyte development and growth. Here, we review the current state of knowledge of de novo DNA methylation mechanisms in the oocyte: how a distinctive gene-body methylation pattern is created, and the extent to which the DNA methylation machinery reads chromatin states. Recent epigenomic studies building on advances in ultra-low input chromatin profiling methods, coupled with genetic studies, have started to allow a detailed interrogation of the interplay between DNA methylation establishment and chromatin states; however, a full mechanistic description awaits.

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Essays in biochemistry , 63 , 6 ,

PMID: 31782490

Open Access

Protocols for Northern Analysis of Exosome Substrates and Other Noncoding RNAs.
Cruz C, Houseley J

Over the past decade a plethora of noncoding RNAs (ncRNAs) have been identified, initiating an explosion in RNA research. Although RNA sequencing methods provide unsurpassed insights into ncRNA distribution and expression, detailed information on structure and processing are harder to extract from sequence data. In contrast, northern blotting methods provide uniquely detailed insights into complex RNA populations but are rarely employed outside specialist RNA research groups. Such techniques are generally considered difficult for nonspecialists, which is unfortunate as substantial technical advances in the past few decades have solved the major challenges. Here we present simple, reproducible and highly robust protocols for separating glyoxylated RNA on agarose gels and heat denatured RNA on polyacrylamide-urea gels using standard laboratory electrophoresis equipment. We also provide reliable transfer and hybridization protocols that do not require optimization for most applications. Together, these should allow any molecular biology lab to elucidate the structure and processing of ncRNAs of interest.

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Methods in molecular biology (Clifton, N.J.) , 2062 , 1 ,

PMID: 31768973

Open Access

BNC1 regulates cell heterogeneity in human pluripotent stem cell-derived epicardium.
Gambardella L, McManus SA, Moignard V, Sebukhan D, Delaune A, Andrews S, Bernard WG, Morrison MA, Riley PR, Göttgens B, Gambardella Le Novère N, Sinha S

The murine developing epicardium heterogeneously expresses the transcription factors TCF21 and WT1. Here, we show that this cell heterogeneity is conserved in human epicardium, regulated by BNC1 and associated with cell fate and function. Single cell RNA sequencing of epicardium derived from human pluripotent stem cells (hPSC-epi) revealed that distinct epicardial subpopulations are defined by high levels of expression for the transcription factors BNC1 or TCF21. WT1 cells are included in the BNC1 population, which was confirmed in human foetal hearts. THY1 emerged as a membrane marker of the TCF21 population. We show that THY1 cells can differentiate into cardiac fibroblasts (CFs) and smooth muscle cells (SMCs), whereas THY1 cells were predominantly restricted to SMCs. Knocking down BNC1 during the establishment of the epicardial populations resulted in a homogeneous, predominantly TCF21 population. Network inference methods using transcriptomic data from the different cell lineages derived from the hPSC-epi delivered a core transcriptional network organised around WT1, TCF21 and BNC1. This study unveils a list of epicardial regulators and is a step towards engineering subpopulations of epicardial cells with selective biological activities.

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Development (Cambridge, England) , 146 , 24 ,

PMID: 31767620

Open Access

DNA methylation aging clocks: challenges and recommendations.
Bell CG, Lowe R, Adams PD, Baccarelli AA, Beck S, Bell JT, Christensen BC, Gladyshev VN, Heijmans BT, Horvath S, Ideker T, Issa JJ, Kelsey KT, Marioni RE, Reik W, Relton CL, Schalkwyk LC, Teschendorff AE, Wagner W, Zhang K, Rakyan VK

Epigenetic clocks comprise a set of CpG sites whose DNA methylation levels measure subject age. These clocks are acknowledged as a highly accurate molecular correlate of chronological age in humans and other vertebrates. Also, extensive research is aimed at their potential to quantify biological aging rates and test longevity or rejuvenating interventions. Here, we discuss key challenges to understand clock mechanisms and biomarker utility. This requires dissecting the drivers and regulators of age-related changes in single-cell, tissue- and disease-specific models, as well as exploring other epigenomic marks, longitudinal and diverse population studies, and non-human models. We also highlight important ethical issues in forensic age determination and predicting the trajectory of biological aging in an individual.

