ABSTRACT: We report the whole-transcriptome profile with total RNA sequencing of postnatal male germ cells. Histone modification levels are dynamically controlled during mammalian spermatogenesis. We found that H3K9 demethylases, Jmjd1a and Jmjd1b catalyze H3K9 demethylation in prospermatogonia. Combined loss of Jmjd1 enzymes disturbed prospermatogonia to spermatogonia transition in mice. To examine a role of Jmjd1 in prospermatogonia to spermatogonia transition, we performed RNA-seq and ChIP-seq analyses using postnatal germ cells at P3 and P7.
Project description:We report the H3K9me2 distribution profile with ChIP sequencing of postnatal male germ cells. Histone modification levels are dynamically controlled during mammalian spermatogenesis. We found that H3K9 demethylases, Jmjd1a and Jmjd1b catalyze H3K9 demethylation in prospermatogonia. Combined loss of Jmjd1 enzymes disturbed prospermatogonia to spermatogonia transition in mice. To examine a role of Jmjd1 in prospermatogonia to spermatogonia transition, we performed RNA-seq and ChIP-seq analyses using postnatal germ cells at P3 and P7.
Project description:Histone H3 lysine 9 (H3K9) methylation is an epigenetic mark of transcriptionally repressed chromatin. During mammalian development, H3K9 methylation levels seem to be spatiotemporally regulated by the opposing activities of methyltransferases and demethylases to govern correct gene expression. However, the combination(s) of H3K9 methyltransferase(s) and demethylase(s) that contribute to this regulation and the genes regulated by them remain unclear. Herein, we demonstrate the essential roles of H3K9 demethylases Jmjd1a and Jmjd1b in the embryogenesis and viability control of embryonic stem (ES) cells. Mouse embryos lacking Jmjd1a/Jmjd1b died after implantation. Depletion of Jmjd1a/Jmjd1b in mouse ES cells induced rapid cell death accompanied with a massive increase in H3K9 methylation. Jmjd1a/Jmjd1b depletion induced an increase in H3K9 methylation in the gene-rich regions of the chromosomes, indicating that Jmjd1a/Jmjd1b removes H3K9 methylation marks in the euchromatin. Importantly, the additional disruption of the H3K9 methyltransferase G9a in a Jmjd1a/Jmjd1b-deficient background rescued not only the H3K9 hypermethylation phenotype but also the cell death phenotype. We also found that Jmjd1a/Jmjd1b removes H3K9 methylation marks deposited by G9a in the Oct4 and Ccnd1 loci to activate transcription. In conclusion, Jmjd1a/Jmjd1b ensures ES cell viability by antagonizing G9a-mediated H3K9 hypermethylation in the gene-rich euchromatin.
Project description:Establishment of spermatogonia throughout the fetal and postnatal period is essential for production of spermatozoa and male fertility. Here, we established a protocol for in vitro reconstitution of human prospermatogonial specification whereby human primordial germ cell (PGC)-like cells (hPGCLCs) differentiated from human induced pluripotent stem cells were further induced into M-prospermatogonia-like cells (MLCs) and T1 prospermatogonia-like cells (T1LCs) using long-term cultured xenogeneic reconstituted testes. Single cell RNA-sequencing was used to delineate the lineage trajectory leading to T1LCs, which closely resemble human T1-prospermatogonia in vivo and exhibited gene expression related to spermatogenesis and diminished proliferation, a hallmark of quiescent T1 prospermatogonia. Notably, this system enabled us to visualize the dynamic and stage-specific regulation of transposable elements during human prospermatogonial specification. Together, our findings pave the way for understanding and reconstructing human male germline development in vitro.
