Histone deacetylase inhibition accelerates the early events of stem cell differentiation: transcriptomic and epigenetic analysis

BACKGROUND: Epigenetic mechanisms regulate gene expression patterns affecting cell function and differentiation. In this report, we examine the role of histone acetylation in gene expression regulation in mouse embryonic stem cells employing transcriptomic and epigenetic analysis. RESULTS: Embryonic stem cells treated with the histone deacetylase inhibitor Trichostatin A (TSA), undergo morphological and gene expression changes indicative of differentiation. Gene profiling utilizing Affymetrix microarrays revealed the suppression of important pluripotency factors, including Nanog, a master regulator of stem cell identity, and the activation of differentiation-related genes. Transcriptional and epigenetic changes induced after 6-12 hours of TSA treatment mimic those that appear during embryoid body differentiation. We show here that the early steps of stem cell differentiation are marked by the enhancement of bulk activatory histone modifications. At the individual gene level, we found that transcriptional reprogramming triggered by histone deacetylase inhibition correlates with rapid changes in activating K4 trimethylation and repressive K27 trimethylation of histone H3. The establishment of H3K27 trimethylation is required for stable gene suppression whereas in its absence, genes can be reactivated upon TSA removal. CONCLUSION: Our data suggest that inhibition of histone deacetylases accelerates the early events of differentiation by regulating the expression of pluripotency- and differentiation-associated genes in an opposite manner. This analysis provides information about genes that are important for embryonic stem cell function and the epigenetic mechanisms that regulate their expression

Evolution and implications of de novo genes in humans

Genes and translated open reading frames (ORFs) that emerged de novo from previously non-coding sequences provide species with opportunities for adaptation. When aberrantly activated, some human-specific de novo genes and ORFs have disease-promoting properties—for instance, driving tumour growth. Thousands of putative de novo coding sequences have been described in humans, but we still do not know what fraction of those ORFs has readily acquired a function. Here, we discuss the challenges and controversies surrounding the detection, mechanisms of origin, annotation, validation and characterization of de novo genes and ORFs. Through manual curation of literature and databases, we provide a thorough table with most de novo genes reported for humans to date. We re-evaluate each locus by tracing the enabling mutations and list proposed disease associations, protein characteristics and supporting evidence for translation and protein detection. This work will support future explorations of de novo genes and ORFs in humans

Heritability and Tissue Specificity of Expression Quantitative Trait Loci

Variation in gene expression is heritable and has been mapped to the genome in humans and model organisms as expression quantitative trait loci (eQTLs). We applied integrated genome-wide expression profiling and linkage analysis to the regulation of gene expression in fat, kidney, adrenal, and heart tissues using the BXH/HXB panel of rat recombinant inbred strains. Here, we report the influence of heritability and allelic effect of the quantitative trait locus on detection of cis- and trans-acting eQTLs and discuss how these factors operate in a tissue-specific context. We identified several hundred major eQTLs in each tissue and found that cis-acting eQTLs are highly heritable and easier to detect than trans-eQTLs. The proportion of heritable expression traits was similar in all tissues; however, heritability alone was not a reliable predictor of whether an eQTL will be detected. We empirically show how the use of heritability as a filter reduces the ability to discover trans-eQTLs, particularly for eQTLs with small effects. Only 3% of cis- and trans-eQTLs exhibited large allelic effects, explaining more than 40% of the phenotypic variance, suggestive of a highly polygenic control of gene expression. Power calculations indicated that, across tissues, minor differences in genetic effects are expected to have a significant impact on detection of trans-eQTLs. Trans-eQTLs generally show smaller effects than cis-eQTLs and have a higher false discovery rate, particularly in more heterogeneous tissues, suggesting that small biological variability, likely relating to tissue composition, may influence detection of trans-eQTLs in this system. We delineate the effects of genetic architecture on variation in gene expression and show the sensitivity of this experimental design to tissue sampling variability in large-scale eQTL studies

α1A-Adrenergic Receptor-Directed Autoimmunity Induces Left Ventricular Damage and Diastolic Dysfunction in Rats

