Molecular determinants of sex-specific DNA methylation signature acquisition in the mammalian germline

点击翻译按钮获取中文摘要

Project abstract Mammalian reproduction requires biparental genetic contributions due to the highly dimorphic nature of gamete epigenomes. DNA methylation (DNAme) is one of the most sexually dimorphic epigenetic marks in gametes, being hypermethylated in the sperm and alternatingly hypo- and hypermethylated in the oocyte. Aberrant DNAme in the germline can negatively impact fertility and offspring development. To prevent transmission of epimutations and establish the germline fate, primordial germ cells (PGCs) undergo global DNAme erasure following specification. While most of the genome achieves demethylation through replication-coupled passive dilution, the active demethylation pathway using the TET1 enzyme is required for methylation erasure of a subset of loci. I recently discovered that sperm-specific hypomethylated regions, while rare, require TET1 for reprogramming. Tissue-specific hypomethylation signatures often correlate with binding of developmentally relevant transcriptional factors, lending to the significance of these sperm-specific hypomethylated regions. Mechanisms of how sperm or oocytes acquire sex-specific DNAme remain a knowledge gap with relevance to fertility and development. While biochemically histone post-translational modifications (PTMs) have been shown to correlate with DNA methyltransferase (DNMT) accessibility, it remains unknown how these epigenetic marks become non-uniformly enriched within the germline genome. I hypothesize that histone PTMs enrichment and DNAme patterning in the germline are determined 1) intrinsically by the demethylation pathway used during PGC reprogramming and 2) extrinsically by the signaling milleu of the gonadal supporting cells. To test this hypothesis in vivo, genetic mouse models and multi-omics analyses will be used to elucidate what cellular signals are responsible for the acquisition of sex-specific DNAme signatures in the sperm and the oocyte. Aim 1 (K99) will test the catalytic and non-catalytic requirement for TET1 during PGC development for the establishment of the methylation signature of the oocyte genome. Aim 2 (R00) will employ genetic sex- reversal models of Dmrt1 overexpression in pre-granulosa cells (female-to-male) and constitutively active Wnt signaling in pre-Sertoli cells (male-to-female) to test the impact of altering the somatic signaling environment for DNAme acquisition in germ cells. In these models, I will integrate and identify correlations between changes in methylome and the relevant histone PTMs enrichment (methylation of H3K4 and H3K36). Single cell transcriptomics will be used to identify instructive cues for the establishment of sex-specific DNAme signatures in gametes. I will receive extensive training in advanced bioinformatics and single-cell genomics during the mentored phase of this proposal under the mentorship of Dr. Bartolomei, a pioneer in DNAme and genomic imprinting, within the UPenn Epigenetics Institute. With the additional guidance from my advisory committee, which includes leaders in the field of germ cell epigenetics and gonadal signaling pathways, I will be well prepared to become an independent investigator in the field reproductive epigenetics.