Cell surface beta-1,4-galactosyltransferase-I activates G protein-dependent exocytotic signaling.
ZP3 is a protein in the mammalian egg coat (zona pellucida) that binds sperm and stimulates acrosomal exocytosis, enabling sperm to penetrate the zona pellucida. The nature of the ZP3 receptor/s on sperm is a matter of considerable debate, but most evidence suggests that ZP3 binds to beta-1,4-galactosyltransferase-I (GalTase) on the sperm surface. It has been suggested that ZP3 induces the acrosome reaction by crosslinking GalTase, activating a heterotrimeric G protein. In this regard, acrosomal exocytosis is sensitive to pertussis toxin and the GalTase cytoplasmic domain can precipitate G(i) from sperm lysates. Sperm from mice that overexpress GalTase bind more soluble ZP3 and show accelerated G protein activation, whereas sperm from mice with a targeted deletion in GalTase have markedly less ability to bind soluble ZP3, undergo the ZP3-induced acrosome reaction, and penetrate the zona pellucida. We have examined the ability of GalTase to function as a ZP3 receptor and to activate heterotrimeric G proteins using Xenopus laevis oocytes as a heterologous expression system. Oocytes that express GalTase bound ZP3 but did not bind other zona pellucida glycoproteins. After oocyte maturation, ZP3 or GalTase antibodies were able to trigger cortical granule exocytosis and activation of GalTase-expressing eggs. Pertussis toxin inhibited GalTase-induced egg activation. Consistent with G protein activation, both ZP3 and anti-GalTase antibodies increased GTP-gamma[(35)S] binding as well as GTPase activity in membranes from eggs expressing GalTase. Finally, mutagenesis of a putative G protein activation motif within the GalTase cytoplasmic domain eliminated G protein activation in response to ZP3 or anti-GalTase antibodies. These results demonstrate directly that GalTase functions as a ZP3 receptor and following aggregation, is capable of activating pertussis toxin-sensitive G proteins leading to exocytosis.
Shi
X
X
Department of Animal Sciences, University of Illinois at Urbana-Champaign, Urbana, IL 61801, USA.
Amindari
S
S
Paruchuru
K
K
Skalla
D
D
Burkin
H
H
Shur
B D
BD
Miller
D J
DJ
D020816:Amino Acid Motifs; D000818:Animals; D002462:Cell Membrane / Q000201:enzymology; D004527:Egg Proteins / Q000378:metabolism*; D005089:Exocytosis / Q000502:physiology*; D005260:Female; D005700:Galactosyltransferases / Q000235:genetics / Q000378:metabolism*; D020962:Heterotrimeric GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D008297:Male; D008562:Membrane Glycoproteins / Q000378:metabolism*; D008845:Microinjections; D008856:Microscopy, Fluorescence; D009865:Oocytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D037342:Pertussis Toxin; D012313:RNA / Q000378:metabolism; D011956:Receptors, Cell Surface / Q000378:metabolism*; D015398:Signal Transduction; D013094:Spermatozoa / Q000502:physiology; D010566:Virulence Factors, Bordetella / Q000494:pharmacology; D014982:Xenopus laevis; D015044:Zona Pellucida / Q000737:chemistry / Q000378:metabolism*; D000072216:Zona Pellucida Glycoproteins
D004527:Egg Proteins; D008562:Membrane Glycoproteins; D011956:Receptors, Cell Surface; D010566:Virulence Factors, Bordetella; D000072216:Zona Pellucida Glycoproteins; C000606521:Zp3 protein, mouse; D012313:RNA; D005700:Galactosyltransferases; C119701:beta-1,4-galactosyltransferase I; D037342:Pertussis Toxin; D020962:Heterotrimeric GTP-Binding Proteins