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

Single cell transcriptome analysis of human, marmoset and mouse embryos reveals common and divergent features of preimplantation development.
Boroviak T, Stirparo GG, Dietmann S, Hernando-Herraez I, Mohammed H, Reik W, Smith A, Sasaki E, Nichols J, Bertone P

The mouse embryo is the canonical model for mammalian preimplantation development. Recent advances in single cell profiling allow detailed analysis of embryogenesis in other eutherian species, including human, to distinguish conserved from divergent regulatory programs and signalling pathways in the rodent paradigm. Here, we identify and compare transcriptional features of human, marmoset and mouse embryos by single cell RNA-seq. Zygotic genome activation correlates with the presence of polycomb repressive complexes in all three species, while ribosome biogenesis emerges as a predominant attribute in primate embryos, supporting prolonged translation of maternally deposited RNAs. We find that transposable element expression signatures are species, stage and lineage specific. The pluripotency network in the primate epiblast lacks certain regulators that are operative in mouse, but encompasses WNT components and genes associated with trophoblast specification. Sequential activation of GATA6, SOX17 and GATA4 markers of primitive endoderm identity is conserved in primates. Unexpectedly, OTX2 is also associated with primitive endoderm specification in human and non-human primate blastocysts. Our cross-species analysis demarcates both conserved and primate-specific features of preimplantation development, and underscores the molecular adaptability of early mammalian embryogenesis.

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Development (Cambridge, England), 145, 1477-9129, , 2018

PMID: 30413530


Regulation of the Germinal Center Response.
Stebegg M, Kumar SD, Silva-Cayetano A, Fonseca VR, Linterman MA, Graca L

The germinal center (GC) is a specialized microstructure that forms in secondary lymphoid tissues, producing long-lived antibody secreting plasma cells and memory B cells, which can provide protection against reinfection. Within the GC, B cells undergo somatic mutation of the genes encoding their B cell receptors which, following successful selection, can lead to the emergence of B cell clones that bind antigen with high affinity. However, this mutation process can also be dangerous, as it can create autoreactive clones that can cause autoimmunity. Because of this, regulation of GC reactions is critical to ensure high affinity antibody production and to enforce self-tolerance by avoiding emergence of autoreactive B cell clones. A productive GC response requires the collaboration of multiple cell types. The stromal cell network orchestrates GC cell dynamics by controlling antigen delivery and cell trafficking. T follicular helper (Tfh) cells provide specialized help to GC B cells through cognate T-B cell interactions while Foxp3 T follicular regulatory (Tfr) cells are key mediators of GC regulation. However, regulation of GC responses is not a simple outcome of Tfh/Tfr balance, but also involves the contribution of other cell types to modulate the GC microenvironment and to avoid autoimmunity. Thus, the regulation of the GC is complex, and occurs at multiple levels. In this review we outline recent developments in the biology of cell subsets involved in the regulation of GC reactions, in both secondary lymphoid tissues, and Peyer's patches (PPs). We discuss the mechanisms which enable the generation of potent protective humoral immunity whilst GC-derived autoimmunity is avoided.

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Frontiers in immunology, 9, 1664-3224, 2469, 2018

PMID: 30410492


Open Access

Alpha-synuclein fibrils recruit TBK1 and OPTN to lysosomal damage sites and induce autophagy in microglial cells.
Bussi C, Peralta Ramos JM, Arroyo DS, Gallea JI, Ronchi P, Kolovou A, Wang JM, Florey O, Celej MS, Schwab Y, Ktistakis NT, Iribarren P

Autophagic dysfunction and protein aggregation have been linked to several neurodegenerative disorders, but the exact mechanisms and causal connections are not clear and most work was done in neurons and not in microglial cells. Here we report that exogenous fibrillar but not monomeric alpha-synuclein (AS) induces autophagy in microglial cells. We extensively studied the dynamics of this response by both live-cell imaging and correlative light-electron microscopy (CLEM) and found that it correlates with lysosomal damage and is characterised by the recruitment of the selective autophagy-associated proteins TANK-binding kinase 1 (TBK1) and Optineurin (OPTN) to ubiquitinated lysosomes. In addition, we observed that LC3 recruitment to damaged lysosomes was dependent on TBK1 activity. In these fibrillar AS-treated cells, autophagy inhibition impairs mitochondrial function and leads to microglial cell death. Our results suggest that microglial autophagy is induced in response to lysosomal damage caused by persistent accumulation of AS fibrils. Importantly, triggering of the autophagic response appears to be an attempt at lysosomal quality control and not for engulfment of fibrillar AS.

