Evidence for the involvement of PECAM-1 in a receptor mediated signal-transduction pathway regulating capacitation-associated tyrosine phosphorylation in human spermatozoa.
Mammalian spermatozoa must become ;capacitated' in the female reproductive tract before they gain the ability to fertilize the oocyte. The attainment of a capacitated state has been correlated with a number of biochemical changes, the most notable of which is a dramatic increase in the tyrosine phosphorylation status of these cells. Despite its biological importance, the mechanisms responsible for initiating this tyrosine phosphorylation cascade in vivo are unknown. Here, we report that this signalling pathway can be elicited in a rapid, dose-dependent and lectin-specific manner by wheat germ agglutinin (WGA), but none of 18 other lectins assessed. This response was abrogated by prior enzymatic cleavage of either sialic acid or GlcNAc residues from the sperm surface and by treatment with a range of pharmacological inhibitors directed against protein kinase A, protein tyrosine kinases and intermediates including Src. Proteomic analysis of the WGA-binding sites on the sperm surface identified the putative cognate receptor as platelet cell adhesion molecule 1 (PECAM-1/CD31). This conclusion was supported by the following evidence: (i) anti-PECAM-1 antibodies identified a molecule of the correct molecular mass in human spermatozoa, (ii) PECAM-1 could be isolated from a pool of sperm surface proteins using WGA immobilized on a solid phase support, (iii) PECAM-1 and WGA co-localized to the sperm surface and (iv) anti-PECAM-1 antibodies could completely block the ability of WGA to stimulate tyrosine phosphorylation in these cells. Collectively, these data provide the first evidence that a receptor-mediated signal transduction pathway triggers human sperm capacitation and identifies PECAM-1 as the probable initiator of this second messenger cascade.
Nixon
Brett
B
Reproductive Science Group, University of Newcastle, Callaghan, NSW 2308, Australia.
Paul
Jonathan W
JW
Spiller
Cassy M
CM
Attwell-Heap
Abigail G
AG
Ashman
Leonie K
LK
Aitken
R John
RJ
D000595:Amino Acid Sequence; D000242:Cyclic AMP / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D037102:Lectins / Q000145:classification / Q000378:metabolism; D008297:Male; D008969:Molecular Sequence Data; D010766:Phosphorylation / Q000187:drug effects; D019000:Phosphotyrosine / Q000378:metabolism*; D019408:Platelet Endothelial Cell Adhesion Molecule-1 / Q000378:metabolism*; D011485:Protein Binding; D017382:Reactive Oxygen Species / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology*; D013075:Sperm Capacitation / Q000502:physiology*; D013094:Spermatozoa / Q000166:cytology / Q000378:metabolism*; D014909:Wheat Germ Agglutinins / Q000494:pharmacology; D019061:src-Family Kinases / Q000378:metabolism
D004791:Enzyme Inhibitors; D037102:Lectins; D019408:Platelet Endothelial Cell Adhesion Molecule-1; D017382:Reactive Oxygen Species; D014909:Wheat Germ Agglutinins; D019000:Phosphotyrosine; D000242:Cyclic AMP; D019061:src-Family Kinases; D017868:Cyclic AMP-Dependent Protein Kinases