Mechanism of PP2A affecting ubiquitination pathway in spermatogenesis

The heterotrimeric complex serine-threonine protein phosphatase 2 A (PP2A) is integral to the regulation of essential cellular processes. It is particularly crucial in spermatogenesis, where it is indispensable for meiosis, mitosis, sperm capacitation, and apoptosis. Previous research has concentrated on the knockdown of the catalytic subunit of PP2A, PPP2CA, in germ cells using Ddx4-Cre, resulting in male mouse sterility, disrupted meiotic recombination, and meiotic arrest of spermatocytes. To further elucidate the role of PP2A in spermatogenesis, we performed transcriptomic and proteomic sequencing analyses on the testes of knockout and control mice. A nine-quadrant map was developed to depict the differential expression of genes and proteins. Our analyses identified 1732 differentially expressed genes, which exhibited a strong positive correlation with the trends in differential protein expression. Gene Ontology (GO) enrichment analysis indicated a significant downregulation of genes involved in spermatogenesis, sperm cell development, and sperm cell differentiation. Furthermore, KEGG enrichment analysis revealed a notable enrichment of differentially expressed genes within the ubiquitin-mediated proteolysis pathway. In knockout mouse testicular tissue, testicular expression of the ubiquitin-related gene, UBE2K, was markedly downregulated, which was associated with the accumulation of histone H3, upregulation of the methyltransferase SETDB1, and increased levels of H3K9me3. Similarly, knockdown of Ppp2ca in GC2 cells resulted in decreased UBE2K expression, histone H3 accumulation, SETDB1 upregulation, and elevated H3K9me3 levels, consistent with mirroring the phenotype observed in the knockout mice. Notably, the ubiquitin-related gene UBE2K was identified as a significant outlier in the nine-quadrant map, and real-time quantitative PCR confirmed that UBE2K transcript levels were significantly reduced in knockout mice compared to wild-type controls. These findings suggest that PP2A may regulate histon.