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Journal of cell science, , 1477-9137, , 2018

PMID: 30404831


The ATG5-binding and coiled coil domains of ATG16L1 maintain autophagy and tissue homeostasis in mice independently of the WD domain required for LC3-associated phagocytosis.
Rai S, Arasteh M, Jefferson M, Pearson T, Wang Y, Zhang W, Bicsak B, Divekar D, Powell PP, Nauman R, Beraza N, Carding SR, Florey O, Mayer U, Wileman T

Macroautophagy/autophagy delivers damaged proteins and organelles to lysosomes for degradation, and plays important roles in maintaining tissue homeostasis by reducing tissue damage. The translocation of LC3 to the limiting membrane of the phagophore, the precursor to the autophagosome, during autophagy provides a binding site for autophagy cargoes, and facilitates fusion with lysosomes. An autophagy-related pathway called LC3-associated phagocytosis (LAP) targets LC3 to phagosome and endosome membranes during uptake of bacterial and fungal pathogens, and targets LC3 to swollen endosomes containing particulate material or apoptotic cells. We have investigated the roles played by autophagy and LAP in vivo by exploiting the observation that the WD domain of ATG16L1 is required for LAP, but not autophagy. Mice lacking the linker and WD domains, activate autophagy, but are deficient in LAP. The LAP mice survive postnatal starvation, grow at the same rate as littermate controls, and are fertile. The liver, kidney, brain and muscle of these mice maintain levels of autophagy cargoes such as LC3 and SQSTM1/p62 similar to littermate controls, and prevent accumulation of SQSTM1 inclusions and tissue damage associated with loss of autophagy. The results suggest that autophagy maintains tissue homeostasis in mice independently of LC3-associated phagocytosis. Further deletion of glutamate E230 in the coiled-coil domain required for WIPI2 binding produced mice with defective autophagy that survived neonatal starvation. Analysis of brain lysates suggested that interactions between WIPI2 and ATG16L1 were less critical for autophagy in the brain, which may allow a low level of autophagy to overcome neonatal lethality. Abbreviations: CCD: coiled-coil domain; CYBB/NOX2: cytochrome b-245: beta polypeptide; GPT/ALT: glutamic pyruvic transaminase: soluble; LAP: LC3-associated phagocytosis; LC3: microtubule-associated protein 1 light chain 3; MEF: mouse embryonic fibroblast; NOD: nucleotide-binding oligomerization domain; NADPH: nicotinamide adenine dinucleotide phosphate; RUBCN/Rubicon: RUN domain and cysteine-rich domain containing Beclin 1-interacting protein; SLE: systemic lupus erythematosus; SQSTM1/p62: sequestosome 1; TLR: toll-like receptor; TMEM: transmembrane protein; TRIM: tripartite motif-containing protein; UVRAG: UV radiation resistance associated gene; WD: tryptophan-aspartic acid; WIPI: WD 40 repeat domain: phosphoinositide interacting.

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Autophagy, , 1554-8635, 1-14, 2018

PMID: 30403914


Disease-relevant transcriptional signatures identified in individual smooth muscle cells from healthy mouse vessels.
Dobnikar L, Taylor AL, Chappell J, Oldach P, Harman JL, Oerton E, Dzierzak E, Bennett MR, Spivakov M, Jørgensen HF

Vascular smooth muscle cells (VSMCs) show pronounced heterogeneity across and within vascular beds, with direct implications for their function in injury response and atherosclerosis. Here we combine single-cell transcriptomics with lineage tracing to examine VSMC heterogeneity in healthy mouse vessels. The transcriptional profiles of single VSMCs consistently reflect their region-specific developmental history and show heterogeneous expression of vascular disease-associated genes involved in inflammation, adhesion and migration. We detect a rare population of VSMC-lineage cells that express the multipotent progenitor marker Sca1, progressively downregulate contractile VSMC genes and upregulate genes associated with VSMC response to inflammation and growth factors. We find that Sca1 upregulation is a hallmark of VSMCs undergoing phenotypic switching in vitro and in vivo, and reveal an equivalent population of Sca1-positive VSMC-lineage cells in atherosclerotic plaques. Together, our analyses identify disease-relevant transcriptional signatures in VSMC-lineage cells in healthy blood vessels, with implications for disease susceptibility, diagnosis and prevention.

