Promoter-Adjacent DNA Hypermethylation Can Downmodulate Gene Expression: TBX15 in the Muscle Lineage

Area of Science:

• Epigenetics
• Molecular Biology
• Gene Regulation

Background:

• TBX15 is a transcription factor crucial for differentiation.
• Promoter-adjacent DNA hypermethylation is observed in TBX15-expressing myoblasts and skeletal muscle.

Purpose of the Study:

• To investigate the functional role of DNA hypermethylation bordering the TBX15 promoter.
• To determine if this hypermethylation impacts gene expression in muscle cells.

Main Methods:

• Whole-genome bisulfite sequencing (WGBS) and enzymatic methyl-seq (EM-seq) to identify hypermethylated regions.
• Cloning differentially methylated regions (DMRs) into reporter vectors for promoter and enhancer assays.
• In vitro CpG methylation of cloned DMRs to assess its effect on activity.

Main Results:

• Hypermethylated regions flanking the TBX15 promoter exhibit promoter and enhancer activity in myoblasts.
• In vitro methylation of these regions abolished 86-100% of their activity.
• This promoter-border hypermethylation downmodulates, but does not silence, TBX15 expression in muscle cells.

Conclusions:

• DNA hypermethylation adjacent to the TBX15 promoter acts as a regulatory mechanism to fine-tune gene expression by suppressing enhancer/promoter activity.
• This epigenetic mark is specific to expressing cell types and differs from the repressive chromatin observed in silent cells.
• Promoter-adjacent DNA hypermethylation may be a more common regulatory strategy for preventing gene overexpression than previously understood, as seen in other TBX family genes.