The egg membrane microdomain-associated uroplakin III-Src system becomes functional during oocyte maturation and is required for bidirectional gamete signaling at fertilization in Xenopus laevis.
In Xenopus laevis, sperm-egg interaction promotes partial proteolysis and/or tyrosine phosphorylation of uroplakin III (UPIII) and the tyrosine kinase Src, which both localize to the cholesterol-enriched egg membrane microdomains (MDs). Here we show that sperm promote proteolysis and/or tyrosine phosphorylation of UPIII and Src in MDs isolated from ovulated and unfertilized eggs (UF-MDs). An antibody against the extracellular domain of UPIII interferes with these events. Inhibition of fertilization by anti-UPIII antibody is rescued by co-incubation with UF-MDs. This suggests that, like MDs in intact eggs, the isolated UF-MDs are capable of interacting with sperm, an interaction that does not interfere with normal fertilization but rather augments the ability of sperm to fertilize eggs pretreated with anti-UPIII antibody. This unexpected effect of UF-MDs on sperm requires UPIII function in UF-MDs and protein kinase activity in sperm. MDs isolated from progesterone-treated mature oocytes, but not ovarian immature oocytes, are similarly functional as UF-MDs. The anti-UPIII extracellular domain antibody binds more effectively to the surface of mature than immature ovarian oocytes. We propose that the structural and functional competency of the UPIII-Src signaling system in MDs is strictly regulated during oocyte maturation and subsequently in sperm-mediated egg activation and fertilization. The fertilization-related signaling properties seen in UF-MDs can be partially reconstituted in MDs of human embryonic kidney 293 cells (293-MDs) expressing UPIII, Src and uroplakin Ib. However, 293-MDs expressing a proteolysis-resistant mutant of UPIII are less functional, suggesting that the availability of UPIII to protease action is important for MD function.
Laboratory of Cell Signaling and Development, Department of Molecular Biosciences, Faculty of Life Sciences, Kyoto Sangyo University, Kyoto 603-8555, Japan.
A K M
AK
Mahbub Hasan
Aki
A
Hashimoto
Yuka
Y
Maekawa
Takashi
T
Matsumoto
Shota
S
Kushima
Takashi W
TW
Ijiri
Yasuo
Y
Fukami
Ken-ichi
K
Sato
D000818:Animals; D000906:Antibodies / Q000494:pharmacology; D002401:Cathepsin B / Q000378:metabolism; D002454:Cell Differentiation / Q000187:drug effects; D005260:Female; D005306:Fertilization / Q000187:drug effects; D057809:HEK293 Cells; D006801:Humans; D008297:Male; D021962:Membrane Microdomains / Q000187:drug effects / Q000378:metabolism*; D008954:Models, Biological; D009154:Mutation / Q000235:genetics; D009865:Oocytes / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D010063:Ovum / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D019000:Phosphotyrosine / Q000378:metabolism; D011374:Progesterone / Q000494:pharmacology; D015398:Signal Transduction / Q000187:drug effects; D013094:Spermatozoa / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D060171:Uroplakin III / Q000378:metabolism*; D060169:Uroplakin Ib / Q000378:metabolism; D014982:Xenopus laevis / Q000378:metabolism*; D019061:src-Family Kinases / Q000378:metabolism*
D000906:Antibodies; D060171:Uroplakin III; D060169:Uroplakin Ib; D019000:Phosphotyrosine; D011374:Progesterone; D019061:src-Family Kinases; D002401:Cathepsin B
Fertilization; Gamete interaction; In vitro reconstitution; Membrane microdomains; Oocyte maturation; Proteolysis; Signal transduction; Src; Tyrosine phosphorylation; Uroplakin III