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

IMPLICON: an ultra-deep sequencing method to uncover DNA methylation at imprinted regions.
Klobučar T, Kreibich E, Krueger F, Arez M, Pólvora-Brandão D, von Meyenn F, da Rocha ST, Eckersley-Maslin M

Genomic imprinting is an epigenetic phenomenon leading to parental allele-specific expression. Dosage of imprinted genes is crucial for normal development and its dysregulation accounts for several human disorders. This unusual expression pattern is mostly dictated by differences in DNA methylation between parental alleles at specific regulatory elements known as imprinting control regions (ICRs). Although several approaches can be used for methylation inspection, we lack an easy and cost-effective method to simultaneously measure DNA methylation at multiple imprinted regions. Here, we present IMPLICON, a high-throughput method measuring DNA methylation levels at imprinted regions with base-pair resolution and over 1000-fold coverage. We adapted amplicon bisulfite-sequencing protocols to design IMPLICON for ICRs in adult tissues of inbred mice, validating it in hybrid mice from reciprocal crosses for which we could discriminate methylation profiles in the two parental alleles. Lastly, we developed a human version of IMPLICON and detected imprinting errors in embryonic and induced pluripotent stem cells. We also provide rules and guidelines to adapt this method for investigating the DNA methylation landscape of any set of genomic regions. In summary, IMPLICON is a rapid, cost-effective and scalable method, which could become the gold standard in both imprinting research and diagnostics.

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Nucleic acids research , 1 , 1 ,

PMID: 32621604

Replicative aging is associated with loss of genetic heterogeneity from extrachromosomal circular DNA in Saccharomyces cerevisiae.
Prada-Luengo I, Møller HD, Henriksen RA, Gao Q, Larsen CE, Alizadeh S, Maretty L, Houseley J, Regenberg B

Circular DNA can arise from all parts of eukaryotic chromosomes. In yeast, circular ribosomal DNA (rDNA) accumulates dramatically as cells age, however little is known about the accumulation of other chromosome-derived circles or the contribution of such circles to genetic variation in aged cells. We profiled circular DNA in Saccharomyces cerevisiae populations sampled when young and after extensive aging. Young cells possessed highly diverse circular DNA populations but 94% of the circular DNA were lost after ∼15 divisions, whereas rDNA circles underwent massive accumulation to >95% of circular DNA. Circles present in both young and old cells were characterized by replication origins including circles from unique regions of the genome and repetitive regions: rDNA and telomeric Y' regions. We further observed that circles can have flexible inheritance patterns: [HXT6/7circle] normally segregates to mother cells but in low glucose is present in up to 50% of cells, the majority of which must have inherited this circle from their mother. Interestingly, [HXT6/7circle] cells are eventually replaced by cells carrying stable chromosomal HXT6 HXT6/7 HXT7 amplifications, suggesting circular DNAs are intermediates in chromosomal amplifications. In conclusion, the heterogeneity of circular DNA offers flexibility in adaptation, but this heterogeneity is remarkably diminished with age.

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Nucleic acids research , 1 , 1 ,

PMID: 32609810

Open Access

Membrane characteristics tune activities of endosomal and autophagic human VPS34 complexes.
Ohashi Y, Tremel S, Masson GR, McGinney L, Boulanger J, Rostislavleva K, Johnson CM, Niewczas I, Clark J, Williams RL

The lipid kinase VPS34 orchestrates diverse processes, including autophagy, endocytic sorting, phagocytosis, anabolic responses and cell division. VPS34 forms various complexes that help adapt it to specific pathways, with complexes I and II being the most prominent ones. We found that physicochemical properties of membranes strongly modulate VPS34 activity. Greater unsaturation of both substrate and non-substrate lipids, negative charge and curvature activate VPS34 complexes, adapting them to their cellular compartments. Hydrogen/deuterium exchange mass spectrometry (HDX-MS) of complexes I and II on membranes elucidated structural determinants that enable them to bind membranes. Among these are the Barkor/ATG14L autophagosome targeting sequence (BATS), which makes autophagy-specific complex I more active than the endocytic complex II, and the Beclin1 BARA domain. Interestingly, even though Beclin1 BARA is common to both complexes, its membrane-interacting loops are critical for complex II, but have only a minor role for complex I.

