Date of Award

Fall 1-1-2025

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Cell Biology

First Advisor

Lin, Haifan

Abstract

Epigenetic information refers to inheritable information that is not directly encoded in the genomic locus. Among various epigenetic mechanisms, DNA methylation play a crucial role in development and homeostasis and undergoes dynamic remodeling during mouse embryogenesis. In the mouse germline, primordial germ cells experience a global erasure of DNA methylation, a process important for the re-establishment of parent-specific methyl-imprint in the late stage of gametogenesis. However, how DNA methylation occurs at specific genomic loci remains an open question. Miwi2, one of three murine PIWI proteins, has been implicated in guiding de novo DNA methylation, yet the molecular machinery of Miwi2-mediated DNA methylation remains elusive. In this thesis, I investigate the possible mechanism of Miwi2-mediated DNA methylation. I show that both de novo DNA methyltransferase Dnmt3C and its cofactor Dnmt3L directly interact with Miwi2, via the ADD in Dnmt3C and 3L. Furthermore, Dnmt3C and Dnmt3L interact with Miwi2 in a mutually exclusive manner and assemble into a heterotetramer methyltransferase complex that bind to Spocd1. Loss of Spocd1 results in the increase of Dnmt3L deposition at 5' UTR regions of active LINE-1 (L1) elements in postnatal day 2 (P2) testes. Preliminary data further indicates that Spocd1 may modulate nucleosome accessibility. Based on these data, I propose that Miwi2 recruits Dnmt3C and Dnmt3L separately into the same genomic site, promoting the assembly of Dnmt3C/Dnmt3L methyltransferase complex. The methyltransferase complex is then anchored by Spocd1 to chromatin prior to the removal of H3K4me3 modification. Once H3K4me3 is removed, the complex binds with unmethylated H3 tails, releasing its autoinhibitory form and enabling de novo DNA methylation. This model provides an explanation for how epigenetic factors are guided to the specific genomic loci and how cells rapidly respond to changes in histone modifications.

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