Sourav Ghosh

Genome-wide analysis reveals distinct patterns of epigenetic features in long non-coding RNA loci

A major fraction of the transcriptome of higher organisms comprised an extensive repertoire of long non-coding RNA (lncRNA) which express in a cell type and development stage-specific manner. While lncRNAs are a proven component of epigenetic gene expression modulation, epigenetic regulation of lncRNA itself remains poorly understood. Here we have analysed pan-genomic DNA methylation and histone modification marks (H3K4me3, H3K9me3, H3K27me3 and H3K36me3) associated with transcription start site (TSS) of lncRNA in four different cell types and three different tissue types representing various cellular stages. We observe that histone marks associated with active transcription H3K4me3 and H3K36me3 along with the repressive histone mark H3K27me3 have similar distribution pattern around TSS irrespective of cell types. Also, the density of these marks correlates well with expression of protein-coding and lncRNA genes. In contrast, the lncRNA genes harbour higher methylation density around TSS than protein-coding genes regardless of their expression status. Furthermore, we found that DNA methylation along with the other repressive histone mark H3K9me3 does not seem to play a role in lncRNA expression. Thus, our observation suggests that epigenetic regulation of lncRNA shares common features with mRNA except the role of DNA methylation which is markedly dissimilar.

lncRNome: a comprehensive knowledgebase of human long noncoding RNAs

The advent of high-throughput genome scale technologies has enabled us to unravel a large amount of the previously unknown transcriptionally active regions of the genome. Recent genome-wide studies have provided annotations of a large repertoire of various classes of noncoding transcripts. Long noncoding RNAs (lncRNAs) form a major proportion of these novel annotated noncoding transcripts, and presently known to be involved in a number of functionally distinct biological processes. Over 18 000 transcripts are presently annotated as lncRNA, and encompass previously annotated classes of noncoding transcripts including large intergenic noncoding RNA, antisense RNA and processed pseudogenes. There is a significant gap in the resources providing a stable annotation, cross-referencing and biologically relevant information. lncRNome has been envisioned with the aim of filling this gap by integrating annotations on a wide variety of biologically significant information into a comprehensive knowledgebase. To the best of our knowledge, lncRNome is one of the largest and most comprehensive resources for lncRNAs. Database URL: http://genome.igib.res.in/lncRNome

High resolution methylome map of rat indicates role of intragenic DNA methylation in identification of coding region.

DNA methylation is crucial for gene regulation and maintenance of genomic stability. Rat has been a key model system in understanding mammalian systemic physiology, however detailed rat methylome remains uncharacterized till date. Here, we present the first high resolution methylome of rat liver generated using Methylated DNA immunoprecipitation and high throughput sequencing (MeDIP-Seq) approach. We observed that within the DNA/RNA repeat elements, simple repeats harbor the highest degree of methylation. Promoter hypomethylation and exon hypermethylation were common features in both RefSeq genes and expressed genes (as evaluated by proteomic approach). We also found that although CpG islands were generally hypomethylated, about 6% of them were methylated and a large proportion (37%) of methylated islands fell within the exons. Notably, we obeserved significant differences in methylation of terminal exons (UTRs); methylation being more pronounced in coding/partially coding exons compared to the non-coding exons. Further, events like alternate exon splicing (cassette exon) and intron retentions were marked by DNA methylation and these regions are retained in the final transcript. Thus, we suggest that DNA methylation could play a crucial role in marking coding regions thereby regulating alternative splicing. Apart from generating the first high resolution methylome map of rat liver tissue, the present study provides several critical insights into methylome organization and extends our understanding of interplay between epigenome, gene expression and genome stability.

Comparative analysis of human mitochondrial methylomes shows distinct patterns of epigenetic regulation in mitochondria

DNA methylation and histone modifications across the nuclear genome have been extensively analyzed, but the epigenetic modifications associated with the mitochondrial genome have not yet been analyzed at high resolutions. In the present work, we analyzed methyl-cytosine profiles from methylated DNA immunoprecipitation datasets from 39 different human cell and tissue types from the NIH Roadmap Epigenomics project and validated the data using an orthologous bisulfite sequencing dataset. We observe a distinct distribution of zmethyl-cytosine in mitochondrial genomes which are conserved across all cell and tissue types. This study thus describes the first comprehensive map of methyl cytosines across the human mitochondrial genome.

Mitoepigenetics: The different shades of grey.

Epigenetic modifications of the nuclear genome have been well studied and it is established that these modifications play a key role in nuclear gene expression. However, the status of mitochondrial epigenetic modifications has not been delved in detail. The recent technological advancements in the genome analyzing tools and techniques, have helped in investigating mitochondrial epigenetic modifications with greater resolution and studies have indicated a regulatory role of the mitochondrial epigenome. Association of mitochondrial DNA methylation with various disease conditions, drug treatment, aging, exposure to environmental pollutants etc. has lent credence to this belief. Herein, we have reviewed studies on mitochondrial epigenetic modifications with a focus to comprehend its regulatory role in gene expression and disease association.

Distinct Patterns of Genetic Variations in Potential Functional Elements in Long Noncoding RNAs

Non‐protein‐coding RNAs have increasingly been shown to be an important class of regulatory RNAs having significant roles in regulation of gene expression. The long noncoding RNA (lncRNA) gene family presently constitutes a large number of noncoding RNA (ncRNA) loci almost equaling the number of protein‐coding genes. Nevertheless, the biological roles and mechanisms of the majority of lncRNAs are poorly understood, with exceptions of a very few well‐studied candidates. The availability of genome‐scale variation datasets, and increasing number of variant loci from genome‐wide association studies falling in lncRNA loci have motivated us to understand the patterns of genomic variations in lncRNA loci, their potential functional correlates, and selection in populations. In the present study, we have performed a comprehensive analysis of genomic variations in lncRNA loci. We analyzed for patterns and distributions of genomic variations with respect to potential functional domains in lncRNAs. The analysis reveals a distinct distribution of variations in subclasses of long ncRNAs and in potential functional domains of lncRNAs. We further examined signals of selections and allele frequencies of these prioritized set of lncRNAs. To the best of our knowledge, this is the first and comprehensive large‐scale analysis of genetic variations in long ncRNAs.