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Nature communications, 9, 2041-1723, 4567, 2018

PMID: 30385745


Open Access

5-Formylcytosine organizes nucleosomes and forms Schiff base interactions with histones in mouse embryonic stem cells.
Raiber EA, Portella G, Martínez Cuesta S, Hardisty R, Murat P, Li Z, Iurlaro M, Dean W, Spindel J, Beraldi D, Liu Z, Dawson MA, Reik W, Balasubramanian S

Nucleosomes are the basic unit of chromatin that help the packaging of genetic material while controlling access to the genetic information. The underlying DNA sequence, together with transcription-associated proteins and chromatin remodelling complexes, are important factors that influence the organization of nucleosomes. Here, we show that the naturally occurring DNA modification, 5-formylcytosine (5fC) is linked to tissue-specific nucleosome organization. Our study reveals that 5fC is associated with increased nucleosome occupancy in vitro and in vivo. We demonstrate that 5fC-associated nucleosomes at enhancers in the mammalian hindbrain and heart are linked to elevated gene expression. Our study also reveals the formation of a reversible-covalent Schiff base linkage between lysines of histone proteins and 5fC within nucleosomes in a cellular environment. We define their specific genomic loci in mouse embryonic stem cells and look into the biological consequences of these DNA-histone Schiff base sites. Collectively, our findings show that 5fC is a determinant of nucleosome organization and plays a role in establishing distinct regulatory regions that control transcription.

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Nature chemistry, , 1755-4349, , 2018

PMID: 30349137


A closer look at neuron interaction with track-etched microporous membranes.
George JH, Nagel D, Waller S, Hill E, Parri HR, Coleman MD, Cui Z, Ye H

Microporous membranes support the growth of neurites into and through micro-channels, providing a different type of neural growth platform to conventional dish cultures. Microporous membranes are used to support various types of culture, however, the role of pore diameter in relation to neurite growth through the membrane has not been well characterised. In this study, the human cell line (SH-SY5Y) was differentiated into neuron-like cells and cultured on track-etched microporous membranes with pore and channel diameters selected to accommodate neurite width (0.8 µm to 5 µm). Whilst neurites extended through all pore diameters, the extent of neurite coverage on the non-seeded side of the membranes after 5 days in culture was found to be directly proportional to channel diameter. Neurite growth through membrane pores reduced significantly when neural cultures were non-confluent. Scanning electron microscopy revealed that neurites bridged pores and circumnavigated pore edges - such that the overall likelihood of a neurite entering a pore channel was decreased. These findings highlight the role of pore diameter, cell sheet confluence and contact guidance in directing neurite growth through pores and may be useful in applications that seek to use physical substrates to maintain separate neural populations whilst permitting neurite contact between cultures.

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Scientific reports, 8, 2045-2322, 15552, 2018

PMID: 30341335


Open Access

P7C3-A20 neuroprotection is independent of Wallerian degeneration in primary neuronal culture.
Hill CS, Menon DK, Coleman MP

The antiapoptotic, neuroprotective compound P7C3-A20 reduces neurological deficits when administered to murine in-vivo models of traumatic brain injury. P7C3-A20 is thought to exert its activity through small-molecule activation of the enzyme nicotinamide phosphoribosyltransferase. This enzyme converts nicotinamide to nicotinamide mononucleotide, the precursor to nicotinamide adenine dinucleotide synthesis. Alterations to this bioenergetic pathway have been shown to induce Wallerian degeneration (WD) of the distal neurite following injury. This study aimed to establish whether P7C3-A20, through induction of nicotinamide phosphoribosyltransferase activity, would affect the rate of WD. The model systems used were dissociated primary cortical neurons, dissociated superior cervical ganglion neurons and superior cervical ganglion explants. P7C3-A20 failed to show any protection against WD induced by neurite transection or vincristine administration. Furthermore, there was a concentration-dependent neurotoxicity. These findings are important in understanding the mechanism by which P7C3-A20 mediates its effects - a key step before moving to human clinical trials.This is an open access article distributed under the Creative Commons Attribution License 4.0 (CCBY), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. http://creativecommons.org/licenses/by/4.0/.

