Altered chromatin landscape and 3D interactions associated with primary constitutional MLH1 epimutations

Area of Science:

• Genetics and Epigenetics
• Cancer Predisposition Syndromes
• Molecular Biology

Background:

• Lynch syndrome (LS) increases cancer risk, often due to mismatch repair gene variants.
• Constitutional MLH1 epimutation (CME) is a less common cause of LS, involving MLH1 promoter methylation.
• Primary CMEs arise de novo and can be passed between generations, with unknown underlying mechanisms.

Purpose of the Study:

• To investigate the genetic and gene regulatory alterations in primary CME.
• To elucidate the molecular mechanisms behind the origin and maintenance of primary CME.

Main Methods:

• Utilized linked-read whole-genome sequencing (WGS) to identify genetic alterations in CME carriers.
• Employed RNA-sequencing (RNA-seq), ATAC-seq, H3K27ac CUT&Tag, and UMI-4C to analyze gene expression, chromatin accessibility, and 3D chromatin interactions.
• Used a specific MLH1 promoter SNP (rs1800734) as a reporter to assess allele-specific chromatin conformation.

Main Results:

• Epimutant MLH1 alleles exhibited closed chromatin conformation and reduced H3K27ac levels compared to unmethylated alleles.
• Demonstrated differential 3D chromatin contacts in the epimutant MLH1 promoter, including altered interactions with distal regulatory elements.
• Identified rare genetic alterations in promoter-contacting regions of CME carriers, potentially impacting transcription factor binding.

Conclusions:

• Primary CMEs display allele-specific differences in interactions with neighboring genes and regulatory elements.
• The identified cis-regulatory regions may play a role in the origin and maintenance of CME.
• Further research is needed to fully understand the molecular mechanisms underlying primary CME.