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Genome biology , 20 , 1 ,

PMID: 31767039

Open Access

Dual-mechanism ERK1/2 inhibitors exploit a distinct binding mode to block phosphorylation and nuclear accumulation of ERK1/2.
Kidger AM, Munck JM, Saini HK, Balmanno K, Minihane E, Courtin A, Graham B, O'Reilly M, Odle R, Cook SJ

The RAS-regulated RAF-MEK1/2-ERK1/2 signalling pathway is frequently deregulated in cancer due to activating mutations of growth factor receptors, RAS or BRAF. Both RAF and MEK1/2 inhibitors are clinically approved and various ERK1/2 inhibitors (ERKi) are currently undergoing clinical trials. To date ERKi display two distinct mechanisms of action (MoA); catalytic ERKi solely inhibit ERK1/2 catalytic activity, whereas dual mechanism ERKi additionally prevent the activating phosphorylation of ERK1/2 at its T-E-Y motif by MEK1/2. These differences may impart significant differences in biological activity because T-E-Y phosphorylation is the signal for nuclear entry of ERK1/2, allowing them to access many key transcription factor targets. Here, we characterised the MoA of five ERKi and examined their functional consequences in terms of ERK1/2 signalling, gene expression and anti-proliferative efficacy. We demonstrate that catalytic ERKi promote a striking nuclear accumulation of p-ERK1/2 in KRAS mutant cell lines. In contrast, dual mechanism ERKi exploit a distinct binding mode to block ERK1/2 phosphorylation by MEK1/2, exhibit superior potency and prevent the nuclear accumulation of ERK1/2. Consequently, dual-mechanism ERKi exhibit more durable pathway inhibition and enhanced suppression of ERK1/2-dependent gene expression compared to catalytic ERKi, resulting in increased efficacy across BRAF and RAS mutant cell lines.

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Molecular cancer therapeutics , 1 , 1 ,

PMID: 31748345

A nutritional memory effect counteracts benefits of dietary restriction in old mice.
Hahn O, Drews LF, Nguyen A, Tatsuta T, Gkioni L, Hendrich O, Zhang Q, Langer T, Pletcher S, Wakelam MJO, Beyer A, Grönke S, Partridge L

Dietary restriction (DR) during adulthood can greatly extend lifespan and improve metabolic health in diverse species. However, whether DR in mammals is still effective when applied for the first time at old age remains elusive. Here, we report results of a late-life DR switch experiment employing 800 mice, in which 24 months old female mice were switched from ad libitum (AL) to DR or vice versa. Strikingly, the switch from DR-to-AL acutely increases mortality, whereas the switch from AL-to-DR causes only a weak and gradual increase in survival, suggesting a memory of earlier nutrition. RNA-seq profiling in liver, brown (BAT) and white adipose tissue (WAT) demonstrate a largely refractory transcriptional and metabolic response to DR after AL feeding in fat tissue, particularly in WAT, and a proinflammatory signature in aged preadipocytes, which is prevented by chronic DR feeding. Our results provide evidence for a nutritional memory as a limiting factor for DR-induced longevity and metabolic remodeling of WAT in mammals.

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Nature metabolism , 1 , 11 ,

PMID: 31742247

Mitochondrial impairment activates the Wallerian pathway through depletion of NMNAT2 leading to SARM1-dependent axon degeneration.
Loreto A, Hill CS, Hewitt VL, Orsomando G, Angeletti C, Gilley J, Lucci C, Sanchez-Martinez A, Whitworth AJ, Conforti L, Dajas-Bailador F, Coleman MP