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eLife , 9 , 1 ,

PMID: 32602837

Correction to: DNA methylation changes during preimplantation development reveal interspecies differences and reprogramming events at imprinted genes.
Ivanova E, Canovas S, Garcia-Martínez S, Romar R, Lopes JS, Rizos D, Sanchez-Calabuig MJ, Krueger F, Andrews S, Perez-Sanz F, Kelsey G, Coy P

An amendment to this paper has been published and can be accessed via the original article.

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Clinical epigenetics , 12 , 1 ,

PMID: 32600441

Open Access

The receptor PTPRU is a redox sensitive pseudophosphatase.
Hay IM, Fearnley GW, Rios P, Köhn M, Sharpe HJ, Deane JE

The receptor-linked protein tyrosine phosphatases (RPTPs) are key regulators of cell-cell communication through the control of cellular phosphotyrosine levels. Most human RPTPs possess an extracellular receptor domain and tandem intracellular phosphatase domains: comprising an active membrane proximal (D1) domain and an inactive distal (D2) pseudophosphatase domain. Here we demonstrate that PTPRU is unique amongst the RPTPs in possessing two pseudophosphatase domains. The PTPRU-D1 displays no detectable catalytic activity against a range of phosphorylated substrates and we show that this is due to multiple structural rearrangements that destabilise the active site pocket and block the catalytic cysteine. Upon oxidation, this cysteine forms an intramolecular disulphide bond with a vicinal "backdoor" cysteine, a process thought to reversibly inactivate related phosphatases. Importantly, despite the absence of catalytic activity, PTPRU binds substrates of related phosphatases strongly suggesting that this pseudophosphatase functions in tyrosine phosphorylation by competing with active phosphatases for the binding of substrates.

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Nature communications , 11 , 1 ,

PMID: 32591542

BioModels Parameters: a treasure trove of parameter values from published systems biology models.
Glont M, Arankalle C, Tiwari K, Nguyen TVN, Hermjakob H, Malik Sheriff RS

One of the major bottlenecks in building systems biology models is identification and estimation of model parameters for model calibration. Searching for model parameters from published literature and models is an essential, yet laborious task.

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Bioinformatics (Oxford, England) , 1 , 1 ,

PMID: 32573648

Epigenetic priming by Dppa2 and 4 in pluripotency facilitates multi-lineage commitment.
Eckersley-Maslin MA, Parry A, Blotenburg M, Krueger C, Ito Y, Franklin VNR, Narita M, D'Santos CS, Reik W

How the epigenetic landscape is established in development is still being elucidated. Here, we uncover developmental pluripotency associated 2 and 4 (DPPA2/4) as epigenetic priming factors that establish a permissive epigenetic landscape at a subset of developmentally important bivalent promoters characterized by low expression and poised RNA-polymerase. Differentiation assays reveal that Dppa2/4 double knockout mouse embryonic stem cells fail to exit pluripotency and differentiate efficiently. DPPA2/4 bind both H3K4me3-marked and bivalent gene promoters and associate with COMPASS- and Polycomb-bound chromatin. Comparing knockout and inducible knockdown systems, we find that acute depletion of DPPA2/4 results in rapid loss of H3K4me3 from key bivalent genes, while H3K27me3 is initially more stable but lost following extended culture. Consequently, upon DPPA2/4 depletion, these promoters gain DNA methylation and are unable to be activated upon differentiation. Our findings uncover a novel epigenetic priming mechanism at developmental promoters, poising them for future lineage-specific activation.

