Integrative Proteomic and Transcriptomic Analyses Reveal Multiple Post-transcriptional Regulatory Mechanisms of Mouse Spermatogenesis

Abstract

Mammalian spermatogenesis consists of many cell types and biological processes and serves as an excellent model for studying gene regulation at transcriptional and post-transcriptional levels. Many key proteins, miRNAs, and perhaps piRNAs have been shown to be involved in post-transcriptional regulation of spermatogenesis. However, a systematic method for assessing the relationship between protein and mRNA expression has not been available for studying mechanisms of post-transcriptional regulation. In the present study, we used the iTRAQbased quantitative proteomic approach to identify 2008 proteins in mouse type A spermatogonia, pachytene spermatocytes, round spermatids, and elongative spermatids with high confidence. Of these proteins, 1194 made up four dynamically changing clusters, which reflect the mitotic amplification, meiosis, and post-meiotic development of germ cells. We identified five major regulatory mechanisms termed "transcript only," "transcript degradation," "translation repression," "translation de-repression," and "protein degradation" based on changes in protein level relative to changes in mRNA level at the mitosis/meiosis transition and the meiosis/post-meiotic development transition. We found that post-transcriptional regulatory mechanisms are related to the generation of piRNAs and antisense transcripts. Our results provide a valuable inventory of proteins produced during mouse spermatogenesis and contribute to elucidating the mechanisms of the post-transcriptional regulation of gene expression in mammalian spermatogenesis. Molecular &

The E2 Ubiquitin-conjugating Enzyme UBE2J1 Is Required for Spermiogenesis in Mice

“…UBE2J1 protein was abundantly present in elongating spermatids, specifically from step 12 to 15 of sperm differentiation (31), peaking at step 14. Spermatids at these differentiation steps are largely transcriptionally inactive (32,33), implying that the transcripts are transcribed earlier, and that translation of the Ube2j1 (and presumably other) transcripts is de-repressed in elongate spermatids (34). Thus, UBE2J1 is likely required at a critical time within the elongating spermatid itself for successful completion of the final steps of differentiation and spermiation.…”

Linking transcriptomics and proteomics in spermatogenesis

“…Importantly, the correlation between transcriptome and proteome data was weakest for the testis and highest for the liver (correlation coefficients of 0.138 and 0.432 respectively). To investigate the relation between proteins and mRNA levels during spermatogenesis in more detail, Gan et al (2013a) used isolated type A spermatogonia, pachytene spermatocytes, round spermatids, and elongated spermatids for an iTRAQ-based proteomic analysis of male germ cell differentiation and compared their results with those of a previously published microarray dataset (Namekawa et al 2006). While they found a consistent match for a subset of transcriptomic and proteomic profiles, they also observed that several regulation mechanismsincluding transcript degradation, translation repression, translation de-repression, and protein degradationaffected most genes and may account for the low correlation between mRNAs and proteins, at both the mitosis/meiosis transition (Pearson correlation of 0.55) and the meiosis/post-meiosis transition (Pearson correlation of 0.41).…”

Expression dynamics, relationships, and transcriptional regulations of diverse transcripts in mouse spermatogenic cells

“…Taking advantage of this fact, various types of SPCs can be isolated at different timepoints after birth when they are first generated for genomic, transcriptomic and proteomic studies. [28][29][30] The primitive type A spermatogonia (priSG-A) are earliest stage spermatogonia which undergo mitosis; the preleptotene spermatocytes (plpSC) have completed meiotic DNA replication and are ready for subsequent meiotic events such as DNA recombination and synapsis. The pachytene spermatocytes (pacSC) are meiotic cells, in which synapses are fully established; the round spermatids (rST) are early stage haploids, and undergo a lengthy postmeiotic developmental process to finally generate mature sperm.…”