Project Summary/Abstract The long-term objective of the proposed research is to further the understanding of mammalian spermatogenesis at the molecular level. Results from these studies address issues of both human infertility and contraception. The proposed studies utilize an innovative technique to synchronize spermatogenesis and isolate subtypes of spermatogonia undergoing development, expanding in numbers and acquiring the competence to enter meiosis. Isolation of enough of these cells to carry out molecular studies is possible only because of this innovative technique. Specific spermatogonial subtypes from wild-type and STRA8 knockout mice will be isolated before and after a retinoic acid pulse. The key focus of the proposed studies will be to determine retinoic acid responsive transcripts and then relate the genes for these transcripts to chromatin structure, histone acetylation and enhancer formation. The retinoic acid responsive transcripts will be determined by RNAseq and the chromatin structure will utilize ATAC chromatin accessibility techniques. Retinoic acid receptors and H3K27ac modifications will be determined by CHIP-seq. We will determine the presence of super enhancer sites marked by H3K27ac and examine the epigenetic changes and transcription factor binding sites in these sites. These studies will provide evidence for the molecular changes required for the entry into meiosis and lay the ground-work for future studies relating to the molecular causes of infertility.