Abstract
Purpose :
A key epigenetic regulator of gene expression is gene silencing through DNA methylation at specific cytosines (mCG). mCG methylation governs chromatin configurations and DNA accessibility changes that modulate transcription factor function. It is well established that mCG methylation regulates the activities of individual genes and their cognate transcription factors but few studies have examined its role in global biological processes or employed it as a tool to predict key transcription factors. Here, we employed a multiomics analysis to establish a requirement for mCG methylation in gene expression events that hallmark lens cell differentiation.
Methods :
The genome-wide relationships between mCG methylation, chromatin accessibility and transcript levels were compared between lens epithelial cells and fiber cells of embryonic day 13 chick lenses through integrated analysis of bisulfite sequencing, ATAC Seq and RNA Seq data. Bioinformatic interrogation of the data was used to predict functional DNA regulatory sites and transcription factor consensus representation across the chick genome.
Results :
7621 genomic loci exhibiting significant differences in mCG levels between lens epithelial and fiber cells were correlated with the differentiation state-specific expression of 1285 genes preferentially expressed in either lens fiber or lens epithelial cells (Pearson r = -0.37, p < 1x10-42). mCG levels were inversely correlated with chromatin accessibility as determined by Assay for Transposase-Accessible Sequencing (ATAC-seq) (Pearson r = -0.86, p < 1x10-300). Genes exhibiting altered regions of DNA methylation, chromatin accessibility and gene expression levels in fiber cells relative to epithelial cells include crystallins (CRYBA4, CRYBB1, CRYGN, CRYBB2), lens beaded filament proteins (BFSP1, BFSP2), transcription factors (HSF4, SOX2, HIF1A), and Notch signaling. Regions exhibiting cell-type specific DNA methylation changes were enriched with transcription factor binding sequences including HIF1A, SOX2, and the MAF family of transcription factors.
Conclusions :
The results demonstrate that gene silencing through mCG methylation changes control chromatin accessibility and gene expression events that are required for lens differentiation. They also point to a role for mCG methylation in the regulated binding of transcription factors important for lens cell differentiation.
This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.