Position-effect variegation revisited: HUSHing up heterochromatin in human cells.

Much of what we understand about heterochromatin formation in mammals has been extrapolated from forward genetic screens for modifiers of position-effect variegation (PEV) in the fruit fly Drosophila melanogaster. The recent identification of the HUSH (Human Silencing Hub) complex suggests that more recent evolutionary developments contribute to the mechanisms underlying PEV in human cells. Although HUSH-mediated repression also involves heterochromatin spreading through the reading and writing of the repressive H3K9me3 histone modification, clear orthologues of HUSH subunits are not found in Drosophila but are conserved in vertebrates. Here we compare the insights into the mechanisms of PEV derived from genetic screens in the fly, the mouse and in human cells, review what is currently known about the HUSH complex and discuss the implications of HUSH-mediated silencing for viral latency. Future studies will provide mechanistic insight into HUSH complex function and reveal the relationship between HUSH and other epigenetic silencing complexes.This work was funded by a Wellcome Trust Principal Research Fellowship to P.J.L. (084957/Z/08/Z) and a Wellcome Trust PhD studentship to I.A.T. The Cambridge Institute for Medical Research is in receipt of a Wellcome Trust Strategic Award.This is the author accepted manuscript. The final version is available from Wiley via http://dx.doi.org/10.1002/bies.20150018

Diagnosing GORD in Respiratory Medicine

Gastroesophageal reflux disease (GORD) is increasing in prevalence and is highly associated with several lung diseases such as asthma and COPD. Current diagnostic methods are imperfect, being insensitive, non-specific, expensive, or invasive. An accurate diagnosis of GORD can aid effective treatment and have significant clinical impact. Novel methods such as exhaled breath condensate analysis and electronic nose technology have the potential to improve the accuracy of diagnosing GORD

ATF7IP-Mediated Stabilization of the Histone Methyltransferase SETDB1 Is Essential for Heterochromatin Formation by the HUSH Complex

The histone methyltransferase SETDB1 plays a central role in repressive chromatin processes, but the functional requirement for its binding partner ATF7IP has remained enigmatic. Here, we show that ATF7IP is essential for SETDB1 stability: nuclear SETDB1 protein is degraded by the proteasome upon ablation of ATF7IP. As a result, ATF7IP is critical for repression that requires H3K9 trimethylation by SETDB1, including transgene silencing by the HUSH complex. Furthermore, we show that loss of ATF7IP phenocopies loss of SETDB1 in genome-wide assays. ATF7IP and SETDB1 knockout cells exhibit near-identical defects in the global deposition of H3K9me3, which results in similar dysregulation of the transcriptome. Overall, these data identify a critical functional role for ATF7IP in heterochromatin formation by regulating SETDB1 abundance in the nucleus.This work was supported by the Wellcome Trust through a Principal Research Fellowship to P.J.L. (101835/Z/13/Z) and a Ph.D. studentship to I.A.T. The CIMR is in receipt of a Wellcome Trust strategic award.This is the final version of the article. It first appeared from Elsevier (Cell Press) via https://doi.org/10.1016/j.celrep.2016.09.05

A Genetic Screen Identifies a Critical Role for the WDR81-WDR91 Complex in the Trafficking and Degradation of Tetherin.

Tetherin (BST2/CD317) is a viral restriction factor that anchors enveloped viruses to host cells and limits viral spread. The HIV-1 Vpu accessory protein counteracts tetherin by decreasing its cell surface expression and targeting it for ubiquitin-dependent endolysosomal degradation. Although the Vpu-mediated downregulation of tetherin has been extensively studied, the molecular details are not completely elucidated. We therefore used a forward genetic screen in human haploid KBM7 cells to identify novel genes required for tetherin trafficking. Our screen identified WDR81 as a novel gene required for tetherin trafficking and degradation in both the presence and absence of Vpu. WDR81 is a BEACH-domain containing protein that is also required for the degradation of EGF-stimulated epidermal growth factor receptor (EGFR) and functions in a complex with the WDR91 protein. In the absence of WDR81 the endolysosomal compartment appears swollen, with enlarged early and late endosomes and reduced delivery of endocytosed dextran to cathepsin-active lysosomes. Our data suggest a role for the WDR81-WDR91 complex in the fusion of endolysosomal compartments and the absence of WDR81 leads to impaired receptor trafficking and degradation.This work was supported by the Wellcome Trust, through a Principal Research Fellowship to PJL (084957/Z/08/Z) and Ph.D studentship to RR (079895/Z/06/Z), by MRC research grant MR/M010007/1 to JPL and by a BBSRC industrial CASE studentship with GSK Research and Development Ltd to LJD. The CIMR is in receipt of a Wellcome Trust strategic award 100140.This is the final version of the article. It first appeared from Wiley via https://doi.org/10.1111/tra.1240

