Testis-specific protein HSF5 is essential for proper chromatin organization and transcriptional reprogramming to drive pachynema progression.

Chromatin remodeling and transcriptional reprogramming play critical roles during mammalian meiotic prophase I; however, the precise mechanisms regulating these processes remain poorly understood. Our previous work demonstrated that deletion of heat shock factor 5 (HSF5), a member of the heat shock factor family, induces meiotic arrest and male infertility. However, the molecular pathways through which HSF5 governs meiotic progression have not yet been fully elucidated. In this study, a comprehensive multi-omics approach was applied to investigate the role of HSF5 in modulating chromatin dynamics and transcriptional reprogramming during pachynema progression. Analysis of ATAC-seq and single-cell RNA sequencing data revealed significant alterations in chromatin accessibility and disruption of the transcriptional regulatory network (TRN) in Hsf5 -/- spermatocytes. Additionally, HSF5 deficiency resulted in defective XY body formation and altered histone modifications. Notably, Hsf5 -/- spermatocytes also exhibited abnormal spermatoproteasome activity specifically on sex chromosomes, with evidence indicating that HSF5 may form a complex with USP7 in vivo to suppress H2AK119ub on meiotic sex chromosomes. These findings provide new insights into the complex, multifunctional role of HSF5 in regulating key meiotic events and advancing our understanding of its function during pachynema progression.