Life Sciences Research for Lifelong Health

Publications

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

From the stem of the placental tree: trophoblast stem cells and their progeny.
Latos PA, Hemberger M

Trophoblast stem cells (TSCs) retain the capacity to self-renew indefinitely and harbour the potential to differentiate into all trophoblast subtypes of the placenta. Recent studies have shown how signalling cascades integrate with transcription factor circuits to govern the fine balance between TSC self-renewal and differentiation. In addition, breakthroughs in reprogramming strategies have enabled the generation of TSCs from fibroblasts, opening up exciting new avenues that may allow the isolation of this stem cell type from other species, notably humans. Here, we review these recent advances in light of their importance for understanding placental pathologies and developing personalised medicine approaches for pregnancy complications.

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Development (Cambridge, England), 143, 1477-9129, 3650-3660, 2016

PMID: 27802134


Capture Hi-C identifies a novel causal gene, IL20RA, in the pan-autoimmune genetic susceptibility region 6q23.
McGovern A, Schoenfelder S, Martin P, Massey J, Duffus K, Plant D, Yarwood A, Pratt AG, Anderson AE, Isaacs JD, Diboll J, Thalayasingam N, Ospelt C, Barton A, Worthington J, Fraser P, Eyre S, Orozco G

The identification of causal genes from genome-wide association studies (GWAS) is the next important step for the translation of genetic findings into biologically meaningful mechanisms of disease and potential therapeutic targets. Using novel chromatin interaction detection techniques and allele specific assays in T and B cell lines, we provide compelling evidence that redefines causal genes at the 6q23 locus, one of the most important loci that confers autoimmunity risk.

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Genome biology, 17, 1474-760X, 212, 2016

PMID: 27799070


Open Access

Survival of mature T cells in the periphery is intrinsically dependent on GIMAP1 in mice.
Datta P, Webb LM, Avdo I, Pascall J, Butcher GW

An effective immune system depends upon the survival of mature T cells in the periphery. Members of the GIMAP family of GTPases have been proposed to regulate this homeostasis, supported by the paucity of peripheral T cells in rodents deficient for either GIMAP1 or GIMAP5. It is unclear whether this lack of T cells is a consequence of an ontological defect, causing the thymus to generate and export T cells incapable of surviving in the periphery, or whether (alternatively or additionally) mature T cells intrinsically require GIMAP1 for survival. Using the ER(T2) Cre(+) transgene, we conditionally deleted Gimap1 in C57BL/6 mice and demonstrate that GIMAP1 is intrinsically required for the survival of mature T cells in the periphery. We show that, in contrast to GIMAP5, this requirement is independent of the T cell's activation status. We investigated the nature of the survival defect in GIMAP1-deficient CD4(+) T cells and show that the death occurring after GIMAP1 ablation is accompanied by mitochondrial depolarisation and activation of the extrinsic apoptotic pathway. This study shows that GIMAP1 is critical for maintaining the peripheral T-cell pool in mice and offers a potent target for the treatment of T-cell-mediated diseases. This article is protected by copyright. All rights reserved.

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European journal of immunology, , 1521-4141, , 2016

PMID: 27792288


Open Access

Insulin resistance uncoupled from dyslipidemia due to C-terminal PIK3R1 mutations.
Huang-Doran I, Tomlinson P, Payne F, Gast A, Sleigh A, Bottomley W, Harris J, Daly A, Rocha N, Rudge S, Clark J, Kwok A, Romeo S, McCann E, Müksch B, Dattani M, Zucchini S, Wakelam M, Foukas LC, Savage DB, Murphy R, O'Rahilly S, Barroso I, Semple RK