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Neuroreport, , 1473-558X, , 2018

PMID: 30334859


Mice Deficient in Nucleoporin Nup210 Develop Peripheral T Cell Alterations.
van Nieuwenhuijze A, Burton O, Lemaitre P, Denton AE, Cascalho A, Goodchild RE, Malengier-Devlies B, Cauwe B, Linterman MA, Humblet-Baron S, Liston A

The nucleopore is an essential structure of the eukaryotic cell, regulating passage between the nucleus and cytoplasm. While individual functions of core nucleopore proteins have been identified, the role of other components, such as Nup210, are poorly defined. Here, through the use of an unbiased ENU mutagenesis screen for mutations effecting the peripheral T cell compartment, we identified a Nup210 mutation in a mouse strain with altered CD4/CD8 T cell ratios. Through the generation of Nup210 knockout mice we identified Nup210 as having a T cell-intrinsic function in the peripheral homeostasis of T cells. Remarkably, despite the deep evolutionary conservation of this key nucleopore complex member, no other major phenotypes developed, with viable and healthy knockout mice. These results identify Nup210 as an important nucleopore complex component for peripheral T cells, and raise further questions of why this nucleopore component shows deep evolutionary conservation despite seemingly redundant functions in most cell types.

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Frontiers in immunology, 9, 1664-3224, 2234, 2018

PMID: 30323813


Open Access

Regulation of Placental Development and Its Impact on Fetal Growth-New Insights From Mouse Models.
Woods L, Perez-Garcia V, Hemberger M

The placenta is the chief regulator of nutrient supply to the growing embryo during gestation. As such, adequate placental function is instrumental for developmental progression throughout intrauterine development. One of the most common complications during pregnancy is insufficient growth of the fetus, a problem termed intrauterine growth restriction (IUGR) that is most frequently rooted in a malfunctional placenta. Together with conventional gene targeting approaches, recent advances in screening mouse mutants for placental defects, combined with the ability to rapidly induce mutations and by CRISPR-Cas9 technology, has provided new insights into the contribution of the genome to normal placental development. Most importantly, these data have demonstrated that far more genes are required for normal placentation than previously appreciated. Here, we provide a summary of common types of placental defects in established mouse mutants, which will help us gain a better understanding of the genes impacting on human placentation. Based on a recent mouse mutant screen, we then provide examples on how these data can be mined to identify novel molecular hubs that may be critical for placental development. Given the close association between placental defects and abnormal cardiovascular and brain development, these functional nodes may also shed light onto the etiology of birth defects that co-occur with placental malformations. Taken together, recent insights into the regulation of mouse placental development have opened up new avenues for research that will promote the study of human pregnancy conditions, notably those based on defects in placentation that underlie the most common pregnancy pathologies such as IUGR and pre-eclampsia.

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Frontiers in endocrinology, 9, 1664-2392, 570, 2018

PMID: 30319550


Open Access

Transgenerational transmission of hedonic behaviors and metabolic phenotypes induced by maternal overnutrition.
Sarker G, Berrens R, von Arx J, Pelczar P, Reik W, Wolfrum C, Peleg-Raibstein D

Maternal overnutrition has been associated with increased susceptibility to develop obesity and neurological disorders later in life. Most epidemiological as well as experimental studies have focused on the metabolic consequences across generations following an early developmental nutritional insult. Recently, it has been shown that maternal high-fat diet (HFD) affects third-generation female body mass via the paternal lineage. We showed here that the offspring born to HFD ancestors displayed addictive-like behaviors as well as obesity and insulin resistance up to the third generation in the absence of any further exposure to HFD. These findings, implicate that the male germ line is a major player in transferring phenotypic traits. These behavioral and physiological alterations were paralleled by reduced striatal dopamine levels and increased dopamine 2 receptor density. Interestingly, by the third generation a clear gender segregation emerged, where females showed addictive-like behaviors while male HFD offspring showed an obesogenic phenotype. However, methylome profiling of F1 and F2 sperm revealed no significant difference between the offspring groups, suggesting that the sperm methylome might not be the major carrier for the transmission of the phenotypes observed in our mouse model. Together, our study for the first time demonstrates that maternal HFD insult causes sustained alterations of the mesolimbic dopaminergic system suggestive of a predisposition to develop obesity and addictive-like behaviors across multiple generations.

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Translational psychiatry, 8, 2158-3188, 195, 2018

PMID: 30315171


Open Access

Divergent wiring of repressive and active chromatin interactions between mouse embryonic and trophoblast lineages.
Schoenfelder S, Mifsud B, Senner CE, Todd CD, Chrysanthou S, Darbo E, Hemberger M, Branco MR

The establishment of the embryonic and trophoblast lineages is a developmental decision underpinned by dramatic differences in the epigenetic landscape of the two compartments. However, it remains unknown how epigenetic information and transcription factor networks map to the 3D arrangement of the genome, which in turn may mediate transcriptional divergence between the two cell lineages. Here, we perform promoter capture Hi-C experiments in mouse trophoblast (TSC) and embryonic (ESC) stem cells to understand how chromatin conformation relates to cell-specific transcriptional programmes. We find that key TSC genes that are kept repressed in ESCs exhibit interactions between H3K27me3-marked regions in ESCs that depend on Polycomb repressive complex 1. Interactions that are prominent in TSCs are enriched for enhancer-gene contacts involving key TSC transcription factors, as well as TET1, which helps to maintain the expression of TSC-relevant genes. Our work shows that the first developmental cell fate decision results in distinct chromatin conformation patterns establishing lineage-specific contexts involving both repressive and active interactions.