BACKGROUND: Agonistic autoantibodies to the alpha(1)-adrenergic receptor occur in nearly half of patients with refractory hypertension; however, their relevance is uncertain. METHODS/PRINCIPAL FINDINGS: We immunized Lewis rats with the second extracellular-loop peptides of the human alpha(1A)-adrenergic receptor and maintained them for one year. Alpha(1A)-adrenergic antibodies (alpha(1A)-AR-AB) were monitored with a neonatal cardiomyocyte contraction assay by ELISA, and by ERK1/2 phosphorylation in human alpha(1A)-adrenergic receptor transfected Chinese hamster ovary cells. The rats were followed with radiotelemetric blood pressure measurements and echocardiography. At 12 months, the left ventricles of immunized rats had greater wall thickness than control rats. The fractional shortening and dp/dt(max) demonstrated preserved systolic function. A decreased E/A ratio in immunized rats indicated a diastolic dysfunction. Invasive hemodynamics revealed increased left ventricular end-diastolic pressures and decreased dp/dt(min). Mean diameter of cardiomyocytes showed hypertrophy in immunized rats. Long-term blood pressure values and heart rates were not different. Genes encoding sarcomeric proteins, collagens, extracellular matrix proteins, calcium regulating proteins, and proteins of energy metabolism in immunized rat hearts were upregulated, compared to controls. Furthermore, fibrosis was present in immunized hearts, but not in control hearts. A subset of immunized and control rats was infused with angiotensin (Ang) II. The stressor raised blood pressure to a greater degree and led to more cardiac fibrosis in immunized, than in control rats. CONCLUSIONS/SIGNIFICANCE: We show that alpha(1A)-AR-AB cause diastolic dysfunction independent of hypertension, and can increase the sensitivity to Ang II. We suggest that alpha(1A)-AR-AB could contribute to cardiovascular endorgan damage

histoneHMM:Differential analysis of histone modifications with broad genomic footprints

BACKGROUND: ChIP-seq has become a routine method for interrogating the genome-wide distribution of various histone modifications. An important experimental goal is to compare the ChIP-seq profiles between an experimental sample and a reference sample, and to identify regions that show differential enrichment. However, comparative analysis of samples remains challenging for histone modifications with broad domains, such as heterochromatin-associated H3K27me3, as most ChIP-seq algorithms are designed to detect well defined peak-like features. RESULTS: To address this limitation we introduce histoneHMM, a powerful bivariate Hidden Markov Model for the differential analysis of histone modifications with broad genomic footprints. histoneHMM aggregates short-reads over larger regions and takes the resulting bivariate read counts as inputs for an unsupervised classification procedure, requiring no further tuning parameters. histoneHMM outputs probabilistic classifications of genomic regions as being either modified in both samples, unmodified in both samples or differentially modified between samples. We extensively tested histoneHMM in the context of two broad repressive marks, H3K27me3 and H3K9me3, and evaluated region calls with follow up qPCR as well as RNA-seq data. Our results show that histoneHMM outperforms competing methods in detecting functionally relevant differentially modified regions. CONCLUSION: histoneHMM is a fast algorithm written in C++ and compiled as an R package. It runs in the popular R computing environment and thus seamlessly integrates with the extensive bioinformatic tool sets available through Bioconductor. This makeshistoneHMM an attractive choice for the differential analysis of ChIP-seq data. Software is available from http://histonehmm.molgen.mpg.de

Overexpression of Full-Length Centrobin Rescues Limb Malformation but Not Male Fertility of the Hypodactylous (hd) Rats

