Putative promoters within gene bodies control exon expression via TET1-mediated H3K36 methylation

Area of Science:

• Epigenetics
• Molecular Biology
• Cancer Genomics

Background:

• Gene promoter hypermethylation is linked to gene silencing and cancer, with DNA demethylating drugs like 5-aza-2'-deoxycytidine (DAC) used clinically.
• The presence of hypermethylated promoters correlating with high gene expression and the limited success of DAC highlight gaps in understanding DNA methylation's role in gene regulation.

Purpose of the Study:

• To investigate the mismatch between promoter methylation levels and gene expression.
• To identify and characterize novel promoter regions within gene bodies.
• To elucidate the role of TET1 and histone modifications in regulating gene expression through intragenic methylation.

Main Methods:

• RNA-sequencing (RNA-seq) and reduced representation bisulfite sequencing (RRBS) to analyze methylation and expression.
• Chromatin immunoprecipitation-quantitative polymerase chain reaction (ChIP-qPCR) and luciferase reporter assays to identify and test gene body promoter activity.
• Analysis of TET1, H3K36me3, 5-hydroxymethylcytosine (5hmC), MeCP2, and CREB1 interactions.

Main Results:

• A discrepancy was observed between promoter methylation levels and gene expression.
• Putative promoters were identified in gene bodies, and their activity was modulated by nearby CpG island (CGI) methylation.
• DAC effectively demethylated promoter CGIs but not gene body CGIs, while TET1 demethylated CGIs in both locations.
• A novel mechanism involving H3K36me3, 5hmC, MeCP2, and CREB1 was revealed to enhance SETD2 gene expression and H3K36me3 levels in gene bodies.

Conclusions:

• Gene bodies contain functional promoters that influence transcriptional activity.
• TET1 plays a crucial role in regulating gene expression by demethylating intragenic CGIs.
• These findings reveal a new layer of epigenetic regulation impacting gene expression and offer potential targets for cancer therapy.