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Nature communications, 9, 2041-1723, 4189, 2018

PMID: 30305613


Open Access

Low levels of NMNAT2 compromise axon development and survival.
Gilley J, Mayer P, Yu G, Coleman MP

NMNAT2 is an endogenous axon maintenance factor that preserves axon health by blocking Wallerian-like axon degeneration. Mice lacking NMNAT2 die at birth with severe axon defects in both the PNS and CNS so a complete absence of NMNAT2 in humans is likely to be similarly harmful, but probably rare. However, there is evidence of widespread natural variation in human NMNAT2 mRNA expression so it is important to establish whether reduced levels of NMNAT2 have consequences that impact health. Whilst mice that express reduced levels of NMNAT2, either those heterozygous for a silenced Nmnat2 allele, or compound heterozygous for one silenced and one partially silenced Nmnat2 allele, remain overtly normal into old age, we now report that Nmnat2 compound heterozygote mice present with early and age-dependent peripheral nerve axon defects. Compound heterozygote mice already have reduced numbers of myelinated sensory axons at 1.5 months and lose more axons, likely motor axons, between 18 and 24 months and, crucially, these changes correlate with early temperature insensitivity and a later-onset decline in motor performance. Slower neurite outgrowth and increased sensitivity to axonal stress are also evident in primary cultures of Nmnat2 compound heterozygote superior cervical ganglion neurons. These data reveal that reducing NMNAT2 levels below a particular threshold compromises the development of peripheral axons and increases their vulnerability to stresses. We discuss the implications for human neurological phenotypes where axons are longer and have to be maintained over a much longer lifespan.

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Human molecular genetics, , 1460-2083, , 2018

PMID: 30304512


Enzymatic Assays of Histone Decrotonylation on Recombinant Histones.
Fellows R, Varga-Weisz P

Class I histone deacetylases (HDACs) are efficient histone decrotonylases, broadening the enzymatic spectrum of these important (epi-)genome regulators and drug targets. Here, we describe an approach to assaying class I HDACs with different acyl-histone substrates, including crotonylated histones and expand this to examine the effect of inhibitors and estimate kinetic constants.

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Bio-protocol, 8, 2331-8325, , 2018

PMID: 30283810


Open Access

Tri-methylation of histone H3 lysine 4 facilitates gene expression in ageing cells.
Cruz C, Della Rosa M, Krueger C, Gao Q, Horkai D, King M, Field L, Houseley J

Transcription of protein coding genes is accompanied by recruitment of COMPASS to promoter-proximal chromatin, which methylates histone H3 lysine 4 (H3K4) to form H3K4me1, H3K4me2 and H3K4me3. Here, we determine the importance of COMPASS in maintaining gene expression across lifespan in budding yeast. We find that COMPASS mutations reduce replicative lifespan and cause expression defects in almost 500 genes. Although H3K4 methylation is reported to act primarily in gene repression, particularly in yeast, repressive functions are progressively lost with age while hundreds of genes become dependent on H3K4me3 for full expression. Basal and inducible expression of these genes is also impaired in young cells lacking COMPASS components Swd1 or Spp1. Gene induction during ageing is associated with increasing promoter H3K4me3, but H3K4me3 also accumulates in non-promoter regions and the ribosomal DNA. Our results provide clear evidence that H3K4me3 is required to maintain normal expression of many genes across organismal lifespan.