Obesity-related insulin resistance is associated with fatty liver, dyslipidemia, and low plasma adiponectin. Insulin resistance due to insulin receptor (INSR) dysfunction is associated with none of these, but when due to dysfunction of the downstream kinase AKT2 phenocopies obesity-related insulin resistance. We report 5 patients with SHORT syndrome and C-terminal mutations in PIK3R1, encoding the p85α/p55α/p50α subunits of PI3K, which act between INSR and AKT in insulin signaling. Four of 5 patients had extreme insulin resistance without dyslipidemia or hepatic steatosis. In 3 of these 4, plasma adiponectin was preserved, as in insulin receptor dysfunction. The fourth patient and her healthy mother had low plasma adiponectin associated with a potentially novel mutation, p.Asp231Ala, in adiponectin itself. Cells studied from one patient with the p.Tyr657X PIK3R1 mutation expressed abundant truncated PIK3R1 products and showed severely reduced insulin-stimulated association of mutant but not WT p85α with IRS1, but normal downstream signaling. In 3T3-L1 preadipocytes, mutant p85α overexpression attenuated insulin-induced AKT phosphorylation and adipocyte differentiation. Thus, PIK3R1 C-terminal mutations impair insulin signaling only in some cellular contexts and produce a subphenotype of insulin resistance resembling INSR dysfunction but unlike AKT2 dysfunction, implicating PI3K in the pathogenesis of key components of the metabolic syndrome.

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JCI insight, 1, , e88766, 2016

PMID: 27766312


Open Access

Crosstalk between pluripotency factors and higher-order chromatin organization.
Lopes Novo C, Rugg-Gunn PJ

Pluripotent cells are characterized by a globally open and accessible chromatin organization that is thought to contribute to cellular plasticity and developmental decision-making. We recently identified the pluripotency factor Nanog as a key regulator of this form of chromatin architecture in mouse embryonic stem cells. In particular, we demonstrated that the transcription factors Nanog and Sall1 co-dependently mediate the epigenetic state of pericentromeric heterochromatin to reinforce a more open and accessible organization in pluripotent cells. Here, we summarize our main findings and place the work into a broader context. We explore how heterochromatin domains could be targets of transcriptional networks in pluripotent cells and are coordinated with cell state. We propose this integration may be to balance the requirement for a dynamic and plastic chromatin organization in pluripotent cells, together with priming for a more restrictive nuclear compartmentalization that is triggered rapidly upon lineage commitment.

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Nucleus (Austin, Tex.), , 1949-1042, 0, 2016

PMID: 27759487


Open Access

Epigenetic inheritance of proteostasis and ageing.
Li C, Casanueva O

Abundant evidence shows that the genome is not as static as once thought and that gene expression can be reversibly modulated by the environment. In some cases, these changes can be transmitted to the next generation even if the environment has reverted. Such transgenerational epigenetic inheritance requires that information be stored in the germline in response to exogenous stressors. One of the most elusive questions in the field of epigenetic inheritance is the identity of such inherited factor(s). Answering this question would allow us to understand how the environment can shape human populations for multiple generations and may help to explain the rapid rise in obesity and neurodegenerative diseases in modern society. It will also provide clues on how we might be able to reprogramme the epigenome to prevent transmission of detrimental phenotypes and identify individuals who might be at increased risk of disease. In this article, we aim to review recent developments in this field, focusing on research conducted mostly in the nematode Caenorhabditis elegans and mice, that link environmental modulators with the transgenerational inheritance of phenotypes that affect protein-folding homoeostasis and ageing.

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Essays in biochemistry, 60, 1744-1358, 191-202, 2016

PMID: 27744335


Open Access

Retinol and ascorbate drive erasure of epigenetic memory and enhance reprogramming to naïve pluripotency by complementary mechanisms.
Hore TA, von Meyenn F, Ravichandran M, Bachman M, Ficz G, Oxley D, Santos F, Balasubramanian S, Jurkowski TP, Reik W

Epigenetic memory, in particular DNA methylation, is established during development in differentiating cells and must be erased to create naïve (induced) pluripotent stem cells. The ten-eleven translocation (TET) enzymes can catalyze the oxidation of 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC) and further oxidized derivatives, thereby actively removing this memory. Nevertheless, the mechanism by which the TET enzymes are regulated, and the extent to which they can be manipulated, are poorly understood. Here we report that retinoic acid (RA) or retinol (vitamin A) and ascorbate (vitamin C) act as modulators of TET levels and activity. RA or retinol enhances 5hmC production in naïve embryonic stem cells by activation of TET2 and TET3 transcription, whereas ascorbate potentiates TET activity and 5hmC production through enhanced Fe(2+) recycling, and not as a cofactor as reported previously. We find that both ascorbate and RA or retinol promote the derivation of induced pluripotent stem cells synergistically and enhance the erasure of epigenetic memory. This mechanistic insight has significance for the development of cell treatments for regenenerative medicine, and enhances our understanding of how intrinsic and extrinsic signals shape the epigenome.

