Among C. elegans spe (sperm-defective) genes, those required for fertilization are categorized into
spe-9 class. By reverse genetic approach, we have recently identified
spe-45/oig-7, which encodes an immunoglobulin (Ig)-like transmembrane protein.
spe-45 likely belongs to
spe-9 class, since ~300 self-sperm are not consumed by fertilization in
spe-45(
tm3715) hermaphrodites. However, we don't have easy, direct methods like in vitro fertilization to evaluate how
spe-45(
tm3715) spermatozoa are defective during fertilization. In this study, we carried out the live imaging analysis of fertilization by
spe-45(
tm3715) spermatozoa and the determination of SPE-45 localization by using transgenic worms. First of all, we made
him-5(
e1490) and
him-5(
e1490);
spe-45(
tm3715) male strains that produce spermatids whose nuclei are brightened by mCherry. We also used
fem-1(
hc17) females where the oocyte plasma membranes are labeled with GFP. After outcrossing the female and male worms, movies of the spermatheca were captured during fertilization. As GFP-labeled oocytes were fertilized by
him-5(
e1490) spermatozoa, a part of the GFP signal was missing like a patch, where gamete fusion probably had occurred. Contrarily,
him-5(
e1490);
spe-45(
tm3715) spermatozoa barely produced the dark patch on the oocyte plasma membrane. Therefore,
spe-45(
tm3715) spermatozoa seemed incapable of fusing with oocytes. If SPE-45 directly functions in sperm-oocyte fusion, this protein likely resides on the sperm pseudopods. We thus constructed two transgenes, a gfp gene that is flanked by native 5'- and 3'-UTRs of
spe-45, and a modified
spe-45 gene that encodes SPE-45 protein where a FLAG tag is inserted into the cytoplasmic tail (SPE45-CTFLAG). As the gfp reporter gene was expressed in
him-5(
e1490) male, germ cells became GFP-positive in the proximal gonads. To figure out the subcellular localization of SPE-45, we integrated a single copy of the SPE45-CTFLAG transgene into the
spe-45(
tm3715) genome. Currently, immunofluorescence of male germ cells expressing the SPE45-CTFLAG transgene is ongoing by using anti-FLAG monoclonal antibody. Through this study, it was found that the male germline-specific gene
spe-45 is essentially required for gamete fusion. In mice, the sperm-specific Ig-like protein IZUMO1 acts during sperm-oocyte fusion. Hence, findings that are obtained from functional analysis of
spe-45 might provide clues to elucidate the sperm fusogenic machinery that is evolutionally conserved.