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eLife, 7, 2050-084X, , 2018

PMID: 30274593


Open Access

CD151 regulates expression of FGFR2 in breast cancer cells via PKC-dependent pathways.
Sadej R, Lu X, Turczyk L, Novitskaya V, Lopez-Clavijo AF, Kordek R, Potemski P, Wakelam MJO, Romanska H, Berditchevski F

Expression of the tetraspanin CD151 is frequently upregulated in epithelial malignancies and correlates with poor prognosis. Here we report that CD151 is involved in regulation of the expression of fibroblast growth factor receptor 2 (FGFR2). Depletion of CD151 in breast cancer cells resulted in an increased level of FGFR2. Accordingly, an inverse correlation between CD151 and FGFR2 was observed in breast cancer tissues. CD151-dependent regulation of the FGFR2 expression relies on post-transcriptional mechanisms involving HuR/ELAVL1, a multifunctional RNA binding protein, and the assembly of processing bodies (P-bodies). Depletion of CD151 correlated with inhibition of PKC, a well-established downstream target of CD151. Accordingly, the levels of dialcylglycerol species were decreased in CD151-negative cells, and inhibition of PKC resulted in the increased expression of FGFR2. Whilst expression of FGFR2 itself did not correlate with any of the clinicopathological data, the FGFR2-/CD151+ patients are more likely to develop lymph node metastasis. Conversely, FGFR2-/CD151- patients demonstrated better overall survival. These results illustrate functional interdependency between CD151 complexes and FGFR2 and suggest a previously unsuspected role of CD151 in breast tumourigenesis.

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Journal of cell science, , 1477-9137, , 2018

PMID: 30257985


FastQ Screen: A tool for multi-genome mapping and quality control.
Wingett SW, Andrews S

DNA sequencing analysis typically involves mapping reads to just one reference genome. Mapping against multiple genomes is necessary, however, when the genome of origin requires confirmation. Mapping against multiple genomes is also advisable for detecting contamination or for identifying sample swaps which, if left undetected, may lead to incorrect experimental conclusions. Consequently, we present FastQ Screen, a tool to validate the origin of DNA samples by quantifying the proportion of reads that map to a panel of reference genomes. FastQ Screen is intended to be used routinely as a quality control measure and for analysing samples in which the origin of the DNA is uncertain or has multiple sources.

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F1000Research, 7, 2046-1402, 1338, 2018

PMID: 30254741


Open Access

Genome organization and chromatin analysis identify transcriptional downregulation of insulin-like growth factor signaling as a hallmark of aging in developing B cells.
Koohy H, Bolland DJ, Matheson LS, Schoenfelder S, Stellato C, Dimond A, Várnai C, Chovanec P, Chessa T, Denizot J, Manzano Garcia R, Wingett SW, Freire-Pritchett P, Nagano T, Hawkins P, Stephens L, Elderkin S, Spivakov M, Fraser P, Corcoran AE, Varga-Weisz PD

Aging is characterized by loss of function of the adaptive immune system, but the underlying causes are poorly understood. To assess the molecular effects of aging on B cell development, we profiled gene expression and chromatin features genome-wide, including histone modifications and chromosome conformation, in bone marrow pro-B and pre-B cells from young and aged mice.

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Genome biology, 19, 1474-760X, 126, 2018

PMID: 30180872


Open Access

Genetic regulation of antibody responsiveness to immunization in substrains of BALB/c mice.
Poyntz HC, Jones A, Jauregui R, Young W, Gestin A, Mooney A, Lamiable O, Altermann E, Schmidt A, Gasser O, Weyrich L, Jolly CJ, Linterman MA, Le Gros G, Hawkins ED, Forbes-Blom E

Antibody-mediated immunity is highly protective against disease. The majority of current vaccines confer protection through humoral immunity, but there is high variability in responsiveness across populations. Identifying immune mechanisms that mediate low antibody responsiveness may provide potential strategies to boost vaccine efficacy. Here, we report diverse antibody responsiveness to unadjuvanted as well as adjuvanted immunization in substrains of BALB/c mice, resulting in high and low antibody response phenotypes. Furthermore, these antibody phenotypes were not affected by changes in environmental factors such as the gut microbiota composition. Antigen-specific B cells following immunization had a marked difference in capability to class-switch, resulting in perturbed IgG isotype antibody production. In vitro, a B cell intrinsic defect in the regulation of class-switch recombination was identified in mice with low IgG antibody production. Whole genome sequencing identified polymorphisms associated with the magnitude of antibody produced, and we propose candidate genes that may regulate isotype class-switching capability. This study highlights that mice sourced from different vendors can have significantly altered humoral immune response profiles, and provides a resource to interrogate genetic regulators of antibody responsiveness. Together these results further our understanding of immune heterogeneity and suggest additional research on the genetic influences of adjuvanted vaccine strategies is warranted for enhancing vaccine efficacy. This article is protected by copyright. All rights reserved.