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Proceedings of the National Academy of Sciences of the United States of America, , 1091-6490, , 2016

PMID: 27729528


Open Access

Comparative Principles of DNA Methylation Reprogramming during Human and Mouse In Vitro Primordial Germ Cell Specification.
von Meyenn F, Berrens RV, Andrews S, Santos F, Collier AJ, Krueger F, Osorno R, Dean W, Rugg-Gunn PJ, Reik W

Primordial germ cell (PGC) development is characterized by global epigenetic remodeling, which resets genomic potential and establishes an epigenetic ground state. Here we recapitulate PGC specification in vitro from naive embryonic stem cells and characterize the early events of epigenetic reprogramming during the formation of the human and mouse germline. Following rapid de novo DNA methylation during priming to epiblast-like cells, methylation is globally erased in PGC-like cells. Repressive chromatin marks (H3K9me2/3) and transposable elements are enriched at demethylation-resistant regions, while active chromatin marks (H3K4me3 or H3K27ac) are more prominent at regions that demethylate faster. The dynamics of specification and epigenetic reprogramming show species-specific differences, in particular markedly slower reprogramming kinetics in the human germline. Differences in developmental kinetics may be explained by differential regulation of epigenetic modifiers. Our work establishes a robust and faithful experimental system of the early events of epigenetic reprogramming and regulation in the germline.

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Developmental cell, 39, 1878-1551, 104-115, 2016

PMID: 27728778


Open Access

Dynamics of mTORC1 activation in response to amino acids.
Manifava M, Smith M, Rotondo S, Walker S, Niewczas I, Zoncu R, Clark J, Ktistakis NT

Amino acids are essential activators of mTORC1 via a complex containing RAG GTPases, RAGULATOR and the vacuolar ATPase. Sensing of amino acids causes translocation of mTORC1 to lysosomes, an obligate step for activation. To examine the spatial and temporal dynamics of this translocation, we used live imaging of the mTORC1 component RAPTOR and a cell permeant fluorescent analogue of di-leucine methyl ester. Translocation to lysosomes is a transient event, occurring within 2 min of aa addition and peaking within 5 min. It is temporally coupled with fluorescent leucine appearance in lysosomes and is sustained in comparison to aa stimulation. Sestrin2 and the vacuolar ATPase are negative and positive regulators of mTORC1 activity in our experimental system. Of note, phosphorylation of canonical mTORC1 targets is delayed compared to lysosomal translocation suggesting a dynamic and transient passage of mTORC1 from the lysosomal surface before targetting its substrates elsewhere.

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eLife, 5, 2050-084X, , 2016

PMID: 27725083


Open Access

Shaping Variation in the Human Immune System.
Liston A, Carr EJ, Linterman MA

Immune responses demonstrate a high level of intra-species variation, compensating for the specialization capacity of pathogens. The recent advent of in-depth immune phenotyping projects in large-scale cohorts has allowed a first look into the factors that shape the inter-individual diversity of the human immune system. Genetic approaches have identified genetic diversity as drivers of 20-40% of the variation between the immune systems of individuals. The remaining 60-80% is shaped by intrinsic factors, with age being the predominant factor, as well as by environmental influences, where cohabitation and chronic viral infections were identified as key mediators. We review and integrate the recent in-depth large-scale studies on human immune diversity and its potential impact on health. VIDEO ABSTRACT.

