The Poorest of Poor Suffer the Greatest Burden From Smokeless Tobacco Use: A Study From 140 Countries.

Background: The full extent of global smokeless tobacco (SLT) use and its association with key demographic factors such as gender, place of residence, and household or country income status is not yet known. Methods: The global burden of SLT use among adults was estimated using nationally representative data of 140 countries by gender and country income group. Countries were grouped in Group 1 (low and low-middle income countries combined) and Group 2 (upper middle and high income countries combined). The number of male and female SLT users was calculated using prevalence and population estimates of corresponding age groups. Results: Nearly one in 10 males and one in 20 females used SLT in some form. SLT use prevalence was significantly higher among males (p < .001) and females (p < .001) in Group 1 countries compared with their counterparts in Group 2 countries. However, for both Group 1 (p < .01) and Group 2 (p < .01), males were more likely to use SLT than females. Nearly 91% of a total 356 million adult SLT users resided in Group 1 countries, with 81.6% in countries of WHO South-East Asia region (SEAR). In SEAR and African region, SLT use was higher in rural areas and poorest communities. Conclusion: The majority of the burden of SLT use is on lower and lower middle income countries with the greatest burden on the poorest segments of the population in these countries. Implications: This study brings the comprehensive information on epidemiology of SLT use among adults at global level. Ninety percent of SLT burden is in low and low-middle income group of countries and more specifically among the poorest group in such countries. These countries need to have strategies to implement different provisions of the WHO Framework Convention on Tobacco Control. The program in such countries should be targeted towards the poorest communities for effective SLT control.

Regulation of toxic contents of smokeless tobacco products.

Effective regulation of contents of tobacco products is one of the primary milestones to reduce negative health effects associated with the use of smokeless tobacco (SLT) products. As per the available sources, testing of some SLT products has been done on ad hoc basis, but there is a lack of comprehensive and periodic analysis of these products. In addition, the available results indicate huge variations among the levels of pH, 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone, N-nitrosonornicotine, benzo[a]pyrene, heavy metals and nicotine within different products as well as within different brands of the same product. This review was aimed to throw light on the variations and gaps in testing of SLT products and emphasize the need for strong policy regulation for monitoring the chemical constituents of these products.

In-silico study of toxicokinetics and disease association of chemicals present in smokeless tobacco products.

Smokeless tobacco (SLT) products are consumed by millions of people in over 130 countries around the world. Consumption of SLT has been estimated to cause a number of diseases accounting to more than 0.65 million deaths per year. There is sufficient epidemiological evidence on the association of SLT products with nicotine addiction, cancers of oral cavity and digestive systems but there is a lack of understanding of the role of toxic chemicals in these diseases. We provide the first comprehensive in-silico analysis of chemical compounds present in different SLT products used worldwide. Many of these compounds are found to have good absorption, solubility and permeability along with mutagenic and toxic properties. They are also found to target more than 350 human proteins involved in a plethora of human biological processes and pathways. Along with all the previously known diseases, the present study has identified the association of compounds of SLT products with a number of unknown diseases like neurodegenerative, immune and cardiac diseases (Left ventricular non compaction, dilated cardiomyopathy etc). These findings indicate far-reaching impact of SLT products on human health than already known which needs further validations using epidemiological, in-vitro and in-vivo methodologies. Thus, this study will provide one stop information for the policy makers in development of regulatory policies on toxic contents of SLT products.

Genomic variations in non-coding RNAs: Structure, function and regulation.

The last decade has seen tremendous improvements in the understanding of human variations and their association with human traits and diseases. The availability of high-resolution map of the human transcriptome and the discovery of a large number of non-protein coding RNA genes has created a paradigm shift in the understanding of functional variations in non-coding RNAs. Several groups in recent years have reported functional variations and trait or disease associated variations mapping to non-coding RNAs including microRNAs, small nucleolar RNAs and long non-coding RNAs. The understanding of the functional consequences of variations in non-coding RNAs has been largely restricted by the limitations in understanding the functionalities of the non-coding RNAs. In this short review, we outline the current state-of-the-art of the field with emphasis on providing a conceptual outline as on how variations could modulate changes in the sequence, structure, and thereby the functionality of non-coding RNAs.