Wallerian degeneration of physically injured axons involves a well-defined molecular pathway linking loss of axonal survival factor NMNAT2 to activation of pro-degenerative protein SARM1. Manipulating the pathway through these proteins led to the identification of non-axotomy insults causing axon degeneration by a Wallerian-like mechanism, including several involving mitochondrial impairment. Mitochondrial dysfunction is heavily implicated in Parkinson's disease, Charcot-Marie-Tooth disease, hereditary spastic paraplegia and other axonal disorders. However, whether and how mitochondrial impairment activates Wallerian degeneration has remained unclear. Here, we show that disruption of mitochondrial membrane potential leads to axonal NMNAT2 depletion in mouse sympathetic neurons, increasing the substrate-to-product ratio (NMN/NAD) of this NAD-synthesising enzyme, a metabolic fingerprint of Wallerian degeneration. The mechanism appears to involve both impaired NMNAT2 synthesis and reduced axonal transport. Expression of WLD and Sarm1 deletion both protect axons after mitochondrial uncoupling. Blocking the pathway also confers neuroprotection and increases the lifespan of flies with Pink1 loss-of-function mutation, which causes severe mitochondrial defects. These data indicate that mitochondrial impairment replicates all the major steps of Wallerian degeneration, placing it upstream of NMNAT2 loss, with the potential to contribute to axon pathology in mitochondrial disorders.

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Neurobiology of disease , 134 , 1 ,

PMID: 31740269

An mTORC1-to-CDK1 Switch Maintains Autophagy Suppression during Mitosis.
Odle RI, Walker SA, Oxley D, Kidger AM, Balmanno K, Gilley R, Okkenhaug H, Florey O, Ktistakis NT, Cook SJ

Since nuclear envelope breakdown occurs during mitosis in metazoan cells, it has been proposed that macroautophagy must be inhibited to maintain genome integrity. However, repression of macroautophagy during mitosis remains controversial and mechanistic detail limited to the suggestion that CDK1 phosphorylates VPS34. Here, we show that initiation of macroautophagy, measured by the translocation of the ULK complex to autophagic puncta, is repressed during mitosis, even when mTORC1 is inhibited. Indeed, mTORC1 is inactive during mitosis, reflecting its failure to localize to lysosomes due to CDK1-dependent RAPTOR phosphorylation. While mTORC1 normally represses autophagy via phosphorylation of ULK1, ATG13, ATG14, and TFEB, we show that the mitotic phosphorylation of these autophagy regulators, including at known repressive sites, is dependent on CDK1 but independent of mTOR. Thus, CDK1 substitutes for inhibited mTORC1 as the master regulator of macroautophagy during mitosis, uncoupling autophagy regulation from nutrient status to ensure repression of macroautophagy during mitosis.

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Molecular cell , S1097-2765 , 19 ,

PMID: 31733992

Open Access

Targeting melanoma's MCL1 bias unleashes the apoptotic potential of BRAF and ERK1/2 pathway inhibitors.
Sale MJ, Minihane E, Monks NR, Gilley R, Richards FM, Schifferli KP, Andersen CL, Davies EJ, Vicente MA, Ozono E, Markovets A, Dry JR, Drew L, Flemington V, Proia T, Jodrell DI, Smith PD, Cook SJ

BRAF and MEK1/2 inhibitors are effective in melanoma but resistance inevitably develops. Despite increasing the abundance of pro-apoptotic BIM and BMF, ERK1/2 pathway inhibition is predominantly cytostatic, reflecting residual pro-survival BCL2 family activity. Here, we show that uniquely low BCL-X expression in melanoma biases the pro-survival pool towards MCL1. Consequently, BRAF or MEK1/2 inhibitors are synthetic lethal with the MCL1 inhibitor AZD5991, driving profound tumour cell death that requires BAK/BAX, BIM and BMF, and inhibiting tumour growth in vivo. Combination of ERK1/2 pathway inhibitors with BCL2/BCL-w/BCL-X inhibitors is stronger in CRC, correlating with a low MCL1:BCL-X ratio; indeed the MCL1:BCL-X ratio is predictive of ERK1/2 pathway inhibitor synergy with MCL1 or BCL2/BCL-w/BCL-X inhibitors. Finally, AZD5991 delays acquired BRAFi/MEKi resistance and enhances the efficacy of an ERK1/2 inhibitor in a model of acquired BRAFi + MEKi resistance. Thus combining ERK1/2 pathway inhibitors with MCL1 antagonists in melanoma could improve therapeutic index and patient outcomes.

