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

Taichi Noda,Yonggang Lu,Yoshitaka Fujihara,Seiya Oura,Takayuki Koyano,Sumire Kobayashi,Martin M Matzuk,Masahito Ikawa

doi:10.1073/pnas.1922650117

Abstract

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 mouse Sof1, Tmem95, and Spaca6 rescued the sterility of Sof1, Tmem95, and Spaca6 KO 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.

Highlights

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
In this study, using CRISPR-Cas9−mediated gene knockouts in mice, we discover that sperm proteins SOF1, TMEM95, and SPACA6 are required for sperm−oocyte fusion and male fertility
We focused on three testis-enriched genes, which are predicted as membrane proteins by in silico analyses

Summary

Introduction

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. 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. 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]. For the sperm−oocyte fusion step, only four factors have been identified: CD9 and JUNO expressed in oocytes and IZUMO1 and FIMP expressed in spermatozoa

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Conclusion