Cardiovascular disease collectively accounts for a quarter of deaths worldwide. Genome-wide association studies across a range of cardiovascular traits and pathologies have highlighted the prevalence of common non-coding genetic variants within candidate loci. Here, we review genetic, epigenomic and molecular approaches to investigate the contribution of non-coding regulatory elements in cardiovascular biology. We then discuss recent insights on the emerging role of non-coding variation in predisposition to cardiovascular disease, with a focus on novel mechanistic examples from functional genomics studies. Lastly, we consider the clinical significance of these findings at present, and some of the current challenges facing the field.
CCN2 is a critical matricellular protein that is expressed in several cells with major implications in physiology and different pathologies. However, the transcriptional regulation of this gene remains obscure. We used the Encyclopaedia of DNA Elements browser (ENCODE) to visualise the region spanning from 300 kb upstream to the CCN2 start site in silico in order to identify enhancer regions that regulate transcription of this gene. Selection was based on three criteria associated with enhancer regions: 1) H3K4me1 and H3K27ac histone modifications, 2) DNase I hypersensitivity of chromatin and 3) inter-species conservation. Reporter constructs were created with sequences spanning each of the regions of interest placed upstream of an Hsp68 silent proximal promoter sequence in order to drive the expression of β-galactosidase transgene. Each of these constructs was subsequently used to create transgenic mice in which reporter gene production was assessed at the E15.5 developmental stage. Four functional enhancers were identified, with each driving distinct, tissue-specific patterns of transgene expression. An enhancer located -100 kb from the CCN2 transcription start site facilitated expression within vascular tissue. An enhancer -135 kb upstream of CCN2 drove expression within the articular chondrocytes of synovial joints. The other two enhancers, located at -198 kb and -229 kb, mediated transgene expression within dermal fibroblasts, however the most prevalent activity was found within hypertrophic chondrocytes and periosteal tissue, respectively. These findings suggest that the global expression of CCN2 during development results from the activity of several tissue-specific enhancer regions in addition to proximal regulatory elements that have previously been demonstrated to drive transcription of the gene during development.Electronic supplementary materialThe online version of this article (10.1007/s12079-017-0440-4) contains supplementary material, which is available to authorized users.
Background and purpose: MAPKs are among the most relevant signalling pathways involved in coordinating cell responses to different stimuli. In this group we can find p38MAPK, constituted by 4 different proteins with a high sequence homology: p38α, p38β, p38γ and p38δ. Despite the high homology, each member shows unique expression patterns and even exclusive functions. Thus, analysing protein-specific functions of MAPK members is necessary to unequivocally uncover the roles of this signalling pathway. Here, we propose to investigate the possible role of p38β (MAPK11) in the cell response to ionizing radiation (IR). Materials and methods: We have developed MAPK11/14 knockdown through shRNA and CRISPR interference gene perturbation approaches, and analysed the implication of this MAPKs in cell response to ionizing radiation in A549, HCT-116 and MCF-7 cancer cell lines. Specifically, we analysed IR toxicity by clonogenic assays; DNA damage response activity by immunocytochemistry; apoptosis and cell cycle by flow cytometry (Annexin V and propidium iodide, respectively); DNA repair by comet assay; and senescence induction by both X-Gal staining and gene expression of senescence-associated genes by RT-qPCR. Results: Our findings demonstrate a critical role of MAPK11 in the cellular response to IR by controlling the associated senescent phenotype, and without observable effects on DDR, apoptosis, cell cycle or DNA damage repair. Conclusion: Our results highlight MAPK11 as a novel mediator of the cellular response to ionising radiation through the control exerted onto IR-associated senescence.
Changes in gene regulation have long been thought to underlie most phenotypic differences between species. Subterranean rodents, and in particular the naked mole rat, have attracted substantial attention due to their proposed phenotypic adaptations, which include hypoxia tolerance, metabolic changes and cancer resistance. However, it is largely unknown what regulatory changes may associate with these phenotypic traits, and whether these are unique to the naked mole rat, the mole rat clade or also present in other mammals. Here, we investigate regulatory evolution in heart and liver from two African mole rat species and two rodent outgroups using genome wide epigenomic profiling. First, we adapted and applied a phylogenetic modeling approach to quantitatively compare epigenomic signals at orthologous regulatory elements, and identified thousands of promoter and enhancer regions with differential epigenomic activity in mole rats. These elements associate with known mole rat adaptation in metabolic and functional pathways, and suggest candidate genetic loci that may underlie mole rat innovations. Second, we evaluated ancestral and species specific regulatory changes in the study phylogeny, and report several candidate pathways experiencing stepwise remodeling during the evolution of mole rats (such as the insulin and hypoxia response pathways). Third, we report non orthologous regulatory elements overlap with lineage specific repetitive elements and appear to modify metabolic pathways by rewiring of HNF4 and RAR/RXR transcription factor binding sites in mole rats. These comparative analyses reveal how mole-rat regulatory evolution informs previously reported phenotypic adaptations. Moreover, the phylogenetic modeling framework we propose here improves upon the state-of-the-art by addressing known limitations of inter-species comparisons of epigenomic profiles, and has broad implications in the field of comparative functional genomics.
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