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Nature communications , 10 , 1 ,

PMID: 31727888

Open Access

Autophagosome biogenesis machinery.
Walker SA, Ktistakis NT

We review current knowledge of the process of autophagosome formation with special emphasis on the very early steps: turning on the autophagy pathway, assembling the autophagy machinery, and building the autophagosome. The pathway is remarkably well co-ordinated spatially and temporally, and it shows broad conservation across species and cell types, including neurons. In addition, although much current knowledge derives mostly from settings of non-selective autophagy, recent work also indicates that selective autophagy, and more specifically mitophagy, shows similar dynamics. Having an understanding of this remarkable process may help the design of novel therapeutics for neurodegeneration and other pathologies.

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Journal of molecular biology , , 1089-8638 , 2019

PMID: 31705882

BioModels-15 years of sharing computational models in life science.
Malik-Sheriff RS, Glont M, Nguyen TVN, Tiwari K, Roberts MG, Xavier A, Vu MT, Men J, Maire M, Kananathan S, Fairbanks EL, Meyer JP, Arankalle C, Varusai TM, Knight-Schrijver V, Li L, Dueñas-Roca C, Dass G, Keating SM, Park YM, Buso N, Rodriguez N, Hucka M, Hermjakob H

Computational modelling has become increasingly common in life science research. To provide a platform to support universal sharing, easy accessibility and model reproducibility, BioModels (, a repository for mathematical models, was established in 2005. The current BioModels platform allows submission of models encoded in diverse modelling formats, including SBML, CellML, PharmML, COMBINE archive, MATLAB, Mathematica, R, Python or C++. The models submitted to BioModels are curated to verify the computational representation of the biological process and the reproducibility of the simulation results in the reference publication. The curation also involves encoding models in standard formats and annotation with controlled vocabularies following MIRIAM (minimal information required in the annotation of biochemical models) guidelines. BioModels now accepts large-scale submission of auto-generated computational models. With gradual growth in content over 15 years, BioModels currently hosts about 2000 models from the published literature. With about 800 curated models, BioModels has become the world's largest repository of curated models and emerged as the third most used data resource after PubMed and Google Scholar among the scientists who use modelling in their research. Thus, BioModels benefits modellers by providing access to reliable and semantically enriched curated models in standard formats that are easy to share, reproduce and reuse.

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Nucleic acids research , 1 , 1362-4962 , 2019

PMID: 31701150

Open Access

Myeloid Tribbles 1 induces early atherosclerosis via enhanced foam cell expansion.
Johnston JM, Angyal A, Bauer RC, Hamby S, Suvarna SK, Baidžajevas K, Hegedus Z, Dear TN, Turner M, , Wilson HL, Goodall AH, Rader DJ, Shoulders CC, Francis SE, Kiss-Toth E

Macrophages drive atherosclerotic plaque progression and rupture; hence, attenuating their atherosclerosis-inducing properties holds promise for reducing coronary heart disease (CHD). Recent studies in mouse models have demonstrated that Tribbles 1 (Trib1) regulates macrophage phenotype and shows that deficiency increases plasma cholesterol and triglyceride levels, suggesting that reduced expression mediates the strong genetic association between the locus and increased CHD risk in man. However, we report here that myeloid-specific (m) deficiency reduces early atheroma formation and that m transgene expression increases atherogenesis. Mechanistically, m increased macrophage lipid accumulation and the expression of a critical receptor (OLR1), promoting oxidized low-density lipoprotein uptake and the formation of lipid-laden foam cells. As and RNA levels were also strongly correlated in human macrophages, we suggest that a conserved, TRIB1-mediated mechanism drives foam cell formation in atherosclerotic plaque and that inhibiting mTRIB1 could be used therapeutically to reduce CHD.

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Science advances , 5 , 2375-2548 , 2019

PMID: 31692955

Open Access

Endogenous retroviral insertions drive non-canonical imprinting in extra-embryonic tissues.
Hanna CW, Pérez-Palacios R, Gahurova L, Schubert M, Krueger F, Biggins L, Andrews S, Colomé-Tatché M, Bourc'his D, Dean W, Kelsey G

Genomic imprinting is an epigenetic phenomenon that allows a subset of genes to be expressed mono-allelically based on the parent of origin and is typically regulated by differential DNA methylation inherited from gametes. Imprinting is pervasive in murine extra-embryonic lineages, and uniquely, the imprinting of several genes has been found to be conferred non-canonically through maternally inherited repressive histone modification H3K27me3. However, the underlying regulatory mechanisms of non-canonical imprinting in post-implantation development remain unexplored.

