Identification of multiple male reproductive tract-specific proteins that regulate sperm migration through the oviduct in mice

Fujihara, Yoshitaka; Noda, Taichi; Kobayashi, Kiyonori; Oji, Asami; Kobayashi, Sumire; Matsumura, Takafumi; Larasati, Tamara; Oura, Seiya; Kojima‐Kita, Kanako; Yu, Zhifeng; Matzuk, Martin M.; Ikawa, Masahito

doi:10.1073/pnas.1908736116

Cited by 59 publications

(53 citation statements)

References 55 publications

(93 reference statements)

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“…Since Spint4/5 null male mice are severely subfertile, without an apparent difference in epididymis histology, sperm number, sperm morphology, and sperm motility parameters in comparison to the wild-type (WT) mice, this phenocopies the reproductive phenotype of several null mice of testis-, epididymis-, or prostate-specific genes (Sof1, Tmem95, and Spaca6; Pate8, and Pate10), which reveal a requirement in regulating sperm migration through the oviduct and sperm-oocyte fusion in mice [92,93]. A severe fertility defect associated with normal sperm number, morphology, and motility is also shared among mice lacking the sperm membrane protein ADAM3, thought to be crucial in sperm-ZP binding and sperm migration through the uterotubular junction (UTJ) [94,95].…”

Section: Drug Target Specificity Of Novel Targetsmentioning

confidence: 78%

Large-scale discovery of male reproductive tract-specific genes through analysis of RNA-seq datasets

et al. 2020

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Background: The development of a safe, effective, reversible, non-hormonal contraceptive method for men has been an ongoing effort for the past few decades. However, despite significant progress on elucidating the function of key proteins involved in reproduction, understanding male reproductive physiology is limited by incomplete information on the genes expressed in reproductive tissues, and no contraceptive targets have so far reached clinical trials. To advance product development, further identification of novel reproductive tract-specific genes leading to potentially druggable protein targets is imperative. Results: In this study, we expand on previous single tissue, single species studies by integrating analysis of publicly available human and mouse RNA-seq datasets whose initial published purpose was not focused on identifying male reproductive tract-specific targets. We also incorporate analysis of additional newly acquired human and mouse testis and epididymis samples to increase the number of targets identified. We detected a combined total of 1178 genes for which no previous evidence of male reproductive tract-specific expression was annotated, many of which are potentially druggable targets. Through RT-PCR, we confirmed the reproductive tract-specific expression of 51 novel orthologous human and mouse genes without a reported mouse model. Of these, we ablated four epididymis-specific genes (Spint3, Spint4, Spint5, and Ces5a) and two testis-specific genes (Pp2d1 and Saxo1) in individual or double knockout mice generated through the CRISPR/Cas9 system. Our results validate a functional requirement for Spint4/5 and Ces5a in male mouse fertility, while demonstrating that Spint3, Pp2d1, and Saxo1 are each individually dispensable for male mouse fertility. Conclusions: Our work provides a plethora of novel testis-and epididymis-specific genes and elucidates the functional requirement of several of these genes, which is essential towards understanding the etiology of male infertility and the development of male contraceptives.

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Section: Drug Target Specificity Of Novel Targetsmentioning

confidence: 78%

Large-scale discovery of male reproductive tract-specific genes through analysis of RNA-seq datasets

et al. 2020

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“…The CRISPR-KO approach efficiently screens male fertility genes in vivo. In fact, we revealed that more than 90 genes are dispensable (22)(23)(24)(25)(26), while 10 genes and two clusters are required (21,(27)(28)(29)(30) for male fertility. Recently, we reported that a testis-specific gene, 4930451I11Rik (renamed as Fertilization Influencing Membrane Protein [Fimp]), is another sperm factor critical for sperm−oocyte fusion (31).…”

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confidence: 91%

Sperm proteins SOF1, TMEM95, and SPACA6 are required for sperm−oocyte fusion in mice

Noda

Fujihara

et al. 2020

Proc. Natl. Acad. Sci. U.S.A.