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Trends in immunology, , 1471-4981, , 2016

PMID: 27693120


MERVL/Zscan4 Network Activation Results in Transient Genome-wide DNA Demethylation of mESCs.
Eckersley-Maslin MA, Svensson V, Krueger C, Stubbs TM, Giehr P, Krueger F, Miragaia RJ, Kyriakopoulos C, Berrens RV, Milagre I, Walter J, Teichmann SA, Reik W

Mouse embryonic stem cells are dynamic and heterogeneous. For example, rare cells cycle through a state characterized by decondensed chromatin and expression of transcripts, including the Zscan4 cluster and MERVL endogenous retrovirus, which are usually restricted to preimplantation embryos. Here, we further characterize the dynamics and consequences of this transient cell state. Single-cell transcriptomics identified the earliest upregulated transcripts as cells enter the MERVL/Zscan4 state. The MERVL/Zscan4 transcriptional network was also upregulated during induced pluripotent stem cell reprogramming. Genome-wide DNA methylation and chromatin analyses revealed global DNA hypomethylation accompanying increased chromatin accessibility. This transient DNA demethylation was driven by a loss of DNA methyltransferase proteins in the cells and occurred genome-wide. While methylation levels were restored once cells exit this state, genomic imprints remained hypomethylated, demonstrating a potential global and enduring influence of endogenous retroviral activation on the epigenome.

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Cell reports, 17, 2211-1247, 179-92, 2016

PMID: 27681430


Establishment and functions of DNA methylation in the germline.
Stewart KR, Veselovska L, Kelsey G

Epigenetic modifications established during gametogenesis regulate transcription and other nuclear processes in gametes, but also have influences in the zygote, embryo and postnatal life. This is best understood for DNA methylation which, established at discrete regions of the oocyte and sperm genomes, governs genomic imprinting. In this review, we describe how imprinting has informed our understanding of de novo DNA methylation mechanisms, highlight how recent genome-wide profiling studies have provided unprecedented insights into establishment of the sperm and oocyte methylomes and consider the fate and function of gametic methylation and other epigenetic modifications after fertilization.

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Epigenomics, , 1750-192X, , 2016

PMID: 27659720


The Phospholipase D2 Knock Out Mouse Has Ectopic Purkinje Cells and Suffers from Early Adult-Onset Anosmia.
Vermeren MM, Zhang Q, Smethurst E, Segonds-Pichon A, Schrewe H, Wakelam MJ

Phospholipase D2 (PLD2) is an enzyme that produces phosphatidic acid (PA), a lipid messenger molecule involved in a number of cellular events including, through its membrane curvature properties, endocytosis. The PLD2 knock out (PLD2KO) mouse has been previously reported to be protected from insult in a model of Alzheimer's disease. We have further analysed a PLD2KO mouse using mass spectrophotometry of its lipids and found significant differences in PA species throughout its brain. We have examined the expression pattern of PLD2 which allowed us to define which region of the brain to analyse for defect, notably PLD2 was not detected in glial-rich regions. The expression pattern lead us to specifically examine the mitral cells of olfactory bulbs, the Cornus Amonis (CA) regions of the hippocampus and the Purkinje cells of the cerebellum. We find that the change to longer PA species correlates with subtle architectural defect in the cerebellum, exemplified by ectopic Purkinje cells and an adult-onset deficit of olfaction. These observations draw parallels to defects in the reelin heterozygote as well as the effect of high fat diet on olfaction.

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PloS one, 11, 1932-6203, e0162814, 0

PMID: 27658289


Open Access

Targeting PI3K in Cancer: Impact on Tumor Cells, Their Protective Stroma, Angiogenesis, and Immunotherapy.
Okkenhaug K, Graupera M, Vanhaesebroeck B

The PI3K pathway is hyperactivated in most cancers, yet the capacity of PI3K inhibitors to induce tumor cell death is limited. The efficacy of PI3K inhibition can also derive from interference with the cancer cells' ability to respond to stromal signals, as illustrated by the approved PI3Kδ inhibitor idelalisib in B-cell malignancies. Inhibition of the leukocyte-enriched PI3Kδ or PI3Kγ may unleash antitumor T-cell responses by inhibiting regulatory T cells and immune-suppressive myeloid cells. Moreover, tumor angiogenesis may be targeted by PI3K inhibitors to enhance cancer therapy. Future work should therefore also explore the effects of PI3K inhibitors on the tumor stroma, in addition to their cancer cell-intrinsic impact.

