Expression of uncharacterized male germ cell-specific genes and discovery of novel sperm-tail proteins in mice

Abstract

The identification and characterization of germ cell-specific genes are essential if we hope to comprehensively understand the mechanisms of spermatogenesis and fertilization. Here, we searched the mouse UniGene databases and identified 13 novel genes as being putatively testis-specific or -predominant. Our in silico and in vitro analyses revealed that the expressions of these genes are testis- and germ cell-specific, and that they are regulated in a stage-specific manner during spermatogenesis. We generated antibodies against the proteins encoded by seven of the genes to facilitate their characterization in male germ cells. Immunoblotting and immunofluorescence analyses revealed that one of these proteins was expressed only in testicular germ cells, three were expressed in both testicular germ cells and testicular sperm, and the remaining three were expressed in sperm of the testicular stages and in mature sperm from the epididymis. Further analysis of the latter three proteins showed that they were all associated with cytoskeletal structures in the sperm flagellum. Among them, MORN5, which is predicted to contain three MORN motifs, is conserved between mouse and human sperm. In conclusion, we herein identify 13 authentic genes with male germ cell-specific expression, and provide comprehensive information about these genes and their encoded products. Our finding will facilitate future investigations into the functional roles of these novel genes in spermatogenesis and sperm functions.

Mouse TMCO5 is localized to the manchette microtubules involved in vesicle transfer in the elongating spermatids

“…Conversely, because monoclonal antibodies recognize restricted antigenic determinants, it is possible that the monoclonal antibody (RTm01) recognizes the three-dimensional structure of TMCO5 protein so that the antibody could not recognize the protein in the round and differentiated spermatids. However, immunoblotting analysis in Fig 2B as well as that of Kwon et al [17] shows that TMCO5 expression starts from 4 weeks-old and 28-postnatal days, respectively, which indicates that TMCO5 expression starts from elongating spermatids in mice [17, 28, 36]. Taken together, the difference in the expression may be due to the difference in mouse and rat spermatogenesis.…”

“…Gene Database (Gene ID: 67356, https://www.ncbi.nlm.nih.gov/gene/) confirmed that the expression of the mRNA is restricted in the adult testes and that the deduced amino acid sequence has a coiled-coil domain in the N-terminal region and a transmembrane domain in the C-terminal region with the calculated molecular weight of the protein 35,850. Regarding the expression of the gene, Kwon et al reported the expression of thirteen testis-specific genes including Tmco5 by reverse transcriptase, PCR, and immunoblotting [17]. The function of this gene has not yet been reported.…”

Spermatogenesis is normal in Tex33 knockout mice

“…Tex33 is an evolutionary conserved and testis-specific gene. It is transcribed in round spermatids and it does not exist in testicular elongated spermatids and epididymal sperm (Kwon et al, 2017). In this study, we generated a 62bp in frame deletion on Exon2 of Tex33 in C57B/L6 mouse model by using the CRISPR/Cas9 system.…”

“…Tex33 is a recently discovered evolutionarily conserved gene present in vertebrates, which is initially expressed in the cytoplasm of round spermatids, and is diminished in elongated spermatids (Kwon et al, 2017). Until now, Tex33 knockout animal model has not been reported.…”

The SpliZ generalizes “Percent Spliced In” to reveal regulated splicing at single-cell resolution

“…The five genes with highest magnitude correlation between pseudotime and the SZS are TPPP2, a gene regulating tubulin polymerization implicated in male infertility 30 , FAM71E1, identified in mouse as testes-predominantly expressed 31 , SPATA42 a long noncoding RNA implicated in azoospermia 32 , MTFR1, a gene regulating mitochondrial fission and MLF1, an oncogene known to be regulated in drosophila testes 33 (Fig. 3A).…”