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Sperm−oocyte membrane fusion is one of the most important events for fertilization. So far, IZUMO1 and Fertilization Influencing Membrane Protein (FIMP) on the sperm membrane and CD9 and JUNO (IZUMO1R/FOLR4) on the oocyte membrane have been identified as fusion-required proteins. However, the molecular mechanisms for sperm−oocyte fusion are still unclear. Here, we show that testis-enriched genes, sperm−oocyte fusion required 1 (Sof1/Llcfc1/1700034O15Rik), transmembrane protein 95 (Tmem95), and sperm acrosome associated 6 (Spaca6), encode sperm proteins required for sperm−oocyte fusion in mice. These knockout (KO) spermatozoa carry IZUMO1 but cannot fuse with the oocyte plasma membrane, leading to male sterility. Transgenic mice which expressed mouseSof1,Tmem95,andSpaca6rescued the sterility ofSof1,Tmem95, andSpaca6KO males, respectively. SOF1 and SPACA6 remain in acrosome-reacted spermatozoa, and SPACA6 translocates to the equatorial segment of these spermatozoa. The coexpression of SOF1, TMEM95, and SPACA6 in IZUMO1-expressing cultured cells did not enhance their ability to adhere to the oocyte membrane or allow them to fuse with oocytes. SOF1, TMEM95, and SPACA6 may function cooperatively with IZUMO1 and/or unknown fusogens in sperm−oocyte fusion.

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“…Due to the power of the CRISPR/Cas9 system, we have the ability to efficiently and accurately delete family cluster genes. While the Oosp family was not found to be critical for fertility, we have identified multiple male reproductive tract specific family cluster proteins that are [41]. The PATE and CST families are crucial for male fertility [41].…”

Section: Discussionmentioning

confidence: 75%

“…While the Oosp family was not found to be critical for fertility, we have identified multiple male reproductive tract specific family cluster proteins that are [41]. The PATE and CST families are crucial for male fertility [41]. Interestingly, a few of these genes were not essential individually but rendered the male mice to be infertile when knocked out together.…”

Section: Discussionmentioning

confidence: 86%

CRISPR/Cas9-Mediated Genome Editing Reveals Oosp Family Genes are Dispensable for Female Fertility in Mice

Abbasi

Kodani

Emori

et al. 2020

Cells

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There are over 200 genes that are predicted to be solely expressed in the oocyte and ovary, and thousands more that have expression patterns in the female reproductive tract. Unfortunately, many of their physiological functions, such as their roles in oogenesis or fertilization, have yet to be elucidated. Previous knockout (KO) mice studies have proven that many of the genes that were once thought to be essential for fertility are dispensable in vivo. Therefore, it is extremely important to confirm the roles of all genes before spending immense time studying them in vitro. To do this, our laboratory analyzes the functions of ovary and oocyte-enriched genes in vivo through generating CRISPR/Cas9 KO mice and examining their fertility. In this study, we have knocked out three Oosp family genes (Oosp1, Oosp2, and Oosp3) that have expression patterns linked to the female reproductive system and found that the triple KO (TKO) mutant mice generated exhibited decreased prolificacy but were not infertile; thus, these genes may potentially be dispensable for fertility. We also generated Cd160 and Egfl6 KO mice and found these genes are individually dispensable for female fertility. KO mice with no phenotypic data are seldom published, but we believe that this information must be shared to prevent unnecessary experimentation by other laboratories.

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Identification of multiple male reproductive tract-specific proteins that regulate sperm migration through the oviduct in mice

Cited by 59 publications

References 55 publications

Large-scale discovery of male reproductive tract-specific genes through analysis of RNA-seq datasets

Large-scale discovery of male reproductive tract-specific genes through analysis of RNA-seq datasets

Sperm proteins SOF1, TMEM95, and SPACA6 are required for sperm−oocyte fusion in mice

CRISPR/Cas9-Mediated Genome Editing Reveals Oosp Family Genes are Dispensable for Female Fertility in Mice

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