Project description:Undifferentiated spermatogonia were sorted from Miwi2Tom/FL; ERT-Cre/+ (Miwi2-iKO) and Miwi2Tom/+; ERT-Cre/+ (Miwi2-CTL) mice injected with tamoxifen two weeks after last tamoxifen injection to understand an impact of acute Miwi2 deletion on spermatogenic homeostasis.
Project description:Spermatogonial stem cells are responsible for sustaining gametes production and male fertiliy in men. However, germ cells including SSCs are highly sensitive to chemotherapies and radiations, placing male cancer patients at high risk of treatment-induce infertility. We have previously shown that the undifferentiated spermatogonial population of mouse testis is functionally heterogeneous and contain both stem cells and committed progenitor cells. Using mouse model of chemotherapy-induced germline damage and recovery, we aim to define molecular charactersitics, dynamics and heterogeneity of undifferentiated spermatogonia during germline regeneration compared to homeostatic undifferentiated spermatogonia.
Project description:To identify the transcripts preferentially expressed in type A spermatogonia in rainbow trout immature testis, we compared the transcriptome between type A spermatogonia and testicular somatic cells by microarry analysis. We used fluorescence-activated cell sorting (FACS) to isolate type A spermatogonia or testicular somatic cells from the fishes carrying the transgene, pvasa-GFP, which expresses GFP in spermatogonia. RNA from type A spermatogonia and from testicular somatic cells were hybridized to a microarray after Cy3 labeling.
Project description:Microarray analysis was performed in order to detail the gene expression profiles in murine b-2M-SPa-6+c-kit-undifferentiated and b-2M-SPa-6+c-kit+ differentiating spermatogonia. These data were used to compare human and mouse transcriptomes of undifferentiated spermatogonia.
Project description:The environmental exposures and lifestyle of parents can alter the development of offspring. How this environmental information is coded into heritable messages to be transmitted by gametes remains unknown, but epigenetic mechanisms have been implicated. We recently determined that disruption of histone H3 di-methylation at lysine 4 (H3K4me2) in sperm has transgenerational consequences in the development of offspring. However, little is known about when in spermatogenesis histone methylation is established and whether epimutations induced in developing sperm are permanent. We hypothesize that epigenetic modifications to histones established in spermatogonia persist through spermatogenesis and can be transmitted to offspring via the sperm. Our objective was to determine what genomic regions bearing histone H3K4me2 in spermatogonia are also present in sperm. Methods: Using transgenic mice expressing Oct4-GFP, we isolated an enriched spermatogonia population and performed ChIP-seq for H3K4me2. Results: Our analysis revealed that H3K4me2 is located throughout the genome in spermatogonia and particularly at the transcription start site region (TSS) of more than 16,000 genes. Remarkably 44% of H3K4me2 peaks present in spermatogonia are conserved in sperm. The greatest overlap between spermatogonia and sperm occurred at the TSS with 83% similarity. Finally, we assessed the expression level of genes enriched in H3K4me2 in spermatogonia and sperm. We observed that genes with the highest enrichment in H3K4me2 in sperm are expressed at higher levels in spermatogenesis and during development. Conclusion: These findings suggest that if epimutations are induced in spermatogonia they may persist in sperm and influence the health and development of offspring.
Project description:Purpose: The goal of this sequencing is to investigate alterations in gene expression that result from impaired retinoid signaling compared with control, and how the RA signaling controls spermatogonia differentiation Methods: THY1+ spermatogonia mRNA profiles of 4-day-old control and germ cell specific impaired retinoid signaling mice were generated by High-throughput sequencing Results: Gene ontology (GO) analysis of the genes at the top of the ranked genes indicated enrichment in genes associated with roles in reproduction, transcription and spermatogenesis. In total, we identified 1633 and 742 transcripts (Reads Per Kilobase of transcript per Million mapped reads (RPKM) > 1) that were significantly (p-value < 0.05, > 1.5-fold difference) down- and up-regulated, respectively, in the germ cell mutants compared with the controls. Most importantly, we found that the majority of transcripts of replication-dependent core histone genes, histone cluster 1 (Hist1) were downregulated in germ cell mutants. THY1+ spermatogonia mRNA profiles of 4-day old germ cell specific impaired retinoid signaling and control mice were generated by deep sequencing, twice, using Illumina HiSeq 2000