Hydroxymethyl cytosine marks in the human mitochondrial genome are dynamic in nature

Apart from DNA methylation, hydroxymethylation has increasingly been studied as an important epigenetic mark. 5- hydroxymethylcytosines, though initially were thought to be an intermediary product of demethylation, recent studies suggest this to be a highly regulated process and modulated by the TET family of enzymes. Recent genome wide studies have shown that hydroxymethylcytosine marks are closely associated with the regulation of important biological processes like transcription and embryonic development. It is also known that aberrant hydroxymethylation marks have been associated with diseases like cancer. The presence of hydroxymethylcytosines in the mitochondrial genome has been earlier suggested, though the genome-scale map has not been laid out. In this present study, we have mapped and analyzed the hydroxymethylcytosine marks in the mitochondrial genome using 23 different publicly available datasets. We cross validated our data by checking for consistency across a subset of genomic regions previously annotated to hydroxymethylcytosines and show good consistency. We observe a dynamic distribution of hydroxymethylation marks in the mitochondrial genome. Unlike the methylcytosine marks, hydroxymethylcytosine marks are characterized by the lack of conservation across the samples considered, though similar cell types shared the pattern. We additionally observed that the hydroxymethylation marks are enriched in the upstream of GSS (gene start site) regions and in gene body as similar as nuclear genes. To the best of our knowledge, this is the first genome-scale map of hydroxymethyl cytosines in the human mitochondrial genome.

Genome-wide regulatory dynamics of G-quadruplexes in human malaria parasite Plasmodium falciparum

The AT-rich genome of P. falciparum has uniquely localized G-rich stretches that have propensity to form G-quadruplexes. However, their global occurrence and potential biological roles in the parasite are poorly explored. Our genome-wide analysis revealed unique enrichment of quadruplexes in P. falciparum genome which was remarkably different from other Plasmodium species. A distinct predominance of quadruplexes was observed in nuclear and organellar genes that participate in antigenic variation, pathogenesis, DNA/RNA regulation, metabolic and protein quality control processes. Data also suggested association of quadruplexes with SNPs and DNA methylation. Furthermore, analysis of steady state mRNA (RNA-seq) and polysome-associated mRNA (Ribosome profiling) data revealed stage-specific differences in translational efficiency of quadruplex harboring genes. Taken together, our findings hint towards existence of regulatory dynamics associated with quadruplexes that may modulate translational efficiency of quadruplex harboring genes to provide survival advantage to the parasite against host immune response and antimalarial drug pressure.

Distinct patterns of epigenetic marks and transcription factor binding sites across promoters of sense-intronic long noncoding RNAs.

Long noncoding RNAs (lncRNAs) are a new class of noncoding RNAs that have been extensively studied in the recent past as a regulator of gene expression, including modulation of epigenetic regulation. The lncRNAs class encompasses a number of subclasses, classified based on their genomic loci and relation to protein-coding genes. Functional differences between subclasses have been increasingly studied in the recent years, though the regulation of expression and biogenesis of lncRNAs have been poorly studied. The availability of genome-scale datasets of epigenetic marks has motivated us to understand the patterns and processes of epigenetic regulation of lncRNAs. Here we analysed the occurrence of expressive and repressive histone marks at the transcription start site (TSS) of lncRNAs and their subclasses, and compared these profiles with that of the protein-coding regions. We observe distinct differences in the density of histone marks across the TSS of a few lncRNA subclasses. The sense-intronic lncRNA subclass showed a paucity for mapped histone marks across the TSS which were significantly different than all the lncRNAs and protein-coding genes in most cases. Similar pattern was also observed for the density of transcription factor binding sites (TFBS). These observations were generally consistent across cell and tissue types. The differences in density across the promoter were significantly associated with the expression level of the genes, but the differences between the densities across long noncoding and protein-coding gene promoters were consistent irrespective of the expression levels. Apart from suggesting general differences in epigenetic regulatory marks across long noncoding RNA promoters, our analysis suggests a possible alternative mechanism of regulation and/or biogenesis of sense-intronic lncRNAs.

Comparative analysis of human mitochondrial methylome show distinct patterns of epigenetic regulation in mitochondria

Method We analyzed methyl-cytosine profiles as evident from Methylated DNA immunoprecipitation across 39 tissues and cell lines that were available as part of the NIH RoadMap Epigenomics project. The mitochondrial reference genome used in the current study was derived from the UCSC human genome build hg19. We used custom scripts to retrieve reads mapping to the mitochondrial genome. Results Our analysis suggests that the general profile of methylated cytosines across the samples show a distinct pattern. This pattern was generally conserved across the datasets considered, with exceptions of a few regions which showed variability in methylation amongst datasets analyzed. We show that certain regions of the mitochondria could be differentially methylated in datasets which show distinct temporal and functional characteristics, like Brain and Blood. One such region harbors the loci associated with mitochondrial encoded NADH dehydrogenase (MTND6), variations in which are associated with neurological disorders. To date this is the first and comprehensive analyses of genome-scale methylation data for Human mitochondria.