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.

Computational approaches towards understanding human long non-coding RNA biology.

Long non-coding RNAs (lncRNAs) form the largest class of non-protein coding genes in the human genome. While a small subset of well-characterized lncRNAs has demonstrated their significant role in diverse biological functions like chromatin modifications, post-transcriptional regulation, imprinting etc., the functional significance of a vast majority of them still remains an enigma. Increasing evidence of the implications of lncRNAs in various diseases including cancer and major developmental processes has further enhanced the need to gain mechanistic insights into the lncRNA functions. Here, we present a comprehensive review of the various computational approaches and tools available for the identification and annotation of long non-coding RNAs. We also discuss a conceptual roadmap to systematically explore the functional properties of the lncRNAs using computational approaches.

Co-evolutionary analysis implies auxiliary functions of HSP110 in Plasmodium falciparum.

Plasmodium falciparum encounters frequent environmental challenges during its life cycle which makes productive protein folding immensely challenging for its metastable proteome. To identify the important components of protein folding machinery involved in maintaining P. falciparum proteome, we performed a proteome-wide phylogenetic profiling across various species. We found that except HSP110, the parasite lost all other cytosolic nucleotide exchange factors essential for regulating HSP70 which is the centrum of the protein folding network. Evolutionary and structural analysis shows that besides its canonical interaction with HSP70, PfHSP110 has acquired sequence insertions for additional dynamic interactions. Molecular co-evolution profile depicts that the co-evolving proteins of PfHSP110 belong to distinct pathways like genetic variation, DNA repair, fatty acid biosynthesis, protein modification/trafficking, molecular motions, and apoptosis. These proteins exhibit unique physiochemical properties like large size, high iso-electric point, low solubility, and antigenicity, hence require PfHSP110 chaperoning to attain functional state. Co-evolving protein interaction network suggests that PfHSP110 serves as an important hub to coordinate protein quality control, survival, and immune evasion pathways in the parasite. Overall, our findings highlight potential accessory roles of PfHSP110 that may provide survival advantage to the parasite during its lifecycle and febrile conditions. The data also open avenues for experimental validation of auxiliary functions of PfHSP110 and their exploration for design of better antimalarial strategies.

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.

The Nodewalk assay to quantitate chromatin fiber interactomes in very small cell populations.

The chromosome conformation capture method and its derivatives, such as circularized chromosome conformation capture, carbon copy chromosome conformation capture, high-throughput chromosome conformation capture and capture high-throughput chromosome conformation capture, have pioneered our understanding of the principles of chromosome folding in the nucleus. These technical advances, however, cannot precisely quantitate interaction frequency in very small input samples. Here we describe a protocol for the Nodewalk assay, which is based on converting chromosome conformation capture DNA samples to RNA and subsequently to cDNA using strategically placed primers. This pipeline enables the quantitative analyses of chromatin fiber interactions without compromising its sensitivity down to <300 cells, making it suitable for MiSeq analyses of higher-order chromatin structures in biopsies, circulating tumor cells and transitional cell states, for example. Importantly, the quality of the Nodewalk sample can be assessed before sequencing to avoid unnecessary costs. Moreover, it enables analyses from hundreds of different restriction enzyme fragment viewpoints within the same initial small input sample to uncover complex, genome-wide networks. Following optimization of the different steps, the entire protocol can be completed within 2 weeks.

Systematic curation and analysis of genomic variations and their potential functional consequences in snoRNA loci.

