The impact of epigenetic modulation on alternative splicing variants of genes involved in mitochondrial biogenesis in high-risk clusters of type 2 diabetes (EMAS)

Abstract

Novel classification using age, BMI, glycaemia, homeostasis model estimates, and islet autoantibodies proposed 5 subtypes of diabetes mellitus. Severe insulin-resistant diabetes (SIRD) and severe insulin-deficient diabetes (SIDD) were identified as high-risk clusters for development of early diabetes-related complications, however, the precise molecular mechanisms explaining elevated risk of complications of these clusters have not yet been elucidated. Differential usage of transcript isoforms, produced through alternative splicing, can alter the function of the resulting proteins. This process has been linked to the development of various disease and may partially explain the molecular mechanisms involved in early development of complications in T2D high risks groups. Like genetic factors, epigenetic mechanisms are proposed to potentially influence specific patterns of alternative splicing. Here, we investigated the interplay between epigenetic alterations and alternative splicing events in skeletal muscle of individuals with severe insulin-resistant diabetes. Additionally, one major goal was to focus on the impact of epigenetic alterations on alternatively spliced mitochondrial genes in SIRD. We collected skeletal muscle biopsies from 15 humans with SIRD as well as from 15 age-, sex- and BMI-matched glucose-tolerant controls to detect full-length transcriptome using Single-molecule Real-time Sequencing (SMRT-Seq) in an unbiased and highly accurate manner. In addition, in the muscle samples we also performed DNA methylation to investigate the potential impact of epigenetic factors on alternative splicing variants detected by the SMRT-Seq. While the data analysis on the SMRT-Seq data is still in progress, we identified close to 78,000 differentially methylated sites in skeletal muscle of individuals with SIRD when compared to CON group. About of ten percent of these sites were located in exons, which are crucial regions for alternative splicing events. Of note, 274 differentially methylated sites were located in exonic regions of mitochondrial genes. We are now proceeding with completing the data analysis from the SMRT-Seq and integrating them with the data from the DNA methylation. In turn, understanding the interplay between epigenetic modulation and alternative splicing may clarify the pathomechanism of early development of severe complications. In the future, these findings may enhance the identification of novel therapeutic targets for individuals belonging to specific diabetes subtype (SIRD) with elevated risk of diabetes-related complications and lead to better disease management.