Epigenomic Reprogramming in Inorganic Arsenic-Mediated Gene Expression Patterns During Carcinogenesis

Arsenic is a ubiquitous metalloid that is not mutagenic but is carcinogenic. The mechanism(s) by which arsenic causes cancer remain unknown. To date, several mechanisms have been proposed, including the arsenic-induced generation of reactive oxygen species (ROS). However, it is also becoming evident that inorganic arsenic (iAs) may exert its carcinogenic effects by changing the epigenome, and thereby modifying chromatin structure and dynamics. These epigenetic changes alter the accessibility of gene regulatory factors to DNA, resulting in specific changes in gene expression both at the levels of transcription initiation and gene splicing. In this review, we discuss recent literature reports describing epigenetic changes induced by iAs exposure and the possible epigenetic mechanisms underlying these changes

Genome-Wide DNA Methylation Reprogramming in Response to Inorganic Arsenic Links Inhibition of CTCF Binding, DNMT Expression and Cellular Transformation

Chronic low dose inorganic arsenic (iAs) exposure leads to changes in gene expression and epithelial-to-mesenchymal transformation. During this transformation, cells adopt a fibroblast-like phenotype accompanied by profound gene expression changes. While many mechanisms have been implicated in this transformation, studies that focus on the role of epigenetic alterations in this process are just emerging. DNA methylation controls gene expression in physiologic and pathologic states. Several studies show alterations in DNA methylation patterns in iAs-mediated pathogenesis, but these studies focused on single genes. We present a comprehensive genome-wide DNA methylation analysis using methyl-sequencing to measure changes between normal and iAs-transformed cells. Additionally, these differential methylation changes correlated positively with changes in gene expression and alternative splicing. Interestingly, most of these differentially methylated genes function in cell adhesion and communication pathways. To gain insight into how genomic DNA methylation patterns are regulated during iAs-mediated carcinogenesis, we show that iAs probably targets CTCF binding at the promoter of DNA methyltransferases, regulating their expression. These findings reveal how CTCF binding regulates DNA methyltransferase to reprogram the methylome in response to an environmental toxin

Quantitative Mass Spectrometry Reveals Changes in Histone H2B Variants as Cells Undergo Inorganic Arsenic-Mediated Cellular Transformation

Exposure to inorganic arsenic, a ubiquitous environmental toxic metalloid, leads to carcinogenesis. However, the mechanism is unknown. Several studies have shown that inorganic arsenic exposure alters specific gene expression patterns, possibly through alterations in chromatin structure. While most studies on understanding the mechanism of chromatin-mediated gene regulation have focused on histone post-translational modifications, the role of histone variants remains largely unknown. Incorporation of histone variants alters the functional properties of chromatin. To understand the global dynamics of chromatin structure and function in arsenic-mediated carcinogenesis, analysis of the histone variants incorporated into the nucleosome and their covalent modifications is required. Here we report the first global mass spectrometric analysis of histone H2B variants as cells undergo arsenic-mediated epithelial to mesenchymal transition. We used electron capture dissociation-based top-down tandem mass spectrometry analysis validated with quantitative reverse transcription real-time polymerase chain reaction to identify changes in the expression levels of H2B variants in inorganic arsenic-mediated epithelial-mesenchymal transition. We identified changes in the expression levels of specific histone H2B variants in two cell types, which are dependent on dose and length of exposure of inorganic arsenic. In particular, we found increases in H2B variants H2B1H/1K/1C/1J/1O and H2B2E/2F, and significant decreases in H2B1N/1D/1B as cells undergo inorganic arsenic-mediated epithelial-mesenchymal transition. The analysis of these histone variants provides a first step toward an understanding of the functional significance of the diversity of histone structures, especially in inorganic arsenic-mediated gene expression and carcinogenesis

The Role of Chromatin Structure in Circular RNA Biogenesis, Function and Disease

Fundamental to cellular differentiation, development, and physiology is the precise regulation of gene expression. At the center of this control lies chromatin, and disruptions in chromatin biology are key contributors to complex human diseases. By convention, gene expression analysis primarily focuses on transcript abundance, however the splicing of transcripts into distinct isoforms is also of biological relevance. One new form of alternative splicing, backsplicing, is responsible for generating circular RNAs (circRNAs), which are emerging as essential components to gene regulatory networks. However, unlike other forms of alternative splicing, the role that chromatin plays in backsplicing and circRNA formation remains poorly understood. This study examines the impact of chromatin on circRNA biogenesis and function. Using a Drosophila model, we investigate how the PARP1-chromatin complex affects backsplicing. Through native elongating transcript sequencing (NETseq), we uncover the role of PARP1 in regulating RNA polymerase II (RNAPII) pausing across circRNA host genes. Our findings reveal that PARP1-chromatin influences circRNAs biogenesis by altering RNAPII pausing within gene bodies, with outcomes contingent upon the architectural context of host genes. We find PARP1 regulates backsplicing by controlling RNAPII pausing, and the outcomes of this regulation are determined by the context of host gene architecture. Furthermore, PARP1\u27s influence extends to RNAPII pausing within introns and exons, influencing the direction of transcriptional output to favor either circRNA or mRNA. We next explore the pathological implications of disrupted chromatin dynamics, particularly in the context of Rett Syndrome, arising from mutations in the chromatin-associated protein MECP2 a PARP1 nucleosome competitor. Employing a single-cell multi-omics approach within patient-derived cortical spheroids, we illuminate cell-type-specific alterations in chromatin accessibility and gene expression mediated by mutant MECP2. These changes correlate with disturbances in the maturation of MEIS2+ neurons and identify potential circRNA candidates associated with these cells in the context of Rett syndrome. The functional presence of these circRNAs in RTT-associated cell types is being tested. Future mechanistic studies will determine how these dysregulated circRNAs function to drive Rett pathology. Finally, we asked how circRNA might function in an epigenetically driven disease—arsenic (iAs)-induced lung cancer. We show that the expression of SATB2, a 3D-chromatin organizer linking chromatin to the nuclear matrix, is upregulated by iAs exposure in bronchial epithelial cells along with its circRNA, circ3915. We show both SATB2 and circ3915 collaborate to promote cancer cell proliferation, migration, and carcinogenic gene expression pathways characterized by acquisition of oncogenic NFE2L2 and KRAS-like gene expression signature. We also show that circ3915 is translated in iAs-transformed cells, where it’s peptide associates with full-length SATB2 to modify chromatin associations in the nucleus. This discovery suggests that iAs triggers SATB2 expression, leading to alterations in chromatin states that govern gene expression to promote oncogenesis. This work advances our understanding of how genome architecture can provide context to fundamental biological processes such as splicing. Our identification of a circRNA that can be translated is exciting and demonstrates how circRNA translation adds to protein biodiversity and function. These findings not only broaden our comprehension of the intricate connections between genome architecture and fundamental biological processes like splicing but also highlight the translational potential of circRNA and its significance in precision medicine as a source of biomarkers and therapeutic targets