Project description:Efferent inhibition of cochlear outer hair cells is mediated by nicotinic cholinergic receptors containing alpha9 (a9) and alpha10 subunits. Mice lacking a9 nicotinic subunits fail to exhibit classic olivocochlear responses and are characterized by abnormal synaptic morphology at the base of outer hair cells. To detail molecular changes induced upon the loss of a9 subunit, we sampled cochlear RNA from wild type and a9 null mice at postnatal (P) days spanning periods of synapse formation and maturation (P3, P7, P13 and P60). Our findings point to a delay in cochlear maturation starting at the onset of hearing (P13), as well as an up-regulation of various GABA receptor subunits in adult mice lacking the a9 nicotinic subunit. Cochleae were removed at postnatal ages P3, P7, P13 and P60. Cochlear tissues from 3-5 mice were pooled per replicate; biological triplicates were performed for each age and genotype.
Project description:Spermatogonial differentiation is a developmental process that is essential for spermatogenesis, but the molecular and cellular changes that germ cells must undergo to transition from undifferentiated spermatogonia to differentiating spermatogonia remain largely undefined. Retinoic acid (RA) is necessary and sufficient for spermatogonial differentiation. Using the postnatal mouse testis, we examine the transcriptome changes that accompany spermatogonial differentiation. Spermatogenesis was synchronized by administration of potent and selective RA synthesis inhibitor; as a result, testes contained only undifferentiated spermatogonia. Then, the inhibitor was discontinued, and mice were given a single dose of exogenous RA to initiate spermatogonial differentiation. We measured transcriptomes in FACS-enriched germ cells either before RA administration, when the cells correspond to Aal spermatogonia (and a minor contribution of spermatogonial stem cells) or at two points after RA administration, when the cells correspond to A1 or A3 differentiating spermatogonia. The results of this study reveal the full transcriptome changes accompanying spermatogonial differentiation in the mouse.
Project description:In acute cold stress in mammals, JMJD1A, an H3K9 demethylase, up-regulates thermogenic gene expressions through β-adrenergic signaling in brown adipose tissue (BAT). Aside BAT-driven thermogenesis, mammals also have another mechanism to cope with long-term cold stress by inducing the browning of subcutaneous white adipose tissue (scWAT). Here, we show that this occurs through a two-step process that requires both β-adrenergic dependent phosphorylation of S265 and demethylation of H3K9me2 by JMJD1A. The histone demethylation independent acute Ucp1 induction in BAT and demethylation dependent chronic Ucp1 expression in beige-scWAT provide complementary molecular mechanisms to ensure an ordered transition between acute and chronic adaptation to cold stress. JMJD1A mediates two major signaling pathways, namlely β-adrenergic receptor and PPARγ activation, via PRDM16-PPARγ-P-JMJD1A complex for beige adipogenesis.
Project description:In acute cold stress in mammals, JMJD1A, an H3K9 demethylase, up-regulates thermogenic gene expressions through β-adrenergic signaling in brown adipose tissue (BAT). Aside BAT-driven thermogenesis, mammals also have another mechanism to cope with long-term cold stress by inducing the browning of subcutaneous white adipose tissue (scWAT). Here, we show that this occurs through a two-step process that requires both β-adrenergic dependent phosphorylation of S265 and demethylation of H3K9me2 by JMJD1A. The histone demethylation independent acute Ucp1 induction in BAT and demethylation dependent chronic Ucp1 expression in beige-scWAT provide complementary molecular mechanisms to ensure an ordered transition between acute and chronic adaptation to cold stress. JMJD1A mediates two major signaling pathways, namlely β-adrenergic receptor and PPARγ activation, via PRDM16-PPARγ-P-JMJD1A complex for beige adipogenesis.
Project description:MicroRNA is essential for the process of spermatogonesis, however analysis of its change in expression within germ cells during this process has been limited. We set out to examine the change in the miRNA expression profile of highly enriched mouse germ cell populations in vairous developmental stages.These inlcuded postnatal gonocytes, day 7-9 spermatogonia as well as pachytene spermatocytes and round spermatids from adult mice Gonocytes (day1) and spermatogonia (day 7-9), pachytene spermatocytes and round spermadits (adult) were enrichged by 2-4% BSA gradient sedementation. Three biological replicates of each cell type were included.