Small nucleolar RNAs (snoRNAs) are a class of noncoding functional RNAs which are involved in RNA modifications, like methylation and pseudouridylation of other RNAs. The snoRNA species of RNAs are characterized by conserved structural motifs they harbor which are also intricately related to their functionality. Though there have been reports of the involvement of snoRNAs in disease processes and anecdotal reports of genomic variations in snoRNA loci and their effects in modulating snoRNA function, there has been no systematic collection and analysis of variations in snoRNA loci. In this manuscript, we present the most comprehensive curation of genomic single nucleotide variations in human snoRNA loci, and their systematic computational analysis to reveal potential single nucleotide variations which could have functional effects. We show six single nucleotide variations in snoRNA loci could significantly alter snoRNA structure and could have potential implications in their functions. The compilation is available at the snoRNA locus specific variation database: http://genome.igib.res.in/snolovd conforming to the HGVS standards for nomenclature of genomic variants.

Systematic analysis and functional annotation of variations in the genome of an Indian individual.

Whole genome sequencing of personal genomes has revealed a large repertoire of genomic variations and has provided a rich template for identification of common and rare variants in genomes in addition to understanding the genetic basis of diseases. The widespread application of personal genome sequencing in clinical settings for predictive and preventive medicine has been limited due to the lack of comprehensive computational analysis pipelines. We have used next-generation sequencing technology to sequence the whole genome of a self-declared healthy male of Indian origin. We have generated around 28X of the reference human genome with over 99% coverage. Analysis revealed over 3 million single nucleotide variations and about 490,000 small insertion-deletion events including several novel variants. Using this dataset as a template, we designed a comprehensive computational analysis pipeline for the systematic analysis and annotation of functionally relevant variants in the genome. This study follows a systematic and intuitive data analysis workflow to annotate genome variations and its potential functional effects. Moreover, we integrate predictive analysis of pharmacogenomic traits with emphasis on drugs for which pharmacogenomic testing has been recommended. This study thus provides the template for genome-scale analysis of personal genomes for personalized medicine.

Canonical WNT signaling-dependent gating of MYC requires a noncanonical CTCF function at a distal binding site.

Abnormal WNT signaling increases MYC expression in colon cancer cells in part via oncogenic super-enhancer-(OSE)-mediated gating of the active MYC to the nuclear pore in a poorly understood process. We show here that the principal tenet of the WNT-regulated MYC gating, facilitating nuclear export of the MYC mRNA, is regulated by a CTCF binding site (CTCFBS) within the OSE to confer growth advantage in HCT-116 cells. To achieve this, the CTCFBS directs the WNT-dependent trafficking of the OSE to the nuclear pore from intra-nucleoplasmic positions in a stepwise manner. Once the OSE reaches a peripheral position, which is triggered by a CTCFBS-mediated CCAT1 eRNA activation, its final stretch (≤0.7 µm) to the nuclear pore requires the recruitment of AHCTF1, a key nucleoporin, to the CTCFBS. Thus, a WNT/ß-catenin-AHCTF1-CTCF-eRNA circuit enables the OSE to promote pathological cell growth by coordinating the trafficking of the active MYC gene within the 3D nuclear architecture.

miRvar: A comprehensive database for genomic variations in microRNAs.

microRNAs are a recently discovered and well studied class of small noncoding functional RNAs. The regulatory role of microRNAs (miRNAs) has been well studied in a wide variety of biological processes but there have been no systematic effort to understand and analyze the genetic variations in miRNA loci and study its functional consequences. We have comprehensively curated genetic variations in miRNA loci in the human genome and established a computational pipeline to assess potential functional consequences of these variants along with methods for systematic curation and reporting of variations in these loci. The data is made available on the Leiden Open (source) Variation Database (LOVD) platform at http://genome.igib.res.in/mirlovd to provide ease of aggregation and analysis and is open for community curation efforts.

Author Correction: WNT signaling and AHCTF1 promote oncogenic MYC expression through super-enhancer-mediated gene gating.

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

SLTChemDB: A database of chemical compounds present in Smokeless tobacco products.