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Cancer discovery, , 2159-8290, , 2016

PMID: 27655435


Characterization of Atg38 and NRBF2, a fifth subunit of the autophagic Vps34/PIK3C3 complex.
Ohashi Y, Soler N, García Ortegón M, Zhang L, Kirsten ML, Perisic O, Masson GR, Burke JE, Jakobi AJ, Apostolakis AA, Johnson CM, Ohashi M, Ktistakis NT, Sachse C, Williams RL

The phosphatidylinositol 3-kinase Vps34 is part of several protein complexes. The structural organization of heterotetrameric complexes is starting to emerge, but little is known about organization of additional accessory subunits that interact with these assemblies. Combining hydrogen-deuterium exchange mass spectrometry (HDX-MS), X-ray crystallography and electron microscopy (EM), we have characterized Atg38 and its human ortholog NRBF2, accessory components of complex I consisting of Vps15-Vps34-Vps30/Atg6-Atg14 (yeast) and PIK3R4/VPS15-PIK3C3/VPS34-BECN1/Beclin 1-ATG14 (human). HDX-MS shows that Atg38 binds the Vps30-Atg14 subcomplex of complex I, using mainly its N-terminal MIT domain and bridges the coiled-coil I regions of Atg14 and Vps30 in the base of complex I. The Atg38 C-terminal domain is important for localization to the phagophore assembly site (PAS) and homodimerization. Our 2.2 Å resolution crystal structure of the Atg38 C-terminal homodimerization domain shows 2 segments of α-helices assembling into a mushroom-like asymmetric homodimer with a 4-helix cap and a parallel coiled-coil stalk. One Atg38 homodimer engages a single complex I. This is in sharp contrast to human NRBF2, which also forms a homodimer, but this homodimer can bridge 2 complex I assemblies.

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Autophagy, , 1554-8635, 0, 2016

PMID: 27630019


Ionic immune suppression within the tumour microenvironment limits T cell effector function.
Eil R, Vodnala SK, Clever D, Klebanoff CA, Sukumar M, Pan JH, Palmer DC, Gros A, Yamamoto TN, Patel SJ, Guittard GC, Yu Z, Carbonaro V, Okkenhaug K, Schrump DS, Linehan WM, Roychoudhuri R, Restifo NP

Tumours progress despite being infiltrated by tumour-specific effector T cells. Tumours contain areas of cellular necrosis, which are associated with poor survival in a variety of cancers. Here, we show that necrosis releases intracellular potassium ions into the extracellular fluid of mouse and human tumours, causing profound suppression of T cell effector function. Elevation of the extracellular potassium concentration ([K(+)]e) impairs T cell receptor (TCR)-driven Akt-mTOR phosphorylation and effector programmes. Potassium-mediated suppression of Akt-mTOR signalling and T cell function is dependent upon the activity of the serine/threonine phosphatase PP2A. Although the suppressive effect mediated by elevated [K(+)]e is independent of changes in plasma membrane potential (Vm), it requires an increase in intracellular potassium ([K(+)]i). Accordingly, augmenting potassium efflux in tumour-specific T cells by overexpressing the potassium channel Kv1.3 lowers [K(+)]i and improves effector functions in vitro and in vivo and enhances tumour clearance and survival in melanoma-bearing mice. These results uncover an ionic checkpoint that blocks T cell function in tumours and identify potential new strategies for cancer immunotherapy.

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Nature, 537, 1476-4687, 539-543, 2016

PMID: 27626381


PIKfyve Regulates Vacuole Maturation and Nutrient Recovery following Engulfment.
Krishna S, Palm W, Lee Y, Yang W, Bandyopadhyay U, Xu H, Florey O, Thompson CB, Overholtzer M