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Nature structural & molecular biology , 1 , 1 ,

PMID: 32572255

Mechanical stretching changes cross-linking and glycation levels in the collagen of mouse tail tendon.
Stammers M, Niewczas IS, Segonds-Pichon A, Clark J

Collagen I is a major tendon protein whose polypeptide chains are linked by covalent cross-links. It is unknown how the cross-linking contributes to the mechanical properties of tendon or whether cross-linking changes in response to stretching or relaxation. Since their discovery, imine bonds within collagen have been recognized as being important in both cross-link formation and collagen structure. They are often described as acidic or thermally labile, but no evidence is available from direct measurements of cross-link levels whether these bonds contribute to the mechanical properties of collagen. Here, we used MS to analyze these imine bonds after reduction with sodium borohydride while under tension and found that their levels are altered in stretched tendon. We studied the changes in cross-link bonding in tail tendon from 11-week-old C57Bl/6 mice at 4% physical strain, at 10% strain, and at breaking point. The cross-links hydroxy-lysino-norleucine (HLNL), dihydroxy-lysino-norleucine (DHLNL), and lysino-norleucine (LNL) increased or decreased depending on the specific cross-link and amount of mechanical strain. We also noted a decrease in glycated lysine residues in collagen, indicating that the imine formed between circulating glucose and lysine is also stress-labile. We also carried out mechanical testing, including cyclic testing at 4% strain, stress relaxation tests, and stress-strain profiles taken at breaking point, both with and without sodium borohydride reduction. The results from both the MS studies and mechanical testing provide insights into the chemical changes during tendon stretching and directly link these chemical changes to functional collagen properties.

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The Journal of biological chemistry , 1 , 1 ,

PMID: 32546479

Open Access

High-Throughput Quantitative RT-PCR in Single and Bulk C. elegans Samples Using Nanofluidic Technology.
Chauve L, Le Pen J, Hodge F, Todtenhaupt P, Biggins L, Miska EA, Andrews S, Casanueva O

This paper presents a high-throughput reverse transcription quantitative PCR (RT-qPCR) assay for Caenorhabditis elegans that is fast, robust, and highly sensitive. This protocol obtains precise measurements of gene expression from single worms or from bulk samples. The protocol presented here provides a novel adaptation of existing methods for complementary DNA (cDNA) preparation coupled to a nanofluidic RT-qPCR platform. The first part of this protocol, named 'Worm-to-CT', allows cDNA production directly from nematodes without the need for prior mRNA isolation. It increases experimental throughput by allowing the preparation of cDNA from 96 worms in 3.5 h. The second part of the protocol uses existing nanofluidic technology to run high-throughput RT-qPCR on the cDNA. This paper evaluates two different nanofluidic chips: the first runs 96 samples and 96 targets, resulting in 9,216 reactions in approximately 1.5 days of benchwork. The second chip type consists of six 12 x 12 arrays, resulting in 864 reactions. Here, the Worm-to-CT method is demonstrated by quantifying mRNA levels of genes encoding heat shock proteins from single worms and from bulk samples. Provided is an extensive list of primers designed to amplify processed RNA for the majority of coding genes within the C. elegans genome.

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Journal of visualized experiments : JoVE , 1 , 159 ,

PMID: 32538915

Worm-align and Worm_CP, Two Open-Source Pipelines for Straightening and Quantification of Fluorescence Image Data Obtained from Caenorhabditis elegans.
Okkenhaug H, Chauve L, Masoudzadeh F, Okkenhaug L, Casanueva O

An issue often encountered when acquiring image data from fixed or anesthetized C. elegans is that worms cross and cluster with their neighbors. This problem is aggravated with increasing density of worms and creates challenges for imaging and quantification. We developed a FIJI-based workflow, Worm-align, that can be used to generate single- or multi-channel montages of user-selected, straightened and aligned worms from raw image data of C. elegans. Worm-align is a simple and user-friendly workflow that does not require prior training of either the user or the analysis algorithm. Montages generated with Worm-align can aid the visual inspection of worms, their classification and representation. In addition, the output of Worm-align can be used for subsequent quantification of fluorescence intensity in single worms, either in FIJI directly, or in other image analysis software platforms. We demonstrate this by importing the Worm-align output into Worm_CP, a pipeline that uses the open-source CellProfiler software. CellProfiler's flexibility enables the incorporation of additional modules for high-content screening. As a practical example, we have used the pipeline on two datasets: the first dataset are images of heat shock reporter worms that express green fluorescent protein (GFP) under the control of the promoter of a heat shock inducible gene hsp-70, and the second dataset are images obtained from fixed worms, stained for fat-stores with a fluorescent dye.

