| Summary: | 博士 === 國立臺灣大學 === 生物科技研究所 === 102 === The establishment and rearrangement of proper germline epigenome is crucial for generating functional germ cells. Accumulating results demonstrated that the ability of prosprematogonia and spermatogonial stem/progenitor cells (SPCs) to reside in a quiescent state is important for prospermatogonia differentiation and preventing premature exhaustion of the postnatal stem cell pool. However, the intrinsic epigenetic factors that regulate SPC quiescence are largely unknown. Here, we investigated how DNA methyltransferase 3-like (DNMT3L), an epigenetic regulator important for interpreting chromatin context and facilitating de novo DNA methylation, sustains the long-term male germ cell pool. We demonstrated that stem cell-enriched THY1+ SPCs constitute a DNMT3L-expressing population in the postnatal testes. DNMT3L influenced the stability of PLZF, potentially by downregulating Cdk2/CDK2 expression, which sequestered CDK2-mediated PLZF degradation. Increased level of ETS1 was found to bind the DNA hypomethylated and H3K4me3-over-accumulated Cdk2 promoter that extended Cdk2/CDK2 expression in 8 dpp Dnmt3l KO THY1+ cells. In addition, CDK2 interacted with PLZF in THY1+ cells and PLZF was excessively post-translational modification with ubiquitin in Dnmt3l KO THY1+ cells, suggesting that DNMT3L participates in the regulation of PLZF stability in postnatal germ cells. Reduced PLZF in Dnmt3l KO THY1+ cells released its antagonist, SALL4A, which overactivated ERK and AKT signaling cascades to stimulate SPC proliferation. These results indicate that DNMT3L is required for preventing premature SPC exhaustion and to delicately balance the cycling and quiescence of SPCs.
Since a well-known function of DNMT3L is to repress transposable elements (TEs) in order to safeguard the integrity of the genome. PiRNAs are highly involved in TE silencing in male germ cells. Further analysis of piRNA-related components revealed that Dnmt3l KO embryonic germ cells display detachment of some cytoplasmic MAEL from P-body components in E18.5 when genome-wide transposon silencing is undergoing. Furthermore, we observed that postnatal Dnmt3l KO THY1+ germ cells exhibit less MAEL protein in the cytoplasmic fraction compared to wild-type THY1+ germ cells. The aberrant MAEL expression and subcellular distribution may account for the abnormal characteristics of piRNA compositions in Dnmt3l KO germ cells.
Cumulatively, these findings provide novel roles for DNMT3L in mouse male germ cells. DNMT3L is important for germline maintenance via influencing SPC self-renewal/differentiation-associated factors and the piRNA-related pathway.
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