3D genome-based transcriptional program for male germline differentiation

Project Summary Spermatogenesis produces fertilization-competent sperm that ensure the continuity of life. The 3D chromatin structure in male germ cells changes dynamically during spermatogenesis. However, the relationship between the 3D genome and transcription in the male germline remains largely unknown. To elucidate how the 3D genome regulates the transcriptional program during mouse spermatogenesis, I performed high-resolution Hi-C in the male germline. I found that CTCF-mediated 3D genome formation in undifferentiated spermatogonia serves as an epigenetic memory, initially establishing a gene expression program that is subsequently used to direct gene expression during late spermatogenesis. My central hypothesis is that the 3D genome predetermines a transcriptional program that directs subsequent spermatogenic differentiation. Elucidating the molecular mechanism by which this 3D genome is formed will provide insight into how the transcriptional program is established to ensure unidirectional spermatogenic differentiation. Aim 1 will determine whether the 3D genome is established in prospermatogonia by capturing Hi-C and CTCF profiles from PGCs to undifferentiated spermatogonia. Aim 2 will elucidate molecular mechanisms underlying formation of the 3D genome by determining how specific CTCF-binding sites are selected from PGCs to undifferentiated spermatogonia. Completion of these aims will elucidate how the transcriptional program for male germline differentiation is established from the perspective of the 3D genome during the period from PGCs after sex determination to conversion to spermatogonia, thereby providing mechanistic insight into how male germ cell identity is established. This research will contribute to the treatment of male infertility and the improvement of human reproductive success in the future. So far in my postdoctoral career in the Namekawa lab at UC Davis I have received training in experimental and analytical skills for comprehensive epigenomic studies, which is the basis for this research proposal. In the K99 phase, I will receive training in analysis techniques specific to 3D genomes and develop and refine my future directions. With the training plan and support of the K99/R00 award, this project will provide me with a strong foundation for success as an independent investigator pursuing important biological questions in germ cells.