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Journal of visualized experiments : JoVE , 1 , 159 ,

PMID: 32538914

Axon Degeneration: Which Method to Choose?
Coleman MP

Axons are diverse. They have different lengths, different branching patterns, and different biological roles. Methods to study axon degeneration are also diverse. The result is a bewildering range of experimental systems in which to study mechanisms of axon degeneration, and it is difficult to extrapolate from one neuron type and one method to another. The purpose of this chapter is to help readers to do this and to choose the methods most appropriate for answering their particular research question.

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Methods in molecular biology (Clifton, N.J.) , 2143 , 1 ,

PMID: 32524468

BACH2 drives quiescence and maintenance of resting Treg cells to promote homeostasis and cancer immunosuppression.
Grant FM, Yang J, Nasrallah R, Clarke J, Sadiyah F, Whiteside SK, Imianowski CJ, Kuo P, Vardaka P, Todorov T, Zandhuis N, Patrascan I, Tough DF, Kometani K, Eil R, Kurosaki T, Okkenhaug K, Roychoudhuri R

Regulatory T (Treg) cell populations are composed of functionally quiescent resting Treg (rTreg) cells which differentiate into activated Treg (aTreg) cells upon antigen stimulation. How rTreg cells remain quiescent despite chronic exposure to cognate self- and foreign antigens is unclear. The transcription factor BACH2 is critical for early Treg lineage specification, but its function following lineage commitment is unresolved. Here, we show that BACH2 is repurposed following Treg lineage commitment and promotes the quiescence and long-term maintenance of rTreg cells. Bach2 is highly expressed in rTreg cells but is down-regulated in aTreg cells and during inflammation. In rTreg cells, BACH2 binds to enhancers of genes involved in aTreg differentiation and represses their TCR-driven induction by competing with AP-1 factors for DNA binding. This function promotes rTreg cell quiescence and long-term maintenance and is required for immune homeostasis and durable immunosuppression in cancer. Thus, BACH2 supports a "division of labor" between quiescent rTreg cells and their activated progeny in Treg maintenance and function, respectively.

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The Journal of experimental medicine , 217 , 9 ,

PMID: 32515782

A distal enhancer at risk locus 11q13.5 promotes suppression of colitis by T cells.
Nasrallah R, Imianowski CJ, Bossini-Castillo L, Grant FM, Dogan M, Placek L, Kozhaya L, Kuo P, Sadiyah F, Whiteside SK, Mumbach MR, Glinos D, Vardaka P, Whyte CE, Lozano T, Fujita T, Fujii H, Liston A, Andrews S, Cozzani A, Yang J, Mitra S, Lugli E, Chang HY, Unutmaz D, Trynka G, Roychoudhuri R

Genetic variations underlying susceptibility to complex autoimmune and allergic diseases are concentrated within noncoding regulatory elements termed enhancers. The functions of a large majority of disease-associated enhancers are unknown, in part owing to their distance from the genes they regulate, a lack of understanding of the cell types in which they operate, and our inability to recapitulate the biology of immune diseases in vitro. Here, using shared synteny to guide loss-of-function analysis of homologues of human enhancers in mice, we show that the prominent autoimmune and allergic disease risk locus at chromosome 11q13.5 contains a distal enhancer that is functional in CD4 regulatory T (T) cells and required for T-mediated suppression of colitis. The enhancer recruits the transcription factors STAT5 and NF-κB to mediate signal-driven expression of Lrrc32, which encodes the protein glycoprotein A repetitions predominant (GARP). Whereas disruption of the Lrrc32 gene results in early lethality, mice lacking the enhancer are viable but lack GARP expression in Foxp3 T cells, which are unable to control colitis in a cell-transfer model of the disease. In human T cells, the enhancer forms conformational interactions with the promoter of LRRC32 and enhancer risk variants are associated with reduced histone acetylation and GARP expression. Finally, functional fine-mapping of 11q13.5 using CRISPR-activation (CRISPRa) identifies a CRISPRa-responsive element in the vicinity of risk variant rs11236797 capable of driving GARP expression. These findings provide a mechanistic basis for association of the 11q13.5 risk locus with immune-mediated diseases and identify GARP as a potential target in their therapy.