Dynamic Bayesian belief network to model the development of walking and cycling schemes

This paper aims to describe a model which represents the formulation of decision-making processes (over a number of years) affecting the step-changes of walking and cycling (WaC) schemes. These processes can be seen as being driven by a number of causal factors, many of which are associated with the attitudes of a variety of factors, in terms of both determining whether any scheme will be implemented and, if it is implemented, the extent to which it is used. The outputs of the model are pathways as to how the future might unfold (in terms of a number of future time steps) with respect to specific pedestrian and cyclist schemes. The transitions of the decision making processes are formulated using a qualitative simulation method, which describes the step-changes of the WaC scheme development. In this article a Bayesian belief network (BBN) theory is extended to model the influence between and within factors in the dynamic decision making process

GENE SILENCING. Epigenetic silencing by the HUSH complex mediates position-effect variegation in human cells.

Forward genetic screens in Drosophila melanogaster for modifiers of position-effect variegation have revealed the basis of much of our understanding of heterochromatin. We took an analogous approach to identify genes required for epigenetic repression in human cells. A nonlethal forward genetic screen in near-haploid KBM7 cells identified the HUSH (human silencing hub) complex, comprising three poorly characterized proteins, TASOR, MPP8, and periphilin; this complex is absent from Drosophila but is conserved from fish to humans. Loss of HUSH components resulted in decreased H3K9me3 both at endogenous genomic loci and at retroviruses integrated into heterochromatin. Our results suggest that the HUSH complex is recruited to genomic loci rich in H3K9me3, where subsequent recruitment of the methyltransferase SETDB1 is required for further H3K9me3 deposition to maintain transcriptional silencing.This work was supported by a Wellcome Trust Principal Research Fellowship to P.J.L. (084957/Z/08/Z) and studentship to I.A.T., an MRC Centenary Award to R.T.T., and the Cambridge Biomedical Research Centre (UK). The CIMR is in receipt of a Wellcome Trust Strategic Award.This is the author accepted manuscript. The final version is available from AAAS via http://dx.doi.org/10.1126/science.aaa722

Identification of genes expressed by immune cells of the colon that are regulated by colorectal cancer-associated variants.

A locus on human chromosome 11q23 tagged by marker rs3802842 was associated with colorectal cancer (CRC) in a genome-wide association study; this finding has been replicated in case-control studies worldwide. In order to identify biologic factors at this locus that are related to the etiopathology of CRC, we used microarray-based target selection methods, coupled to next-generation sequencing, to study 103 kb at the 11q23 locus. We genotyped 369 putative variants from 1,030 patients with CRC (cases) and 1,061 individuals without CRC (controls) from the Ontario Familial Colorectal Cancer Registry. Two previously uncharacterized genes, COLCA1 and COLCA2, were found to be co-regulated genes that are transcribed from opposite strands. Expression levels of COLCA1 and COLCA2 transcripts correlate with rs3802842 genotypes. In colon tissues, COLCA1 co-localizes with crystalloid granules of eosinophils and granular organelles of mast cells, neutrophils, macrophages, dendritic cells and differentiated myeloid-derived cell lines. COLCA2 is present in the cytoplasm of normal epithelial, immune and other cell lineages, as well as tumor cells. Tissue microarray analysis demonstrates the association of rs3802842 with lymphocyte density in the lamina propria (p = 0.014) and levels of COLCA1 in the lamina propria (p = 0.00016) and COLCA2 (tumor cells, p = 0.0041 and lamina propria, p = 6 × 10(-5)). In conclusion, genetic, expression and immunohistochemical data implicate COLCA1 and COLCA2 in the pathogenesis of colon cancer. Histologic analyses indicate the involvement of immune pathways