The scavenging of extracellular macromolecules by engulfment can sustain cell growth in a nutrient-depleted environment. Engulfed macromolecules are contained within vacuoles that are targeted for lysosome fusion to initiate degradation and nutrient export. We have shown that vacuoles containing engulfed material undergo mTORC1-dependent fission that redistributes degraded cargo back into the endosomal network. Here we identify the lipid kinase PIKfyve as a regulator of an alternative pathway that distributes engulfed contents in support of intracellular macromolecular synthesis during macropinocytosis, entosis, and phagocytosis. We find that PIKfyve regulates vacuole size in part through its downstream effector, the cationic transporter TRPML1. Furthermore, PIKfyve promotes recovery of nutrients from vacuoles, suggesting a potential link between PIKfyve activity and lysosomal nutrient export. During nutrient depletion, PIKfyve activity protects Ras-mutant cells from starvation-induced cell death and supports their proliferation. These data identify PIKfyve as a critical regulator of vacuole maturation and nutrient recovery during engulfment.

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Developmental cell, 38, 1878-1551, 536-47, 2016

PMID: 27623384


PI3Kδ and primary immunodeficiencies.
Lucas CL, Chandra A, Nejentsev S, Condliffe AM, Okkenhaug K

Primary immunodeficiencies are inherited disorders of the immune system, often caused by the mutation of genes required for lymphocyte development and activation. Recently, several studies have identified gain-of-function mutations in the phosphoinositide 3-kinase (PI3K) genes PIK3CD (which encodes p110δ) and PIK3R1 (which encodes p85α) that cause a combined immunodeficiency syndrome, referred to as activated PI3Kδ syndrome (APDS; also known as p110δ-activating mutation causing senescent T cells, lymphadenopathy and immunodeficiency (PASLI)). Paradoxically, both loss-of-function and gain-of-function mutations that affect these genes lead to immunosuppression, albeit via different mechanisms. Here, we review the roles of PI3Kδ in adaptive immunity, describe the clinical manifestations and mechanisms of disease in APDS and highlight new insights into PI3Kδ gleaned from these patients, as well as implications of these findings for clinical therapy.

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Nature reviews. Immunology, , 1474-1741, , 2016

PMID: 27616589


Open Access

The health care and life sciences community profile for dataset descriptions.
Dumontier M, Gray AJ, Marshall MS, Alexiev V, Ansell P, Bader G, Baran J, Bolleman JT, Callahan A, Cruz-Toledo J, Gaudet P, Gombocz EA, Gonzalez-Beltran AN, Groth P, Haendel M, Ito M, Jupp S, Juty N, Katayama T, Kobayashi N, Krishnaswami K, Laibe C, Le Novère N, Lin S, Malone J, Miller M, Mungall CJ, Rietveld L, Wimalaratne SM, Yamaguchi A

Access to consistent, high-quality metadata is critical to finding, understanding, and reusing scientific data. However, while there are many relevant vocabularies for the annotation of a dataset, none sufficiently captures all the necessary metadata. This prevents uniform indexing and querying of dataset repositories. Towards providing a practical guide for producing a high quality description of biomedical datasets, the W3C Semantic Web for Health Care and the Life Sciences Interest Group (HCLSIG) identified Resource Description Framework (RDF) vocabularies that could be used to specify common metadata elements and their value sets. The resulting guideline covers elements of description, identification, attribution, versioning, provenance, and content summarization. This guideline reuses existing vocabularies, and is intended to meet key functional requirements including indexing, discovery, exchange, query, and retrieval of datasets, thereby enabling the publication of FAIR data. The resulting metadata profile is generic and could be used by other domains with an interest in providing machine readable descriptions of versioned datasets.

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PeerJ, 4, 2167-8359, e2331, 2016

PMID: 27602295


Open Access

In B cells, phosphatidylinositol 5-phosphate 4-kinase-α synthesizes PI(4,5)P2 to impact mTORC2 and Akt signaling.
Bulley SJ, Droubi A, Clarke JH, Anderson KE, Stephens LR, Hawkins PT, Irvine RF