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Nature , 1 , 1 ,

PMID: 32499651

Active and repressed biosynthetic gene clusters have spatially distinct chromosome states.
Nützmann HW, Doerr D, Ramírez-Colmenero A, Sotelo-Fonseca JE, Wegel E, Di Stefano M, Wingett SW, Fraser P, Hurst L, Fernandez-Valverde SL, Osbourn A

While colocalization within a bacterial operon enables coexpression of the constituent genes, the mechanistic logic of clustering of nonhomologous monocistronic genes in eukaryotes is not immediately obvious. Biosynthetic gene clusters that encode pathways for specialized metabolites are an exception to the classical eukaryote rule of random gene location and provide paradigmatic exemplars with which to understand eukaryotic cluster dynamics and regulation. Here, using 3C, Hi-C, and Capture Hi-C (CHi-C) organ-specific chromosome conformation capture techniques along with high-resolution microscopy, we investigate how chromosome topology relates to transcriptional activity of clustered biosynthetic pathway genes in Our analyses reveal that biosynthetic gene clusters are embedded in local hot spots of 3D contacts that segregate cluster regions from the surrounding chromosome environment. The spatial conformation of these cluster-associated domains differs between transcriptionally active and silenced clusters. We further show that silenced clusters associate with heterochromatic chromosomal domains toward the periphery of the nucleus, while transcriptionally active clusters relocate away from the nuclear periphery. Examination of chromosome structure at unrelated clusters in maize, rice, and tomato indicates that integration of clustered pathway genes into distinct topological domains is a common feature in plant genomes. Our results shed light on the potential mechanisms that constrain coexpression within clusters of nonhomologous eukaryotic genes and suggest that gene clustering in the one-dimensional chromosome is accompanied by compartmentalization of the 3D chromosome.

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Proceedings of the National Academy of Sciences of the United States of America , 1 , 1 ,

PMID: 32493747

LPS-treatment of bovine endometrial epithelial cells causes differential DNA methylation of genes associated with inflammation and endometrial function.
Jhamat N, Niazi A, Guo Y, Chanrot M, Ivanova E, Kelsey G, Bongcam-Rudloff E, Andersson G, Humblot P

Lipopolysaccharide (LPS) endotoxin stimulates pro-inflammatory pathways and is a key player in the pathological mechanisms involved in the development of endometritis. This study aimed to investigate LPS-induced DNA methylation changes in bovine endometrial epithelial cells (bEECs), which may affect endometrial function. Following in vitro culture, bEECs from three cows were either untreated (0) or exposed to 2 and 8 μg/mL LPS for 24 h.

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BMC genomics , 21 , 1 ,

PMID: 32493210

The dead phosphatases society: a review of the emerging roles of pseudophosphatases.
Reiterer V, Pawłowski K, Desrochers G, Pause A, Sharpe HJ, Farhan H

Phosphatases are a diverse family of enzymes, comprising at least 10 distinct protein folds. Like most other enzyme families, many have sequence variations that predict an impairment or loss of catalytic activity classifying them as pseudophosphatases. Research on pseudoenzymes is an emerging area of interest, with new biological functions repurposed from catalytically active relatives. Here, we provide an overview of the pseudophosphatases identified to date in all major phosphatase families. We will highlight the degeneration of the various catalytic sequence motifs and discuss the challenges associated with the experimental determination of catalytic inactivity. We will also summarize the role of pseudophosphatases in various diseases and discuss the major challenges and future directions in this field.

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The FEBS journal , 1 , 1 ,

PMID: 32484316

Loss of Protects Against the Deleterious Effects of Traumatic Brain Injury in .
Hill CS, Sreedharan J, Loreto A, Menon DK, Coleman MP

Traumatic brain injury is a major global cause of death and disability. Axonal injury is a major underlying mechanism of TBI and could represent a major therapeutic target. We provide evidence that targeting the axonal death pathway known as Wallerian degeneration improves outcome in a model of high impact trauma. This cell-autonomous neurodegenerative pathway is initiated following axon injury, and in Drosophila, involves activity of the E3 ubiquitin ligase . We demonstrate that a loss-of-function mutation in the gene rescues deleterious effects of a traumatic injury, including-improved functional outcomes, lifespan, survival of dopaminergic neurons, and retention of synaptic proteins. This data suggests that represents a potential therapeutic target in traumatic injury.

