SYMPK Is Required for Meiosis and Involved in Alternative Splicing in Male Germ Cells

Abstract

SYMPK is a scaffold protein that supports polyadenylation machinery assembly on nascent transcripts and is also involved in alternative splicing in some mammalian somatic cells. However, the role of SYMPK in germ cells remains unknown. Here, we report that SYMPK is highly expressed in male germ cells, and germ cell-specific knockout (cKO) of Sympk in mouse leads to male infertility. Sympk cKODdx4–cre mice showed reduced spermatogonia at P4 and almost no germ cells at P18. Sympk cKOStra8–Cre spermatocytes exhibit defects in homologous chromosome synapsis, DNA double-strand break (DSB) repair, and meiotic recombination. RNA-Seq analyses reveal that SYMPK is associated with alternative splicing, besides regulating the expressions of many genes in spermatogenic cells. Importantly, Sympk deletion results in abnormal alternative splicing and a decreased expression of Sun1. Taken together, our results demonstrate that SYMPK is pivotal for meiotic progression by regulating pre-mRNA alternative splicing in male germ cells.

Differential susceptibility of male and female germ cells to glucocorticoid-mediated signaling

“…GO term analysis on differentially expressed genes in the germ cells following dex treatment revealed a strong enrichment for genes related to the regulation of mRNA splicing ( Figure 5—figure supplement 1A ). The regulation of RNA splicing in germline cells is crucial for the proper progression of meiosis and spermatogenesis ( Kuroda et al, 2000 ; Li et al, 2007 ; O’Bryan et al, 2013 ; Schmid et al, 2013 ; Zagore et al, 2015 ; Naro et al, 2017 ; Liu et al, 2017 ; Xu et al, 2017 ; Horiuchi et al, 2018 ; Ehrmann et al, 2019 ; Legrand et al, 2019 ; Yuan et al, 2021 ; Wu et al, 2021 ), and is conserved across species ( Mattox and Baker, 1991 ; Wu et al, 2016 ; Chen et al, 2019 ). Downregulated genes also showed an enrichment for mitotic cell cycle progression, which is of interest at this PN1 time point given that the mitotically-arrested male germ cells will begin to re-enter mitosis at approximately PN2 ( Vergouwen et al, 1991 ; Drumond et al, 2011 ; Nagano et al, 2000 ).…”

“…Given this link between GR signaling and splicing and the known role of transcript splicing in regulating spermatogenesis ( Kuroda et al, 2000 ; Li et al, 2007 ; O’Bryan et al, 2013 ; Schmid et al, 2013 ; Zagore et al, 2015 ; Naro et al, 2017 ; Liu et al, 2017 ; Xu et al, 2017 ; Horiuchi et al, 2018 ; Ehrmann et al, 2019 ; Legrand et al, 2019 ; Yuan et al, 2021 ; Wu et al, 2021 ), we next sought to assess whether loss of GR would impact male fertility. Because full body deletion of GR results in lethality at birth due to defects in lung maturation ( Cole et al, 1995 ), we generated a conditional deletion of GR in the germline using a transgenic Prdm1- Cre line ( Ohinata et al, 2005 ).…”

“…To our knowledge, this is the first time GR has been implicated in regulating the expression of genes involved in RNA splicing. Interestingly, it has been well documented in the literature that a full genetic deletion of a single splice factor leads to impairments in spermatogenesis, and ultimately infertility ( Kuroda et al, 2000 ; Li et al, 2007 ; O’Bryan et al, 2013 ; Zagore et al, 2015 ; Liu et al, 2017 ; Xu et al, 2017 ; Horiuchi et al, 2018 ; Ehrmann et al, 2019 ; Legrand et al, 2019 ; Yuan et al, 2021 ; Wu et al, 2021 ). We suspect that dex treatment leads to fewer differential splicing events than a full splice factor deletion, given that dex treatment causes a broader decrease in splice factor expression without entirely abolishing any single splice factor.…”

Differential susceptibility of male and female germ cells to glucocorticoid-mediated signaling

“…GO term analysis on differentially expressed genes in the germ cells following dex treatment revealed a strong enrichment for genes related to the regulation of mRNA splicing ( Figure S5A ). The regulation of RNA splicing in germline cells is crucial for the proper progression of meiosis and spermatogenesis 46–58 , and is conserved across species 59–61 . Downregulated genes also showed an enrichment for mitotic cell cycle progression, which is of interest at this PN1 time point given that the mitotically-arrested male germ cells will begin to re-enter mitosis at approximately PN2 62–64 .…”

“…To our knowledge, this is the first time GR has been implicated in regulating the expression of genes involved in RNA splicing. Interestingly, it has been well documented in the literature that a full genetic deletion of a single splice factor leads to impairments in spermatogenesis, and ultimately infertility [46][47][48]50,[52][53][54][55][56][57][58] . We suspect that dex treatment leads to fewer differential splicing events than a full splice factor deletion, given that dex treatment causes a broader decrease in splice factor expression without entirely abolishing any single splice factor.…”