In males, DDO-3::mCherry was localized in the excretory pore cell, head tip cells, hypodermal cells, seminal vesicle cells, tail cells, and spicule cells. By contrast, DDO-3::mCherry was also localized in the tail fan from the adult stage, suggesting that DDO-3 is secreted in a SP-dependent manner from the cells where it is produced and transferred to the tail fan. Interestingly, DDO-3::mCherry was localized in vesicle-like structures in the seminal vesicle, where spermatids are stored until ejaculation. Because the seminal vesicle is composed of 20 secretory cells, we presumed that DDO-3 was secreted from the seminal vesicle into the seminal fluid and transferred to the hermaphrodite during mating. To test this possibility, we examined the uteri of hermaphrodites just after mating with transgenic males carrying the
ddo-3::mCherry reporter gene. DDO-3::mCherry was dispersed throughout the uterus, suggesting that DDO-3 is a seminal fluid protein. Taken together, these results are consistent with the idea that DDO-3 is secreted from the seminal vesicle into the seminal fluid in a SP-dependent manner, and then transferred to the hermaphrodite uterus through mating. This fusion protein was also present in coelomocytes of males. During larval stages L1 and L2, localization of DDO-3::mCherry outside coelomocytes was limited to the excretory pore cell, implying that DDO-3 expressed in excretory pore cells is secreted into the body fluid. Indeed, at least in the adult stage, DDO-3::mCherry in excretory pore cells was localized to vesicle-like structures.