Poster abstracts
Poster number 103 submitted by Vishal Nanavaty
DNA methylation regulates alternative polyadenylation via CTCF and the cohesin complex
Vishal Nanavaty (Genomic Medicine, Lerner Research Institute, Cleveland Clinic, Cleveland, OH 44195, USA), Elizabeth W. Abrash (Genomic Medicine, Lerner Research Institute, Cleveland Clinic, Cleveland, OH 44195, USA), Changjin Hong, Sunho Park, Tae Hyun Hwang (Quantitative Health Sciences, Lerner Research Institute, Cleveland Clinic, Cleveland, OH 44195, USA), Thomas J. Sweet, Jeffrey M. Bhasin, Srinidhi Singuri (Genomic Medicine, Lerner Research Institute, Cleveland Clinic, Cleveland, OH 44195, USA), Byron H. Lee (Cardiovascular & Metabolic Sciences, Lerner Research Institute, Glickman Urologic Institute, Cleveland Clinic, Cleveland, OH 44195, USA), Angela H. Ting (Genomic Medicine, Lerner Research Institute, Cleveland Clinic, Cleveland, OH 44195, USA, Department of Molecular Medicine, Cleveland Clinic Lerner College of Medicine, Case Western Reserve University)
Abstract:
Dysregulation of DNA methylation and mRNA alternative cleavage and polyadenylation (APA) are both prevalent in cancer, but they have been studied as independent processes. While promoter hypermethylation is known to silence transcription of tumor suppressor genes, functions of DNA methylation in non-promoter regions are poorly understood, hindering a complete understanding of the biological impact of aberrant methylation in tumorigenesis and cancer progression. We discovered a DNA methylation-regulated APA mechanism when we compared genome-wide DNA methylation (MBD-seq), gene expression (RNA-seq) and polyadenylation (poly(A)) site usage between DNA methylation-competent HCT116 and DNA methylation-deficient DKO cells. Using RNA-seq and DNA methylation data from The Cancer Genome Atlas (TCGA) for 11 cancer types, we authenticated the relationship between DNA methylation and mRNA polyadenylation isoform expression in vivo. Through sequence motif analysis and chromatin immunoprecipitation (ChIP), we identified and confirmed CCCTC-binding factor (CTCF), which binds DNA in a DNA methylation-sensitive manner, to be preferentially recruited to unmethylated DNA downstream of the proximal poly(A) site. Removal of DNA methylation enables CTCF binding and recruitment of the cohesin complex, which in turn, promotes proximal polyadenylation site usage. In this DNA de-methylated context, depletion of the RAD21 a cohesin complex component can recover distal poly(A) site usage. Deletion of CTCF binding motifs via CRISPR/CAS9 highlights the importance of CTCF and cohesin complex proteins in chromatin loop formation and alternative polyadenylation. This DNA methylation-regulated APA mechanism demonstrates how aberrant DNA methylation impacts transcriptome diversity and highlights the potential sequelae of global DNA methylation inhibition as a cancer treatment.
Keywords: DNA methylation, Alternative polyadenylation, Chromatin loops