Dynamic regulation and functions of locus-specific DNA methylation

Abstract

The role and regulation of DNA methylation at various genetic elements have gathered tremendous interest over decades. The methylomes of many cell types have been described, revealing a dynamic and tissue-specific pattern of DNA methylation (tissue-specific differentially methylated regions, T-DMRs) in the distal regulatory elements, such as enhancers. The formation of T-DMRs still remain mysterious, however, one of their interesting features observed in mouse ES cells (mESCs) is the low-to-intermediate levels of average DNA methylation resulted from inter-cellular epigenetic heterogeneity. Given the transcriptional repressive role of DNA methylation at promoters, such non-zero levels of enhancer methylation is interesting to characterize.

Prior to this thesis, a reporter for genomic DNA methylation (RGM) has been developed in the Jaenisch lab, when targeted into T-DMRs of interest, the surrounding locus-specific DNA methylation will be reported as on-and-off of fluorescent signals in single cells. We further modified RGM to investigate the regulation of DNA methylation at pluripotency super-enhancers Sox2 and MiR290 at single allele level in mESCs. We found that enhancer DNA methylation is surprisingly dynamic with two alleles independently being demethylated and methylated within days. Such dynamics is the basis of epigenetic and transcriptional heterogeneity and is coupled with changes in histone modifications and transcription factor binding. Furthermore, epigenetic heterogeneity was also observed in the developing preimplantation embryos. Our work provided a paradigm to functionally investigate locus-specific DNA methylation in heterogenous tissues in diseases and development.

The regulation of locus-specific DNA methylation is highly context dependent and sensitive to the environment. Our understanding of how locus-specific DNA methylation is regulated in vivo is still restricted to a few genomic elements. The appendix of this thesis attempts to generate an animal model to expand the scope of research on DNA methylation to retroelement-associated metastable epialleles.