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Genome biology , 20 , 1474-760X , 2019

PMID: 31665063

Open Access

Sarm1 deletion suppresses TDP-43-linked motor neuron degeneration and cortical spine loss.
White MA, Lin Z, Kim E, Henstridge CM, Pena Altamira E, Hunt CK, Burchill E, Callaghan I, Loreto A, Brown-Wright H, Mead R, Simmons C, Cash D, Coleman MP, Sreedharan J

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative condition that primarily affects the motor system and shares many features with frontotemporal dementia (FTD). Evidence suggests that ALS is a 'dying-back' disease, with peripheral denervation and axonal degeneration occurring before loss of motor neuron cell bodies. Distal to a nerve injury, a similar pattern of axonal degeneration can be seen, which is mediated by an active axon destruction mechanism called Wallerian degeneration. Sterile alpha and TIR motif-containing 1 (Sarm1) is a key gene in the Wallerian pathway and its deletion provides long-term protection against both Wallerian degeneration and Wallerian-like, non-injury induced axonopathy, a retrograde degenerative process that occurs in many neurodegenerative diseases where axonal transport is impaired. Here, we explored whether Sarm1 signalling could be a therapeutic target for ALS by deleting Sarm1 from a mouse model of ALS-FTD, a TDP-43, YFP-H double transgenic mouse. Sarm1 deletion attenuated motor axon degeneration and neuromuscular junction denervation. Motor neuron cell bodies were also significantly protected. Deletion of Sarm1 also attenuated loss of layer V pyramidal neuronal dendritic spines in the primary motor cortex. Structural MRI identified the entorhinal cortex as the most significantly atrophic region, and histological studies confirmed a greater loss of neurons in the entorhinal cortex than in the motor cortex, suggesting a prominent FTD-like pattern of neurodegeneration in this transgenic mouse model. Despite the reduction in neuronal degeneration, Sarm1 deletion did not attenuate age-related behavioural deficits caused by TDP-43. However, Sarm1 deletion was associated with a significant increase in the viability of male TDP-43 mice, suggesting a detrimental role of Wallerian-like pathways in the earliest stages of TDP-43-mediated neurodegeneration. Collectively, these results indicate that anti-SARM1 strategies have therapeutic potential in ALS-FTD.

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Acta neuropathologica communications , 7 , 2051-5960 , 2019

PMID: 31661035

Open Access

Stem-cell-derived human microglia transplanted in mouse brain to study human disease.
Mancuso R, Van Den Daele J, Fattorelli N, Wolfs L, Balusu S, Burton O, Liston A, Sierksma A, Fourne Y, Poovathingal S, Arranz-Mendiguren A, Sala Frigerio C, Claes C, Serneels L, Theys T, Perry VH, Verfaillie C, Fiers M, De Strooper B

Although genetics highlights the role of microglia in Alzheimer's disease, one-third of putative Alzheimer's disease risk genes lack adequate mouse orthologs. Here we successfully engraft human microglia derived from embryonic stem cells in the mouse brain. The cells recapitulate transcriptionally human primary microglia ex vivo and show expression of human-specific Alzheimer's disease risk genes. Oligomeric amyloid-β induces a divergent response in human versus mouse microglia. This model can be used to study the role of microglia in neurological diseases.

