LSD1 cooperates with CTIP2 to promote HIV-1 transcriptional silencing

Microglial cells are the main HIV-1 targets in the central nervous system (CNS) and constitute an important reservoir of latently infected cells. Establishment and persistence of these reservoirs rely on the chromatin structure of the integrated proviruses. We have previously demonstrated that the cellular cofactor CTIP2 forces heterochromatin formation and HIV-1 gene silencing by recruiting HDAC and HMT activities at the integrated viral promoter. In the present work, we report that the histone demethylase LSD1 represses HIV-1 transcription and viral expression in a synergistic manner with CTIP2. We show that recruitment of LSD1 at the HIV-1 proximal promoter is associated with both H3K4me3 and H3K9me3 epigenetic marks. Finally, our data suggest that LSD1-induced H3K4 trimethylation is linked to hSET1 recruitment at the integrated provirus

Genome-Wide Association Study Identifies GPC5 as a Novel Genetic Locus Protective against Sudden Cardiac Arrest

BACKGROUND:Existing studies indicate a significant genetic component for sudden cardiac arrest (SCA) and genome-wide association studies (GWAS) provide an unbiased approach for identification of novel genes. We performed a GWAS to identify genetic determinants of SCA. METHODOLOGY/PRINCIPAL FINDINGS:We used a case-control design within the ongoing Oregon Sudden Unexpected Death Study (Oregon-SUDS). Cases (n = 424) were SCAs with coronary artery disease (CAD) among residents of Portland, OR (2002-07, population approximately 1,000,000) and controls (n = 226) were residents with CAD, but no history of SCA. All subjects were of White-European ancestry and GWAS was performed using Affymetrix 500K/5.0 and 6.0 arrays. High signal markers were genotyped in SCA cases (n = 521) identified from the Atherosclerosis Risk in Communities Study (ARIC) and the Cardiovascular Health Study (CHS) (combined n = 19,611). No SNPs reached genome-wide significance (p<5x10(-8)). SNPs at 6 loci were prioritized for follow-up primarily based on significance of p<10(-4) and proximity to a known gene (CSMD2, GPR37L1, LIN9, B4GALNT3, GPC5, and ZNF592). The minor allele of GPC5 (GLYPICAN 5, rs3864180) was associated with a lower risk of SCA in Oregon-SUDS, an effect that was also observed in ARIC/CHS whites (p<0.05) and blacks (p<0.04). In a combined Cox proportional hazards model analysis that adjusted for race, the minor allele exhibited a hazard ratio of 0.85 (95% CI 0.74 to 0.98; p<0.01). CONCLUSIONS/SIGNIFICANCE:A novel genetic locus for SCA, GPC5, was identified from Oregon-SUDS and successfully validated in the ARIC and CHS cohorts. Three other members of the Glypican family have been previously implicated in human disease, including cardiac conditions. The mechanism of this specific association requires further study

Genome-Wide Binding Map of the HIV-1 Tat Protein to the Human Genome

The HIV-1 Trans-Activator of Transcription (Tat) protein binds to multiple host cellular factors and greatly enhances the level of transcription of the HIV genome. While Tat's control of viral transcription is well-studied, much less is known about the interaction of Tat with the human genome. Here, we report the genome-wide binding map of Tat to the human genome in Jurkat T cells using chromatin immunoprecipitation combined with next-generation sequencing. Surprisingly, we found that ∼53% of the Tat target regions are within DNA repeat elements, greater than half of which are Alu sequences. The remaining target regions are located in introns and distal intergenic regions; only ∼7% of Tat-bound regions are near transcription start sites (TSS) at gene promoters. Interestingly, Tat binds to promoters of genes that, in Jurkat cells, are bound by the ETS1 transcription factor, the CBP histone acetyltransferase and/or are enriched for histone H3 lysine 4 tri-methylation (H3K4me3) and H3K27me3. Tat binding is associated with genes enriched with functions in T cell biology and immune response. Our data reveal that Tat's interaction with the host genome is more extensive than previously thought, with potentially important implications for the viral life cycle

Epigenetic Regulation of HIV-1 Latency by Cytosine Methylation

Human immunodeficiency virus type 1 (HIV-1) persists in a latent state within resting CD4+ T cells of infected persons treated with highly active antiretroviral therapy (HAART). This reservoir must be eliminated for the clearance of infection. Using a cDNA library screen, we have identified methyl-CpG binding domain protein 2 (MBD2) as a regulator of HIV-1 latency. Two CpG islands flank the HIV-1 transcription start site and are methylated in latently infected Jurkat cells and primary CD4+ T cells. MBD2 and histone deacetylase 2 (HDAC2) are found at one of these CpG islands during latency. Inhibition of cytosine methylation with 5-aza-2′deoxycytidine (aza-CdR) abrogates recruitment of MBD2 and HDAC2. Furthermore, aza-CdR potently synergizes with the NF-κB activators prostratin or TNF-α to reactivate latent HIV-1. These observations confirm that cytosine methylation and MBD2 are epigenetic regulators of HIV-1 latency. Clearance of HIV-1 from infected persons may be enhanced by inclusion of DNA methylation inhibitors, such as aza-CdR, and NF-κB activators into current antiviral therapies

Histone deacetylases in viral infections

Chromatin remodeling and gene expression are regulated by histone deacetylases (HDACs) that condense the chromatin structure by deacetylating histones. HDACs comprise a group of enzymes that are responsible for the regulation of both cellular and viral genes at the transcriptional level. In mammals, a total of 18 HDACs have been identified and grouped into four classes, i.e., class I (HDACs 1, 2, 3, 8), class II (HDACs 4, 5, 6, 7, 9, 10), class III (Sirt1–Sirt7), and class IV (HDAC11). We review here the role of HDACs on viral replication and how HDAC inhibitors could potentially be used as new therapeutic tools in several viral infections

