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Application of an allele-specific pipeline to study DNA methylation inheritance and dynamics in the early embryo

University of British Columbia

Classic genomic imprinted loci display parent-of-origin transcription in adult cells. Such allele-specific expression is thought to be driven by epigenetic marks, including DNA methylation (DNAme) and histone modifications, established in the gametes. However, the extent of monoallelic transcription in early mammalian development has remained relatively unexplored. As chromatin is highly dynamic before implantation, parent-of- origin directed transcription may be particularly high at this stage. To identify parent-of-origin controlled transcription, including during early embryogenesis, I helped develop MEA, a Methylomic and Epigenomic Allele-specific analysis pipeline designed to integrate whole genome bisulphite (WGBS)-, RNA- and ChIP-sequencing datasets. To demonstrate the utility and sensitivity of MEA, I processed existing mouse and human datasets and uncovered classical as well as novel candidate imprinted genes. Subsequently, I applied MEA to analyze WGBS and ChIP-seq data generated from adult gametes and preimplantation mouse embryos, which yielded female/maternal and male/paternal epigenomic profiles. Surprisingly, despite global DNAme loss following fertilization, I uncovered several dozen CpG island promoters that are de novo methylated on the paternal genome in 2-cell embryos, coincident with H3K4me3 loss on the same allele. A subset of these loci is hypermethylated in androgenetic blastocysts but hypomethylated in parthenogenetic blastocysts, confirming the paternal genome is susceptible to post-fertilization DNAme activity. Notably, this zygotic paternal DNAme gain was ablated following genetic depletion of maternal DNMT3A. Parental DNAme levels at these loci are harmonized in the post-implantation embryo and beyond: thus, unlike classic imprinted regions, these novel DMRs are established on the paternal genome by maternal stores of DNMT3A in the zygote and likely only transiently impart allelic transcription in the embryo. Indeed, expression analysis of DNMT3A maternal KO preimplantation embryos revealed that genes normally gaining paternal DNAme following fertilization are prematurely activated from the paternal allele in 4C embryos. Taken together, the results presented in my thesis illustrate the benefits of increasing the range and sensitivity of allele-specific analyses, and uncover zygotic de novo DNAme activity at CpG island promoters on the paternal genome against a backdrop of acute global demethylation.

Text

2020-04-22

Attribution-NonCommercial-NoDerivatives 4.0 International

http://hdl.handle.net/2429/74111

Medical Genetics

University of British Columbia

UBCV

http://creativecommons.org/licenses/by-nc-nd/4.0/