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Frontiers in neurology , 11 , 1 ,

PMID: 32477254

A distinctive epigenetic ageing profile in human granulosa cells.
Olsen KW, Castillo-Fernandez J, Zedeler A, Freiesleben NC, Bungum M, Chan AC, Cardona A, Perry JRB, Skouby SO, Borup R, Hoffmann ER, Kelsey G, Grøndahl ML

Does women's age affect the DNA methylation (DNAm) profile differently in mural granulosa cells (MGCs) from other somatic cells?

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Human reproduction (Oxford, England) , 1 , 1 ,

PMID: 32474592

TET1 and 5-Hydroxymethylation Preserve the Stem Cell State of Mouse Trophoblast.
Senner CE, Chrysanthou S, Burge S, Lin HY, Branco MR, Hemberger M

The ten-eleven translocation factor TET1 and its conferred epigenetic modification 5-hydroxymethylcytosine (5hmC) have important roles in maintaining the pluripotent state of embryonic stem cells (ESCs). We previously showed that TET1 is also essential to maintain the stem cell state of trophoblast stem cells (TSCs). Here, we establish an integrated panel of absolute 5hmC levels, genome-wide DNA methylation and hydroxymethylation patterns, transcriptomes, and TET1 chromatin occupancy in TSCs and differentiated trophoblast cells. We show that the combined presence of 5-methylcytosine (5mC) and 5hmC correlates with transcriptional activity of associated genes. Hypoxia can slow down the global loss of 5hmC that occurs upon differentiation of TSCs. Notably, unlike in ESCs and epiblast cells, most TET1-bound regions overlap with active chromatin marks and TFAP2C binding sites and demarcate putative trophoblast enhancer regions. These chromatin modification and occupancy patterns are highly informative to identify novel candidate regulators of the TSC state.

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Stem cell reports , 1 , 1 ,

PMID: 32442533

Open Access

Truncation of Pik3r1 causes severe insulin resistance uncoupled from obesity and dyslipidemia by increased energy expenditure.
Kwok A, Zvetkova I, Virtue S, Luijten I, Huang-Doran I, Tomlinson P, Bulger DA, West J, Murfitt S, Griffin J, Alam R, Hart D, Knox R, Voshol P, Vidal-Puig A, Jensen J, O'Rahilly S, Semple RK

Insulin signaling via phosphoinositide 3-kinase (PI3K) requires PIK3R1-encoded regulatory subunits. C-terminal PIK3R1 mutations cause SHORT syndrome, including lipodystrophy and insulin resistance (IR), surprisingly without fatty liver or metabolic dyslipidemia. We sought to investigate this discordance.

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Molecular metabolism , 1 , 1 ,

PMID: 32439336

Open Access

THP-1 macrophage cholesterol efflux is impaired by palmitoleate through Akt activation.
Marshall JD, Courage ER, Elliott RF, Fitzpatrick MN, Kim AD, Lopez-Clavijo AF, Woolfrey BA, Ouimet M, Wakelam MJO, Brown RJ

Lipoprotein lipase (LPL) is upregulated in atherosclerotic lesions and it may promote the progression of atherosclerosis, but the mechanisms behind this process are not completely understood. We previously showed that the phosphorylation of Akt within THP-1 macrophages is increased in response to the lipid hydrolysis products generated by LPL from total lipoproteins. Notably, the free fatty acid (FFA) component was responsible for this effect. In the present study, we aimed to reveal more detail as to how the FFA component may affect Akt signalling. We show that the phosphorylation of Akt within THP-1 macrophages increases with total FFA concentration and that phosphorylation is elevated up to 18 hours. We further show that specifically the palmitoleate component of the total FFA affects Akt phosphorylation. This is tied with changes to the levels of select molecular species of phosphoinositides. We further show that the total FFA component, and specifically palmitoleate, reduces apolipoprotein A-I-mediated cholesterol efflux, and that the reduction can be reversed in the presence of the Akt inhibitor MK-2206. Overall, our data support a negative role for the FFA component of lipoprotein hydrolysis products generated by LPL, by impairing macrophage cholesterol efflux via Akt activation.