Smokeless tobacco (SLT), a cause of potentially preventable diseases, has a diverse chemical composition encompassing toxicants as well as potent carcinogens. Though the chemical profile of SLT products has been analyzed earlier, this information is not available in a comprehensive and easily accessible format. Hence, there is an imperative felt need to develop a one-stop information source providing inclusive information on SLT products. SLTChemDB is the first such database that makes available detailed information on various properties of chemical compounds identified across different brands of SLT products. The primary information for the database was extracted through extensive literature search, which was further curated from popular chemical web servers and databases. At present, SLTChemDB contains comprehensive information on 233 unique chemical compounds and 82 SLT products. The database has been made user-friendly with facility for systematic search and filters. SLTChemDB would provide the initial data on chemical compounds in SLT products to various tobacco testing laboratories. The database also highlights research gaps and thus, would be a guide for researchers interested in chemistry and toxicology of SLT products. With regular update of information in the database, it shall be a valuable evidence base for policymakers to formulate stringent policies for SLT control.

WNT signaling and AHCTF1 promote oncogenic MYC expression through super-enhancer-mediated gene gating.

WNT signaling activates MYC expression in cancer cells. Here we report that this involves an oncogenic super-enhancer-mediated tethering of active MYC alleles to nuclear pores to increase transcript export rates. As the decay of MYC transcripts is more rapid in the nucleus than in the cytoplasm, the oncogenic super-enhancer-facilitated export of nuclear MYC transcripts expedites their escape from the nuclear degradation system in colon cancer cells. The net sum of this process, as supported by computer modeling, is greater cytoplasmic MYC messenger RNA levels in colon cancer cells than in wild type cells. The cancer-cell-specific gating of MYC is regulated by AHCTF1 (also known as ELYS), which connects nucleoporins to the oncogenic super-enhancer via ß-catenin. We conclude that WNT signaling collaborates with chromatin architecture to post-transcriptionally dysregulate the expression of a canonical cancer driver.

OrCanome: a Comprehensive Resource for Oral Cancer.

Oral cancer is one of the most prevalent cancers in India but the underlying mechanisms are minimally unraveled. Cancer research has immensely benefited from genome scale high throughput studies which have contributed to expanding the volume of data. Such datasets also exist for oral cancer genes but there has been no consolidated approach to integrate the data to reveal meaningful biological information. OrCanome is one of the largest and comprehensive, user-friendly databases of oral cancer. It features a compilation of over 900 genes dysregulated in oral cancer and provides detailed annotations of the genes, transcripts and proteins along with additional information encompassing expression, inhibitors, epitopes and pathways. The resource has been envisioned as a one-stop solution for genomic, transcriptomic and proteomic annotation of these genes and the integrated approach will facilitate the identification of potential biomarkers and therapeutic targets.

HGV&TB: a comprehensive online resource on human genes and genetic variants associated with tuberculosis.

Tuberculosis (TB) is an infectious disease caused by fastidious pathogen Mycobacterium tuberculosis. TB has emerged as one of the major causes of mortality in the developing world. Role of host genetic factors that modulate disease susceptibility have not been studied widely. Recent studies have reported few genetic loci that provide impetus to this area of research. The availability of tools has enabled genome-wide scans for disease susceptibility loci associated with infectious diseases. Till now, information on human genetic variations and their associated genes that modulate TB susceptibility have not been systematically compiled. In this work, we have created a resource: HGV&TB, which hosts genetic variations reported to be associated with TB susceptibility in humans. It currently houses information on 307 variations in 98 genes. In total, 101 of these variations are exonic, whereas 78 fall in intronic regions. We also analysed the pathogenicity of the genetic variations, their phenotypic consequences and ethnic origin. Using various computational analyses, 30 variations of the 101 exonic variations were predicted to be pathogenic. The resource is freely available at http://genome.igib.res.in/hgvtb/index.html. Using integrative analysis, we have shown that the disease associated variants are selectively enriched in the immune signalling pathways which are crucial in the pathophysiology of TB. Database URL: http://genome.igib.res.in/hgvtb/index.html