Cell line-dependent variability in HIV activation employing DNMT inhibitors

Long-lived reservoirs of Human Immunodeficiency Virus (HIV) latently infected cells present the main barrier to a cure for HIV infection. Much interest has focused on identifying strategies to activate HIV, which would be used together with antiretrovirals to attack reservoirs. Several HIV activating agents, including Tumor Necrosis Factor alpha (TNFα) and other agents that activate via NF-kB are not fully effective in all latent infection models due to epigenetic restrictions, such as DNA methylation and the state of histone acetylation. DNA methyltransferases (DNMT) inhibitors like 5-aza-2'deoxycytidine (Aza-CdR) and histone deacetylase (HDAC) inhibitors like Trichostatin A (TSA) have been proposed as agents to enhance reactivation and have shown activity in model systems. However, it is not clear how the activities of DNMT and HDAC inhibitors range across different latently infected cell lines, potential models for the many different latently infected cells within an HIV patient. We determined HIV activation following treatment with TNFα, TSA and Aza-CdR across a range of well known latently infected cell lines. We assessed the activity of these compounds in four different Jurkat T cell-derived J-Lat cell lines (6.3, 8.4, 9.2 and 10.6), which have a latent HIV provirus in which GFP replaces Nef coding sequence, and ACH-2 and J1.1 (T cell-derived), and U1 (promonocyte-derived) cell lines with full-length provirus. We found that Aza-CdR plus TNFα activated HIV at least twice as well as TNFα alone for almost all J-Lat cells, as previously described, but not for J-Lat 10.6, in which TNFα plus Aza-CdR moderately decreased activation compared to TNFα alone. Surprisingly, a much greater reduction of TNFα-stimulated activation with Aza-CdR was detected for ACH-2, J1.1 and U1 cells. Reaching the highest reduction in U1 cells with a 75% reduction. Interestingly, Aza-CdR not only decreased TNFα induction of HIV expression in certain cell lines, but also decreased activation by TSA. Since DNMT inhibitors reduce the activity of provirus activators in some HIV latently infected cell lines the use of epigenetic modifying agents may need to be carefully optimized if they are to find clinical utility in therapies aimed at attacking latent HIV reservoirs

Multiscale computational analysis of the bioelectric consequences of myocardial ischaemia and infarction

[EN] Ischaemic heart disease is considered as the single most frequent cause of death, provoking more than 7 000 000 deaths every year worldwide. A high percentage of patients experience sudden cardiac death, caused in most cases by tachyarrhythmic mechanisms associated to myocardial ischaemia and infarction. These diseases are difficult to study using solely experimental means due to their complex dynamics and unstable nature. In the past decades, integrative computational simulation techniques have become a powerful tool to complement experimental and clinical research when trying to elucidate the intimate mechanisms of ischaemic electrophysiological processes and to aid the clinician in the improvement and optimization of therapeutic procedures. The purpose of this paper is to briefly review some of the multiscale computational models of myocardial ischaemia and infarction developed in the past 20 years, ranging from the cellular level to whole-heart simulations.This work was partially supported by the 'VI Plan Nacional de Investigacion Cientifica, Desarrollo e Innovacion Tecnologica' from the Ministerio de Economia y Competitividad of Spain (grant number TIN2012-37546-C03-01) and the European Commission (European Regional Development Funds-ERDF-FEDER), and by the Direccion General de Politica Cientifica de la Generalitat Valenciana (grant number GV/2013/119).Ferrero De Loma-Osorio, JM.; Trénor Gomis, BA.; Romero Pérez, L. (2014). Multiscale computational analysis of the bioelectric consequences of myocardial ischaemia and infarction. EP-Europace. 16(3):405-415. https://doi.org/10.1093/europace/eut405S40541516

Arrhythmogenic actions of the Ca2+ channel agonist FPL-64716 in Langendorff-perfused murine hearts

The experiments explored the extent to which alterations in L-type Ca2+ channel-mediated Ca2+ entry triggers Ca2+-mediated arrhythmogenesis in Langendorff-perfused murine hearts through use of the specific L-type Ca2+ channel modulator FPL-64716 (FPL). Introduction of FPL (1 μm) resulted in a gradual development (>10 min) of diastolic electrical events and alternans in spontaneously beating hearts from which monophasic action potentials were recorded. In regularly paced hearts, they additionally led to non-sustained and sustained ventricular tachycardia (nsVT and sVT). Programmed electrical stimulation (PES) resulted in nsVT and sVT after 5–10 and >10 min perfusion, respectively. Pretreatments with nifedipine, diltiazem and cyclopiazonic acid abolished arrhythmogenic tendency induced by subsequent introduction of FPL, consistent with its dependence upon both extracellular Ca2+ entry and the degree of filling of the sarcoplasmic reticular Ca2+ store. Values for action potential duration at 90% repolarization when any of these agents were applied to FPL-treated hearts became indistinguishable from those shown by untreated control hearts, in contrast to earlier reports of their altering in long QT syndrome type 3 and hypokalaemic murine models for re-entrant arrhythmogenesis. These arrhythmic effects instead correlated with alterations in Ca2+ homeostasis at the single-cell level found in investigations of the effects of both FPL and the same agents in regularly stimulated fluo−3 loaded myocytes. These findings are compatible with a prolonged extracellular Ca2+ entry that potentially results in an intracellular Ca2+ overload and produces the cardiac arrhythmogenecity following addition of FPL