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

哺乳动物种系中性别特异性 DNA 甲基化特征获取的分子决定因素

• 批准号: 10723071
• 负责人: Rexxi Diptya Prasasya
• 金额: $ 12万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-01 至 2025-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10723071
• 关键词: Aberrant DNA Methylation; Advisory Committees; Age; Binding; Biochemical; Bioinformatics; Cell Reprogramming; Cells; Coupled; Cues; DNA; DNA Methylation; DNA Modification Methylases; Deposition; Development; Dioxygenases; Embryo; Embryonic Development; Environment; Enzymes; Epiblast; Epigenetic Process; Event; Exclusion; Exhibits; Female; Fertility; Fertilization; Gametogenesis; Genetic; Genetic Diseases; Genetic Models; Genome; Genomic Imprinting; Genomics; Germ Cells; Gonadal structure; Growth; Histones; Human Genetics; Hypermethylation; Knowledge; Length; Mentors; Mentorship; Methylation; Methyltransferase; Modeling; Modification; Molecular; Mus; Mutation; Nature; Oocytes; Outcome; Ovarian aging; Parents; Pathway interactions; Pattern; Phase; Post-Translational Protein Processing; Process; Reproduction; Research Personnel; Resistance; Role; Signal Pathway; Signal Transduction; Site; Somatic Cell; Specific qualifier value; Spermatogonia; Structure of primordial sex cell; Supporting Cell; Testing; Time; Tissues; Training; WNT Signaling Pathway; bisulfite sequencing; candidate identification; demethylation; epigenome; epigenomics; experimental study; extracellular; follow-up; genotypic sex; granulosa cell; idiopathic infertility; in vivo; infertility treatment; male; methylation pattern; methylome; mouse model; multiple omics; mutant; novel; nuclease; offspring; overexpression; postnatal; prevent; public health relevance; recruit; reproductive; sertoli cell; sex; sexual dimorphism; sperm cell; transcription factor; transcriptomics; transdifferentiation; transmission process; whole genome

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.

项目摘要 哺乳动物的繁殖需要两种遗传贡献，这是由于配子的高态性质 表观基因组。 DNA甲基化（DNAME）是配子中最性别的二态表观遗传标记之一， 在精子中高甲基化，并在卵母细胞中交替地甲基化和高甲基化。异常 种系中的dname可能会对生育能力和后代发展产生负面影响。防止传播 表演并建立种系命运，原始生殖细胞（PGC）经历全局dname擦除 以下规格。大多数基因组通过复制耦合被动实现脱甲基化 稀释，使用TET1酶的活性去甲基化途径是甲基化擦除需要的 基因座的子集。我最近发现，精子特异性的低甲基化区域虽然很少需要TET1 重新编程。组织特异性的低甲基化标志通常与发育的结合相关 相关的转录因子，贷款达到这些精子特异性降甲基化区域的重要性。 精子或卵母细胞如何获得性别特异性dname的机制仍然是一个知识差距，与 生育和发展。虽然生物化学组蛋白后翻译后修饰（PTM）已经 证明与DNA甲基转移酶（DNMT）可访问性相关，仍然未知这些 表观遗传标记在种系基因组中变得不均匀。我假设组蛋白 生殖线中的PTM富集和DNAME构图是1）通过脱甲基的本质确定 PGC重新编程期间使用的途径和2）由性腺支撑的信号毫升外在的途径 细胞。为了在体内检验这一假设，将使用遗传小鼠模型和多矩分析来阐明 哪些蜂窝信号负责获得精子中的性别特异性DNAME签名 卵母细胞。 AIM 1（K99）将在PGC开发过程中测试TET1的催化和非催化需求 建立卵母细胞基因组的甲基化特征。 AIM 2（R00）将采用遗传性别 - 颗粒前细胞（女性到男子）和组成性活性Wnt中DMRT1过表达的反转模型 在静脉前细胞（男性到女性）中的信号传导，以测试改变躯体信号环境的影响 用于生殖细胞中的DNAME获取。在这些模型中，我将集成并确定变化之间的相关性 在甲基组和相关组蛋白PTM富集（H3K4和H3K36的甲基化）中。单细胞 转录组学将用于识别建立性别特定DNAME签名的启发性线索 在配子中。在此期间，我将获得有关高级生物信息学和单细胞基因组学的广泛培训 在Bartolomei博士的指导下，该提案的指导阶段，Bartolomei博士，Dname和Genomic的先驱 在Upenn表观遗传学研究所内的印迹。在我的咨询委员会的额外指导下 其中包括生殖细胞表观遗传学和性腺信号通路领域的领导者，我会很好 准备成为现场生殖表观遗传学的独立研究者。