EIF5A couples translational control with transcriptional reprogramming through chromocenter reorganization during spermiogenesis [RNA-Seq]

Translation initiation factor eIF5A facilitates protein synthesis and impacts diverse biological processes, yet its role in transcriptional regulation is poorly understood. Here we find eIF5A highly expressed in diverse spermatogenic cell types. Conditional knockout of Eif5a (SKO) causes complete infertility in male mice due to round spermatid arrest. Interestingly, eIF5A deletion severely compromises chromocenter integrity in round spermatids. Proteomic profiling reveals widespread dysregulation in eIF5A-deficient round spermatids, downregulated proteins are enriched for chromatin-associated functions, likely contributing to chromocenter dysfunction. Notably, ATAC-seq analysis shows increased chromatin accessibility upon eIF5A depletion, accompanied by transcriptional dysregulation of genes critical for acrosome and manchette formation. Our data underscore that eIF5A not only regulates the translation of chromatin-organizing proteins required for chromocenter stability but also influences transcriptional regulation by modulating chromatin landscape. These findings illuminate a novel, germ cell-specific pathway coupling translational control and transcriptional regulation via chromatin reorganization. Overall design: We performed Smart-seq2 on FACS-isolated round spermatids from control and SKO mice at postnatal day 28 (primarily step 8 stage).For Smart-seq2, cell lysis, mRNA reverse transcription, and cDNA amplification were performed using the Discover-sc WTA Kit V2 (Vazyme, N711). Following this, cDNA fragmentation and adapter ligation were carried out with the TruePrep Flexible DNA Library Prep Kit for Illumina (Vazyme, TD504). All procedures were conducted in strict accordance with the manufacturer's protocols. The Smart-seq2 library sequencing was performed by Novogene on Illumina platforms, generating 150 bp paired-end reads.