Project description:This is a study to explore the transcriptional changes after Adjudin treatment in adult rat testes at three time points (control, 8 hour and 4 day). Adjudin, [1-(2,4-dichlorobenzyl)-1H-indazole-3-carbohydrazide], is a potential male contraceptive that targets the Sertoli-germ cell interface and causes germ cell depletion from the seminiferous epithelium. Adjudin has been proved to be a useful model to study the mechanisms that regulate junction restructuring in the testis. Keywords: Sertoli-germ cell adhesion regulation using an Adjudin model
Project description:To study the dynamic of DNA methylation during male gametogenesis 9 stages of germ cells were purified, then DNA samples (n=4/stage) were analyzed by Rat Methyl seq capture
Project description:Spermatogenesis is a complex process, dependent upon the successive activation and/or repression of thousands of gene products, and ends with the production of haploid male gametes. RNA sequencing of male germ cells in the rat identified thousands of novel testicular unannotated transcripts, named TUTs. Although such RNAs are usually annotated as long non-coding RNAs, it is possible that some of these TUTs code for protein. To test this possibility, we used a “Proteomics Informed by Transcriptomics” strategy, combining shotgun proteomics analyses and RNA sequencing data for enriched populations of rat testicular cells. Among 3559 TUTs and 506 lncRNAs found in meiotic and post-meiotic germ cells, 44 encoded at least one peptide. We show that these novel high-confidence protein-coding loci exhibit several genomic features intermediate between those of lncRNAs and mRNAs. We experimentally validated the testicular expression pattern of two of these novel protein-coding gene candidates, both highly conserved in mammals: one for a vesicle-associated membrane protein, we named VAMP-9, and the other for an enolase domain-containing protein. This study confirms the potential of PIT approaches for the discovery of protein coding transcripts initially thought to be untranslated, or unknown transcripts. Our results contribute to the understanding of spermatogenesis by characterizing two novel proteins, implicated by their strong expression in germ cells.
Project description:In mammals, pluripotent cells transit through a continuum of distinct molecular and functional states en route to initiating lineage specification. Capturing pluripotent stem cells (PSCs) mirroring in vivo pluripotent states provides accessible in vitro models to study the pluripotency program and mechanisms underlying lineage restriction. Here, we develop optimal culture conditions to derive and propagate post-implantation epiblast-derived PSCs (EpiSCs) in rats, a valuable model for biomedical research. We show that rat EpiSCs can be reset toward the naïve pluripotent state with exogenous Klf4, albeit not with the other five candidate genes (Nanog, Klf2, Esrrb, Tfcp2l1, and Tbx3) effective in mice. Finally, we demonstrate that rat EpiSCs retain competency to produce authentic primordial germ cell-like cells that contribute to functional gametogenesis leading to the birth of viable offspring. Our findings in the rat model uncover conserved principles underpinning pluripotency and germline competency across species.
Project description:We took advantage of a transgenic rat expressing the green fluorescent protein (GFP) specifically in germ cells allowing purification of perinatal GFP-positive germ cells. Timed-pregnant rats were exposed to ethinylestradiol (EE2, 2 µg/kg/d), genistein (GE, 10 mg/kg/d) or vehicle by gavage, from gestational days (GD) 13 to 19; testes were sampled at GD20 or post-natal (PND) 5 for sorting of GFP-positive cells. Gene expression was assessed in GFP-positive cells using Affymetrix Rat Gene 2.0 ST microarrays.
Project description:To test the impact of a maternal HFD on embryonic germline DNA methylation erasure, we use a rat strain that expresses green fluorescent protein specifically in germ cells. Germ cells were collected by FACS from male and female F1 gonads at midgestation for DNA methylation analysis by methyl-seq capture.