The objective of this project is to elucidate the regulatory mechanisms and biological significance of the epigenetic programming of the sex chromosomes in the male germline. During male meiosis, unsynapsed sex chromosomes are epigenetically silenced in a process called meiotic sex chromosome inactivation (MSCI), which is necessary for spermatogenesis. During the term of the previous grant, we elucidated the underlying mechanisms of MSCI and demonstrated the role of DNA damage response (DDR) factors as essential regulators. Establishment of MSCI requires phosphorylation of the histone variant H2AX (?H2AX) to spread from the axes to the chromosome-wide domain of the sex chromosomes. This process is directed by MDC1, a binding partner of ?H2AX, at the onset of the pachytene stage. Downstream of MDC1, SCML2, a germline- specific Polycomb protein, is recruited to ?H2AX-containing nucleosomes and required for epigenetic programming. During the last project period, we unexpectedly found that initiation of MSCI is tightly coupled to active DNA demethylation. Initially the DNA demethylation is directed by MDC1 and precedes the establishment of silent histone modifications. In this renewal application, we will test the central hypothesis that the DDR pathway regulates active DNA demethylation, enabling the epigenetic programming of sex chromosomes necessary for male reproduction. While DNA methylation is generally associated with gene silencing, we propose that DNA demethylation is linked to gene silencing in MSCI. Our data suggest that demethylation in MSCI involves two major phases: the initial phase is mediated by the DDR pathway at the early pachytene stage (Aim 1) and the later phase is mediated by SCML2 downstream of the DDR at the mid- pachytene stage (Aim 2). This study will establish a novel link between DDR signaling and active DNA demethylation, and will further elucidate the biological significance of the epigenetic programming of the sex chromosomes, which is essential for male reproduction.

Public Health Relevance

The proposed research is relevant to public health because deficiencies in meiotic sex chromosome inactivation underlie male infertility. The proposed study potentially identifies fundamental mechanisms that regulate sex chromosomes. The outcome of this study will contribute to a better understanding of human male infertility and birth defects associated with dysregulation of sex chromosome-linked genes.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Research Project (R01)
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Cellular, Molecular and Integrative Reproduction Study Section (CMIR)
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Carter, Anthony D
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Cincinnati Children's Hospital Medical Center
United States
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Maezawa, So; Yukawa, Masashi; Alavattam, Kris G et al. (2018) Dynamic reorganization of open chromatin underlies diverse transcriptomes during spermatogenesis. Nucleic Acids Res 46:593-608
Maezawa, So; Hasegawa, Kazuteru; Yukawa, Masashi et al. (2018) Polycomb protein SCML2 facilitates H3K27me3 to establish bivalent domains in the male germline. Proc Natl Acad Sci U S A 115:4957-4962
Sakashita, Akihiko; Yeh, Yu-Han V; Namekawa, Satoshi H et al. (2018) Epigenomic and single-cell profiling of human spermatogonial stem cells. Stem Cell Investig 5:11
Maezawa, So; Hasegawa, Kazuteru; Alavattam, Kris G et al. (2018) SCML2 promotes heterochromatin organization in late spermatogenesis. J Cell Sci 131:
Abe, Hironori; Alavattam, Kris G; Kato, Yasuko et al. (2018) CHEK1 coordinates DNA damage signaling and meiotic progression in the male germline of mice. Hum Mol Genet 27:1136-1149
Maezawa, So; Alavattam, Kris G; Tatara, Mayu et al. (2018) A rapidly evolved domain, the SCML2 DNA-binding (SDB) repeats, contributes to chromatin binding of mouse SCML2. Biol Reprod :
Adams, Shannel R; Maezawa, So; Alavattam, Kris G et al. (2018) RNF8 and SCML2 cooperate to regulate ubiquitination and H3K27 acetylation for escape gene activation on the sex chromosomes. PLoS Genet 14:e1007233
Maezawa, So; Hasegawa, Kazuteru; Yukawa, Masashi et al. (2017) Polycomb directs timely activation of germline genes in spermatogenesis. Genes Dev 31:1693-1703
Zhang, Tingting; Du, Wei; Wilson, Andrew F et al. (2017) Fancd2 in vivo interaction network reveals a non-canonical role in mitochondrial function. Sci Rep 7:45626
Alavattam, Kris G; Kato, Yasuko; Sin, Ho-Su et al. (2016) Elucidation of the Fanconi Anemia Protein Network in Meiosis and Its Function in the Regulation of Histone Modifications. Cell Rep 17:1141-1157

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