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PloS one , 15 , 5 ,

PMID: 32437392

Open Access

Remodeling of light and dark zone follicular dendritic cells governs germinal center responses.
Pikor NB, Mörbe U, Lütge M, Gil-Cruz C, Perez-Shibayama C, Novkovic M, Cheng HW, Nombela-Arrieta C, Nagasawa T, Linterman MA, Onder L, Ludewig B

Efficient generation of germinal center (GC) responses requires directed movement of B cells between distinct microenvironments underpinned by specialized B cell-interacting reticular cells (BRCs). How BRCs are reprogrammed to cater to the developing GC remains unclear, and studying this process is largely hindered by incomplete resolution of the cellular composition of the B cell follicle. Here we used genetic targeting of Cxcl13-expressing cells to define the molecular identity of the BRC landscape. Single-cell transcriptomic analysis revealed that BRC subset specification was predetermined in the primary B cell follicle. Further topological remodeling of light and dark zone follicular dendritic cells required CXCL12-dependent crosstalk with B cells and dictated GC output by retaining B cells in the follicle and steering their interaction with follicular helper T cells. Together, our results reveal that poised BRC-defined microenvironments establish a feed-forward system that determines the efficacy of the GC reaction.

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Nature immunology , 1 , 1 ,

PMID: 32424359

Senescence blurs the line between innate and adaptive immune cells.
Quinn KM, Linterman MA

In Covre et al. and Pereira et al., the authors demonstrate the parallels between senescent NK cells and senescent CD8 T cells, and formalise the mechanism by which senescent CD8 T cells become more NK cell-like, through the action of sestrins.

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Immunology and cell biology , 1 , 1 ,

PMID: 32406096

DNA methylation changes during preimplantation development reveal inter-species differences and reprogramming events at imprinted genes.
Ivanova E, Canovas S, Garcia-Martínez S, Romar R, Lopes JS, Rizos D, Sanchez-Calabuig MJ, Krueger F, Andrews S, Perez-Sanz F, Kelsey G, Coy P

Preimplantation embryos experience profound resetting of epigenetic information inherited from the gametes. Genome-wide analysis at single-base resolution has shown similarities but also species differences between human and mouse preimplantation embryos in DNA methylation patterns and reprogramming. Here, we have extended such analysis to two key livestock species, the pig and the cow. We generated genome-wide DNA methylation and whole-transcriptome datasets from gametes to blastocysts in both species. In oocytes from both species, a distinctive bimodal methylation landscape is present, with hypermethylated domains prevalent over hypomethylated domains, similar to human, while in the mouse the proportions are reversed.An oocyte-like pattern of methylation persists in the cleavage stages, albeit with some reduction in methylation level, persisting to blastocysts in cow, while pig blastocysts have a highly hypomethylated landscape. In the pig, there was evidence of transient de novo methylation at the 8-16 cell stages of domains unmethylated in oocytes, revealing a complex dynamic of methylation reprogramming. The methylation datasets were used to identify germline differentially methylated regions (gDMRs) of known imprinted genes and for the basis of detection of novel imprinted loci. Strikingly in the pig, we detected a consistent reduction in gDMR methylation at the 8-16 cell stages, followed by recovery to the blastocyst stage, suggesting an active period of imprint stabilization in preimplantation embryos. Transcriptome analysis revealed absence of expression in oocytes of both species of ZFP57, a key factor in the mouse for gDMR methylation maintenance, but presence of the alternative imprint regulator ZNF445. In conclusion, our study reveals species differences in DNA methylation reprogramming and suggests that porcine or bovine models may be closer to human in key aspects than in the mouse model.

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Clinical epigenetics , 12 , 1 ,

PMID: 32393379

Open Access