Phosphatidylinositol 5-phosphate 4-kinases (PI5P4Ks) are enigmatic lipid kinases with physiological functions that are incompletely understood, not the least because genetic deletion and cell transfection have led to contradictory data. Here, we used the genetic tractability of DT40 cells to create cell lines in which endogenous PI5P4Kα was removed, either stably by genetic deletion or transiently (within 1 h) by tagging the endogenous protein genomically with the auxin degron. In both cases, removal impacted Akt phosphorylation, and by leaving one PI5P4Kα allele present but mutating it to be kinase-dead or have PI4P 5-kinase activity, we show that all of the effects on Akt phosphorylation were dependent on the ability of PI5P4Kα to synthesize phosphatidylinositol (4,5)-bisphosphate [PI(4,5)P2] rather than to remove PI5P. Although stable removal of PI5P4Kα resulted in a pronounced decrease in Akt phosphorylation at Thr308 and Ser473, in part because of reduced plasma membrane PIP3, its acute removal led to an increase in Akt phosphorylation only at Ser473. This process invokes activation primarily of mammalian target of rapamycin complex 2 (mTORC2), which was confirmed by increased phosphorylation of other mTORC2 substrates. These findings establish PI5P4Kα as a kinase that synthesizes a physiologically relevant pool of PI(4,5)P2 and as a regulator of mTORC2, and show a phenomenon similar to the "butterfly effect" described for phosphatidylinositol 3-kinase Iα [Hart JR, et al. (2015) Proc Natl Acad Sci USA 112(4):1131-1136], whereby through apparently the same underlying mechanism, the removal of a protein's activity from a cell can have widely divergent effects depending on the time course of that removal.

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Proceedings of the National Academy of Sciences of the United States of America, , 1091-6490, , 2016

PMID: 27601656


Open Access

Phosphoproteomic Analyses of Interleukin 2 Signaling Reveal Integrated JAK Kinase-Dependent and -Independent Networks in CD8(+) T Cells.
Ross SH, Rollings C, Anderson KE, Hawkins PT, Stephens LR, Cantrell DA

Interleukin-2 (IL-2) is a fundamental cytokine that controls proliferation and differentiation of T cells. Here, we used high-resolution mass spectrometry to generate a comprehensive and detailed map of IL-2 protein phosphorylations in cytotoxic T cells (CTL). The data revealed that Janus kinases (JAKs) couple IL-2 receptors to the coordinated phosphorylation of transcription factors, regulators of chromatin, mRNA translation, GTPases, vesicle trafficking, and the actin and microtubule cytoskeleton. We identified an IL-2-JAK-independent SRC family Tyr-kinase-controlled signaling network that regulates ∼10% of the CTL phosphoproteome, the production of phosphatidylinositol (3,4,5)-trisphosphate (PIP3), and the activity of the serine/threonine kinase AKT. These data reveal a signaling framework wherein IL-2-JAK-controlled pathways coordinate with IL-2-independent networks of kinase activity and provide a resource toward the further understanding of the networks of protein phosphorylation that program CTL fate.

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Immunity, , 1097-4180, , 2016

PMID: 27566939


Open Access

The RNA-Binding Proteins Zfp36l1 and Zfp36l2 Enforce the Thymic β-Selection Checkpoint by Limiting DNA Damage Response Signaling and Cell Cycle Progression.
Vogel KU, Bell LS, Galloway A, Ahlfors H, Turner M

The RNA-binding proteins Zfp36l1 and Zfp36l2 act redundantly to enforce the β-selection checkpoint during thymopoiesis, yet their molecular targets remain largely unknown. In this study, we identify these targets on a genome-wide scale in primary mouse thymocytes and show that Zfp36l1/l2 regulate DNA damage response and cell cycle transcripts to ensure proper β-selection. Double-negative 3 thymocytes lacking Zfp36l1/l2 share a gene expression profile with postselected double-negative 3b cells despite the absence of intracellular TCRβ and reduced IL-7 signaling. Our findings show that in addition to controlling the timing of proliferation at β-selection, posttranscriptional control by Zfp36l1/l2 limits DNA damage responses, which are known to promote thymocyte differentiation. Zfp36l1/l2 therefore act as posttranscriptional safeguards against chromosomal instability and replication stress by integrating pre-TCR and IL-7 signaling with DNA damage and cell cycle control.