Rat hypodactyly (hd) mutation is characterized by abnormal spermatogenesis and sperm decapitation, limb malformation (missing digits II and III) and growth retardation. We have previously reported centrobin (centrosome BRCA2-interacting protein) truncation at the C-terminus in the hd mutant. Here, we report data from a transgenic rescue experiment carried out to determine a role of centrobin in pathogenesis of hd. The transgenic construct, consisting of full-length-coding cDNA linked to a ubiquitous strong promoter/enhancer combination, was inserted to chromosome 16 into a LINE repeat. No known gene is present in the vicinity of the insertion site. Transgenic centrobin was expressed in all tissues tested, including testis. Transgenic animals show normal body weight and limb morphology as well as average weight of testis and epididymis. Yet, abnormal spermatogenesis and sperm decapitation persisted in the transgenic animals. Western blotting showed the coexistence of full-length and truncated or partially degraded centrobin in sperm of the rescued transgenic animals. Immunocytochemistry showed a buildup of centrobin and ODF2 (outer dense fiber 2) at the sperm decapitation site in the hd mutant and rescued transgenic rats. Additional findings included bulge-like formations and thread-like focal dissociations along the sperm flagellum and the organization of multiple whorls of truncated sperm flagella in the epididymal lumen. We conclude that centrobin is essential for normal patterning of the limb autopod. Centrobin may be required for stabilizing the attachment of the sperm head to flagellum and for maintaining the structural integrity of the sperm flagellum. We postulate that the presence of truncated centrobin, coexisting with full-length centrobin, together with incorrect timing of transgenic centrobin expression may hamper the rescue of fertility in hd male rats

Fine Mapping of the 1p36 Deletion Syndrome Identifies Mutation of PRDM16 as a Cause of Cardiomyopathy

Deletion 1p36 syndrome is recognized as the most common terminal deletion syndrome. Here, we describe the loss of a gene within the deletion that is responsible for the cardiomyopathy associated with monosomy 1p36, and we confirm its role in nonsyndromic left ventricular noncompaction cardiomyopathy (LVNC) and dilated cardiomyopathy (DCM). With our own data and publically available data from array comparative genomic hybridization (aCGH), we identified a minimal deletion for the cardiomyopathy associated with 1p36del syndrome that included only the terminal 14 exons of the transcription factor PRDM16 (PR domain containing 16), a gene that had previously been shown to direct brown fat determination and differentiation. Resequencing of PRDM16 in a cohort of 75 nonsyndromic individuals with LVNC detected three mutations, including one truncation mutant, one frameshift null mutation, and a single missense mutant. In addition, in a series of cardiac biopsies from 131 individuals with DCM, we found 5 individuals with 4 previously unreported nonsynonymous variants in the coding region of PRDM16. None of the PRDM16 mutations identified were observed in more than 6,400 controls. PRDM16 has not previously been associated with cardiac disease but is localized in the nuclei of cardiomyocytes throughout murine and human development and in the adult heart. Modeling of PRDM16 haploinsufficiency and a human truncation mutant in zebrafish resulted in both contractile dysfunction and partial uncoupling of cardiomyocytes and also revealed evidence of impaired cardiomyocyte proliferative capacity. In conclusion, mutation of PRDM16 causes the cardiomyopathy in 1p36 deletion syndrome as well as a proportion of nonsyndromic LVNC and DCM

Wars2 is a determinant of angiogenesis.

Coronary flow (CF) measured ex vivo is largely determined by capillary density that reflects angiogenic vessel formation in the heart in vivo. Here we exploit this relationship and show that CF in the rat is influenced by a locus on rat chromosome 2 that is also associated with cardiac capillary density. Mitochondrial tryptophanyl-tRNA synthetase (Wars2), encoding an L53F protein variant within the ATP-binding motif, is prioritized as the candidate at the locus by integrating genomic data sets. WARS2(L53F) has low enzyme activity and inhibition of WARS2 in endothelial cells reduces angiogenesis. In the zebrafish, inhibition of wars2 results in trunk vessel deficiencies, disordered endocardial-myocardial contact and impaired heart function. Inhibition of Wars2 in the rat causes cardiac angiogenesis defects and diminished cardiac capillary density. Our data demonstrate a pro-angiogenic function for Wars2 both within and outside the heart that may have translational relevance given the association of WARS2 with common human diseases

Author Correction: WWP2 regulates pathological cardiac fibrosis by modulating SMAD2 signaling.

An amendment to this paper has been published and can be accessed via a link at the top of the paper

Научно-исследовательская, культурно-массовая, физкультурно-спортивная и общественная деятельность студентов как факторы успешности их учебно-познавательного процесса

Материалы IV Респ. науч.-метод. конф., посвящ. 120-летию со дня рождения П. О. Сухого, Гомель, 29–30 окт. 2015 г