The Protein Phosphorylation Landscape of Mouse Spermatids during Spermiogenesis

Abstract

The characteristic tadpole shape of sperm is formed from round spermatids via spermiogenesis, a process which results in dramatic morphological changes in the final stage of spermatogenesis in the testis. Protein phosphorylation, as one of the most important post-translational modifications, can regulate spermiogenesis; however, the phosphorylation events taking place during this process have not been systematically analyzed. In order to better understand the role of phosphorylation in spermiogenesis, large-scale phosphoproteome profiling is performed using IMAC and TiO 2 enrichment. In total, 13 835 phosphorylation sites, in 4196 phosphoproteins, are identified in purified mouse spermatids undergoing spermiogenesis in two biological replicates. Overall, 735 testis-specific proteins are identified to be phosphorylated, and are expressed at high levels during spermiogenesis. Gene ontology analysis shows enrichment of the identified phosphoproteins in terms of histone modification, cilium organization, centrosome and the adherens junction. Further characterization of the kinase-substrate phosphorylation network demonstrates enrichment of phosphorylation substrates related to the regulation of spermiogenesis. This global protein phosphorylation landscape of spermiogenesis shows wide phosphoregulation across a diverse range of processes during spermiogenesis and can help to further characterize the process of sperm generation. All MS data are available via ProteomeXchange with the identifier PXD011890.

Cyclin‐dependent kinase 7 is essential for spermatogenesis by regulating retinoic acid signaling pathways and the STAT3 molecular pathway

“…Spermatid differentiation is a complex process closely associated with phosphorylation that results in dramatic morphological and functional changes in spermatids. 44,45 Consistent with the localization characteristics of CDK7 (expressed in steps 1-11 of spermatids) (Figure 1b), our results suggested that the inhibition of CDK7 affects spermatid differentiation, resulting in the observed reduced in sperm count and quality. On the one hand, inhibition of CDK7 affected acrosome formation and nuclear condensation and structure, leading to sperm head malformation (Figure 5).…”

“…Phosphorylation is particularly active during spermiogenesis 44 . We previously constructed the phosphoproteome of mouse spermatids 45 . Based on bioinformatics analysis, we predicted the potential phosphorylated target proteins downstream of CDK7 (Figure 8a).…”

“…44 We previously constructed the phosphoproteome of mouse spermatids. 45 Based on bioinformatics analysis, we predicted the potential phosphorylated target proteins downstream of CDK7 (Figure 8a). Among these candidates, we selected EP300 and STAT3 to test their phosphorylation expression levels.…”

ZDHHC19 Is Dispensable for Spermatogenesis, but Is Essential for Sperm Functions in Mice

“…Multiple protein post-translational modifications (PTMs) occur in germ cells during spermatogenesis, such as acetylation [2,3], crotonylation [4], glycosylation [5][6][7], phosphorylation [8][9][10], SUMOylation [11], and ubiquitination [12]. Many of these PTMs have been identified to play important roles in sustaining mammalian spermatogenesis and male fertility [13,14].…”

“…Ohno et al investigated the ZDHHCs genes expression patterns in human by RT-PCR, and classified ZDHHC genes into roughly three groups according to their tissue-specific expression patterns: highly ubiquitous, nearly ubiquitous, and tissue-specific [27]. The highly ubiquitous and nearly ubiquitous groups each include most of ZDHHC genes (ZDHHC4, 5,7,8,10,12,13,17,and 22;and ZDHHC1,3,6,9,14,16,18, and 21, respectively), indicating that multiple ZDHHC proteins are co-expressed in the same cell. Interestingly, a small number of ZDHHC genes are tissue-specific, including ZDHHC2, 11, 15, 19, and 20.…”