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Journal of immunology (Baltimore, Md. : 1950), , 1550-6606, , 2016

PMID: 27566829


Oxygen Sensing by T Cells Establishes an Immunologically Tolerant Metastatic Niche.
Clever D, Roychoudhuri R, Constantinides MG, Askenase MH, Sukumar M, Klebanoff CA, Eil RL, Hickman HD, Yu Z, Pan JH, Palmer DC, Phan AT, Goulding J, Gattinoni L, Goldrath AW, Belkaid Y, Restifo NP

Cancer cells must evade immune responses at distant sites to establish metastases. The lung is a frequent site for metastasis. We hypothesized that lung-specific immunoregulatory mechanisms create an immunologically permissive environment for tumor colonization. We found that T-cell-intrinsic expression of the oxygen-sensing prolyl-hydroxylase (PHD) proteins is required to maintain local tolerance against innocuous antigens in the lung but powerfully licenses colonization by circulating tumor cells. PHD proteins limit pulmonary type helper (Th)-1 responses, promote CD4(+)-regulatory T (Treg) cell induction, and restrain CD8(+) T cell effector function. Tumor colonization is accompanied by PHD-protein-dependent induction of pulmonary Treg cells and suppression of IFN-γ-dependent tumor clearance. T-cell-intrinsic deletion or pharmacological inhibition of PHD proteins limits tumor colonization of the lung and improves the efficacy of adoptive cell transfer immunotherapy. Collectively, PHD proteins function in T cells to coordinate distinct immunoregulatory programs within the lung that are permissive to cancer metastasis. PAPERCLIP.

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Cell, 166, 1097-4172, 1117-1131.e14, 2016

PMID: 27565342


Exosomes bind autotaxin and act as a physiological delivery mechanism to stimulate LPA receptor signalling in cells.
Jethwa SA, Leah EJ, Zhang Q, Bright NA, Oxley D, Bootman MD, Rudge SA, Wakelam MJ

Autotaxin (ATX) the lysophospholipase responsible for generating the lipid receptor agonist lysophosphatidic acid (LPA) is a secreted enzyme. Here we show that once secreted it can bind to the surface of cell secreted exosomes. Exosome-bound ATX is catalytically active and carries generated LPA. Once bound to a cell, through specific integrin interaction, ATX releases the LPA to activate cell surface G-protein coupled LPA receptors; inhibition of signaling by the receptor antagonist Ki1642 suggests these are either LPAR1 or LPAR3. The binding stimulates downstream signaling including AKT and MAPK phosphorylation, the release of intracellular stored calcium and cell migration. We propose that exosomal binding of LPA-loaded ATX provides a means of efficiently delivering the lipid agonist to cell surface receptors to promote signalling. We further propose that this is a means whereby autotaxin-LPA signaling operates physiologically.

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Journal of cell science, , 1477-9137, , 2016

PMID: 27557622


Clinical spectrum and features of activated phosphoinositide 3-kinase δ syndrome: A large patient cohort study.
Coulter TI, Chandra A, Bacon CM, Babar J, Curtis J, Screaton N, Goodlad JR, Farmer G, Steele CL, Leahy TR, Doffinger R, Baxendale H, Bernatoniene J, Edgar JD, Longhurst HJ, Ehl S, Speckmann C, Grimbacher B, Sediva A, Milota T, Faust SN, Williams AP, Hayman G, Kucuk ZY, Hague R, French P, Brooker R, Forsyth P, Herriot R, Cancrini C, Palma P, Ariganello P, Conlon N, Feighery C, Gavin PJ, Jones A, Imai K, Ibrahim MA, Markelj G, Abinun M, Rieux-Laucat F, Latour S, Pellier I, Fischer A, Touzot F, Casanova JL, Durandy A, Burns SO, Savic S, Kumararatne DS, Moshous D, Kracker S, Vanhaesebroeck B, Okkenhaug K, Picard C, Nejentsev S, Condliffe AM, Cant AJ

Activated phosphoinositide 3-kinase δ syndrome (APDS) is a recently described combined immunodeficiency resulting from gain-of-function mutations in PIK3CD, the gene encoding the catalytic subunit of phosphoinositide 3-kinase δ (PI3Kδ).

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The Journal of allergy and clinical immunology, , 1097-6825, , 2016

PMID: 27555459


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