Abstract
BackgroundDNA methylation is an epigenetic modification that mainly repress expression of genes essential during embryogenesis and development. There are key ATPase-dependent enzymes that read or write DNA methylation to remodel chromatin and regulate gene expression. Structural maintenance of chromosome hinge domain containing 1 (SMCHD1) is an architectural protein that regulates expression of numerous genes, some of which are imprinted, that are sensitive to DNA methylation. In addition, SMCHD1 germline mutations lead to developmental diseases; facioscapulohumoral muscular dystrophy (FSHD), bosma arhinia and micropthalmia (BAMS). Current evidence suggests that SMCHD1 functions through maintenance or de novo DNA methylation required for chromatin compaction. However, it is unclear if DNA methylation is also essential for genomic recruitment of SMCHD1 and its role as an architectural protein. We previously isolated SMCHD1 using a methylated DNA region from mouse pituitary growth hormone (Gh1) promoter, suggesting that methylation is required for SMCHD1 DNA binding. The goal of this study was to further understand DNA methylation directed role of SMCHD1 in regulating gene expression. Therefore, we profiled SMCHD1 genome wide occupancy in human neuroblastoma SH-SY5Y cells and evaluated if DNA methylation is required for SMCHD1 genomic binding by treating cells with the DNA demethylating reagent, 5-azacytidine (5-azaC).ResultsOur data suggest that the majority of SMCHD1 binding occurs in intron and intergenic regions. Gene ontology analysis of genes associated with SMCHD1 genomic occupancy that is sensitive to 5-azaC treatment suggests SMCHD1 involvement in central nervous system development. The potassium voltage-gated channel subfamily Q member1 (KCNQ1) gene that associates with central nervous system is a known SMCHD1 target. We showed SMCHD1 binding to an intronic region of KCNQ1 that is lost following 5-azaC treatment suggesting DNA methylation facilitated binding of SMCHD1. Indeed, deletion of SMCHD1 by CRISPR- Cas9 increases KCNQ1 gene expression confirming its role in regulating KCNQ1 gene expression.ConclusionThese findings provide novel insights on DNA methylation directed function of SMCHD1 in regulating expression of genes associated with central nervous system development that impact future drug development strategies.
Highlights
DNA methylation is an epigenetic modification that mainly repress expression of genes essential during embryogenesis and development
Using chromatin immunoprecipitation in combination with massively parallel sequencing (ChIP-seq), we obtained information on genomic sites bound by SMCHD1 with high resolution and identified SMCHD1 recruitment sites that are sensitive to DNA methylation using 5-azaC
To investigate DNA methylation dependency of SMCHD1 in regulating gene expression, we sought to compare SMCHD1 genomic localization in SH-SY5Y neuroblastoma cells cultured under normal conditions and those treated with 5-azaC to induce global loss of DNA methylation
Summary
DNA methylation is an epigenetic modification that mainly repress expression of genes essential during embryogenesis and development. Structural maintenance of chromosome hinge domain containing 1 (SMCHD1) is an architectural protein that regulates expression of numerous genes, some of which are imprinted, that are sensitive to DNA methylation. Current evidence suggests that SMCHD1 functions through maintenance or de novo DNA methylation required for chromatin compaction. It is unclear if DNA methylation is essential for genomic recruitment of SMCHD1 and its role as an architectural protein. The most consistent signature of BAMS individuals is the complete absence of a nose which might accompany other malformations [12] These findings demonstrate that as an epigenetic modifier, SMCHD1 impacts gene regulation of multiple genomic regions which can result in severe diseases. It is still unclear how SMCHD1 works within the epigenetic machinery
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