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Immunology and cell biology, , 1440-1711, , 2018

PMID: 30152893


Targeting IKKβ in Cancer: Challenges and Opportunities for the Therapeutic Utilisation of IKKβ Inhibitors.
Prescott JA, Cook SJ

Deregulated NF-κB signalling is implicated in the pathogenesis of numerous human inflammatory disorders and malignancies. Consequently, the NF-κB pathway has attracted attention as an attractive therapeutic target for drug discovery. As the primary, druggable mediator of canonical NF-κB signalling the IKKβ protein kinase has been the historical focus of drug development pipelines. Thousands of compounds with activity against IKKβ have been characterised, with many demonstrating promising efficacy in pre-clinical models of cancer and inflammatory disease. However, severe on-target toxicities and other safety concerns associated with systemic IKKβ inhibition have thus far prevented the clinical approval of any IKKβ inhibitors. This review will discuss the potential reasons for the lack of clinical success of IKKβ inhibitors to date, the challenges associated with their therapeutic use, realistic opportunities for their future utilisation, and the alternative strategies to inhibit NF-κB signalling that may overcome some of the limitations associated with IKKβ inhibition.

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Cells, 7, 2073-4409, , 2018

PMID: 30142927


Open Access

Paths to expansion: Differential requirements of IRF4 in CD8 T-cell expansion driven by antigen and homeostatic cytokines.
Lugli E, Brummelman J, Pilipow K, Roychoudhuri R

Interferon regulatory factor 4 (IRF4) regulates the clonal expansion and metabolic activity of activated T cells, but the precise context and mechanisms of its function in these processes are unclear. In this issue of the European Journal of Immunology, Miyakoda et al. [Eur. J. Immunol. 2018. 48: 1319-1328] show that IRF4 is required for activation and expansion of naïve and memory CD8 T cells driven by T-cell receptor (TCR) signaling, but dispensable for memory CD8 T-cell maintenance and homeostatic proliferation driven by homeostatic cytokines. The authors show that the function of IRF4 in CD8 T-cell expansion is partially dependent upon activation of the PI3K/AKT pathway through direct or indirect attenuation of PTEN expression. These data shed light upon the differential intracellular pathways required for naïve and memory T cells to respond to self-antigens and/or homeostatic cytokines, and highlight the potential translational relevance of these findings in the context of immune reconstitution such as following allogeneic stem cell transplantation.

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European journal of immunology, 48, 1521-4141, 1281-1284, 2018

PMID: 30133745


MS-based lipidomics of human blood plasma - a community-initiated position paper to develop accepted guidelines.
Burla B, Arita M, Arita M, Bendt AK, Cazenave-Gassiot A, Dennis EA, Ekroos K, Han X, Ikeda K, Liebisch G, Lin MK, Loh TP, Meikle PJ, Orešič M, Quehenberger O, Shevchenko A, Torta F, Wakelam MJO, Wheelock CE, Wenk MR

Human blood is a self-regenerating, lipid-rich biologic fluid that is routinely collected in hospital settings. The inventory of lipid molecules found in blood plasma (plasma lipidome) offers insights into individual metabolism and physiology in health and disease. Disturbances in lipid metabolism also occur in conditions that are not directly linked to lipid metabolism; therefore, plasma lipidomics based on mass spectrometry (MS) is an emerging tool in an array of clinical diagnostics and disease management. However, challenges exist in the translation of such lipidomic data to clinical applications. These relate to the reproducibility, accuracy, and precision of lipid quantitation, study design, sample handling, and data sharing. This position paper emerged from a workshop that initiated a community-led process to elaborate and define a set of generally accepted guidelines for quantitative MS-based lipidomics of blood plasma or serum, with harmonization of data acquired on different instrumentation platforms in independent laboratories across laboratories as an ultimate goal. We hope that other fields may benefit from and follow such a precedent.

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Journal of lipid research, , 1539-7262, , 2018

PMID: 30115755


Open Access

Transcriptome analysis of infected and bystander type 2 alveolar epithelial cells during influenza A virus infection reveals Wnt pathway downregulation.
Hancock AS, Stairiker CJ, Boesteanu AC, Monzón-Casanova E, Lukasiak S, Mueller YM, Stubbs AP, Garcia-Sastre A, Turner M, Katsikis PD