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Nature neuroscience , , 1546-1726 , 2019

PMID: 31659342

How is the acyl chain composition of phosphoinositides created and does it matter?
Barneda D, Cosulich S, Stephens L, Hawkins P

The phosphoinositide (PIPn) family of signalling phospholipids are central regulators in membrane cell biology. Their varied functions are based on the phosphorylation pattern of their inositol ring, which can be recognized by selective binding domains in their effector proteins and be modified by a series of specific PIPn kinases and phosphatases, which control their interconversion in a spatial and temporal manner. Yet, a unique feature of PIPns remains largely unexplored: their unusually uniform acyl chain composition. Indeed, while most phospholipids present a range of molecular species comprising acyl chains of diverse length and saturation, PIPns in several organisms and tissues show the predominance of a single hydrophobic backbone, which in mammals is composed of arachidonoyl and stearoyl chains. Despite evolution having favoured this specific PIPn configuration, little is known regarding the mechanisms and functions behind it. In this review, we explore the metabolic pathways that could control the acyl chain composition of PIPns as well as the potential roles of this selective enrichment. While our understanding of this phenomenon has been constrained largely by the technical limitations in the methods traditionally employed in the PIPn field, we believe that the latest developments in PIPn analysis should shed light onto this old question.

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Biochemical Society transactions , 47 , 5 ,

PMID: 31657437

Open Access

Lipidomics: Current state of the art in a fast moving field.
O'Donnell VB, Ekroos K, Liebisch G, Wakelam M

Lipids are essential for all facets of life. They play three major roles: energy metabolism, structural, and signaling. They are dynamic molecules strongly influenced by endogenous and exogenous factors including genetics, diet, age, lifestyle, drugs, disease and inflammation. As precision medicine starts to become mainstream, there is a huge burgeoning interest in lipids and their potential to act as unique biomarkers or prognostic indicators. Lipids comprise a large component of all metabolites (around one-third), and our expanding knowledge about their dynamic behavior is fueling the hope that mapping their regulatory biochemical pathways on a systems level will revolutionize our ability to prevent, diagnose, and stratify major human diseases. Up to now, clinical lipid measurements have consisted primarily of total cholesterol or triglycerides, as a measure for cardiovascular risk and response to lipid lowering drugs. Nowadays, we are able to measure thousands of individual lipids that make up the lipidome. nuclear magnetic resonance spectrometry (NMR) metabolomics is also being increasingly used in large cohort studies where it can report on total levels of selected lipid classes, and relative levels of fatty acid saturation. To support the application of lipidomics research, LIPID MAPS was established in 2003, and since then has gone on to become the go-to resource for several lipid databases, lipid drawing tools, data deposition, and more recently lipidomics informatics tools, and a lipid biochemistry encyclopedia, LipidWeb. Alongside this, the recently established Lipidomics Standards Initiative plays a key role in standardization of lipidomics methodologies. This article is categorized under: Laboratory Methods and Technologies > Metabolomics Analytical and Computational Methods > Analytical Methods.

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Wiley interdisciplinary reviews. Systems biology and medicine , , 1939-005X , 2019

PMID: 31646749

Tet3 regulates cellular identity and DNA methylation in neural progenitor cells.
Santiago M, Antunes C, Guedes M, Iacovino M, Kyba M, Reik W, Sousa N, Pinto L, Branco MR, Marques CJ

TET enzymes oxidize 5-methylcytosine (5mC) into 5-hydroxymethylcytosine (5hmC), a process thought to be intermediary in an active DNA demethylation mechanism. Notably, 5hmC is highly abundant in the brain and in neuronal cells. Here, we interrogated the function of Tet3 in neural precursor cells (NPCs), using a stable and inducible knockdown system and an in vitro neural differentiation protocol. We show that Tet3 is upregulated during neural differentiation, whereas Tet1 is downregulated. Surprisingly, Tet3 knockdown led to a de-repression of pluripotency-associated genes such as Oct4, Nanog or Tcl1, with concomitant hypomethylation. Moreover, in Tet3 knockdown NPCs, we observed the appearance of OCT4-positive cells forming cellular aggregates, suggesting de-differentiation of the cells. Notably, Tet3 KD led to a genome-scale loss of DNA methylation and hypermethylation of a smaller number of CpGs that are located at neurogenesis-related genes and at imprinting control regions (ICRs) of Peg10, Zrsr1 and Mcts2 imprinted genes. Overall, our results suggest that TET3 is necessary to maintain silencing of pluripotency genes and consequently neural stem cell identity, possibly through regulation of DNA methylation levels in neural precursor cells.

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Cellular and molecular life sciences : CMLS , 1 , 1420-9071 , 2019

PMID: 31646359

Open Access