Influenza virus outbreaks remain a serious threat to public health. Greater understanding of how cells targeted by the virus respond to the infection can provide insight into the pathogenesis of disease. Here we examined the transcriptional profile of infected and uninfected type 2 alveolar epithelial cells (AEC) in the lungs of influenza virus infected mice. We show for the first time the unique gene expression profiles induced by the infection of AEC as well as the transcriptional response of uninfected bystander cells. This work allows us to distinguish the direct and indirect effects of infection at the cellular level. Transcriptome analysis revealed that although directly infected and bystander AEC from infected animals shared many transcriptome changes when compared to AEC from uninfected animals, directly infected cells compared to bystander uninfected AEC produce more interferon and express lower Wnt signaling associated transcripts, while concurrently expressing more transcripts associated with cell death pathways. The Wnt signaling pathway was downregulated in both infected AEC and infected human lung epithelial A549 cells. Wnt signaling did not affect type I and III interferon production by infected A549 cells. Our results reveal unique transcriptional changes that occur within infected AEC and show that influenza virus downregulates Wnt signaling. In light of recent findings that Wnt signaling is essential for lung epithelial stem cells, our findings reveal a mechanism by which influenza virus may affect host lung repair. Influenza virus infection remains a major public health problem. Utilizing a recombinant green fluorescent protein expressing influenza virus we compared the in vivo transcriptomes of directly infected and uninfected bystander cells from infected mouse lungs and discovered many pathways uniquely regulated in each population. The Wnt signaling pathway was downregulated in directly infected cells and was shown to affect virus but not interferon production. Our study is the first to discern the in vivo transcriptome changes induced by direct viral infection as compared to mere exposure to the lung inflammatory milieu and highlight the downregulation of Wnt signaling. This downregulation has important implications for understanding influenza virus pathogenesis as Wnt signaling is critical for lung epithelial stem cells and lung epithelial cell differentiation. Our findings reveal a mechanism by which influenza virus may affect host lung repair and suggest interventions that prevent damage or accelerate recovery of the lung.

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Journal of virology, , 1098-5514, , 2018

PMID: 30111569


C. elegans Eats Its Own Intestine to Make Yolk Leading to Multiple Senescent Pathologies.
Ezcurra M, Benedetto A, Sornda T, Gilliat AF, Au C, Zhang Q, van Schelt S, Petrache AL, Wang H, de la Guardia Y, Bar-Nun S, Tyler E, Wakelam MJ, Gems D

Aging (senescence) is characterized by the development of numerous pathologies, some of which limit lifespan. Key to understanding aging is discovery of the mechanisms (etiologies) that cause senescent pathology. In C. elegans, a major senescent pathology of unknown etiology is atrophy of its principal metabolic organ, the intestine. Here we identify a cause of not only this pathology but also of yolky lipid accumulation and redistribution (a form of senescent obesity): autophagy-mediated conversion of intestinal biomass into yolk. Inhibiting intestinal autophagy or vitellogenesis rescues both visceral pathologies and can also extend lifespan. This defines a disease syndrome leading to multimorbidity and contributing to late-life mortality. Activation of gut-to-yolk biomass conversion by insulin/IGF-1 signaling (IIS) promotes reproduction and senescence. This illustrates how major, IIS-promoted senescent pathologies in C. elegans can originate not from damage accumulation but from direct effects of futile, continued action of a wild-type biological program (vitellogenesis).

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Current biology : CB, , 1879-0445, , 2018

PMID: 30100339


Open Access

PI3Kδ hyper-activation promotes development of B cells that exacerbate Streptococcus pneumoniae infection in an antibody-independent manner.
Stark AK, Chandra A, Chakraborty K, Alam R, Carbonaro V, Clark J, Sriskantharajah S, Bradley G, Richter AG, Banham-Hall E, Clatworthy MR, Nejentsev S, Hamblin JN, Hessel EM, Condliffe AM, Okkenhaug K

Streptococcus pneumoniae is a major cause of pneumonia and a leading cause of death world-wide. Antibody-mediated immune responses can confer protection against repeated exposure to S. pneumoniae, yet vaccines offer only partial protection. Patients with Activated PI3Kδ Syndrome (APDS) are highly susceptible to S. pneumoniae. We generated a conditional knock-in mouse model of this disease and identify a CD19B220 B cell subset that is induced by PI3Kδ signaling, resides in the lungs, and is correlated with increased susceptibility to S. pneumoniae during early phases of infection via an antibody-independent mechanism. We show that an inhaled PI3Kδ inhibitor improves survival rates following S. pneumoniae infection in wild-type mice and in mice with activated PI3Kδ. These results suggest that a subset of B cells in the lung can promote the severity of S. pneumoniae infection, representing a potential therapeutic target.

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Nature communications, 9, 2041-1723, 3174, 2018

PMID: 30093657


Open Access