Regulation of protein tyrosine phosphorylation during bovine sperm capacitation by a cyclic adenosine 3'5'-monophosphate-dependent pathway.
Mammalian sperm capacitation, defined as an obligatory maturational process leading to the development of the fertilization-competent state, results from a poorly understood series of morphological and molecular events. We report here that ejaculated bovine sperm, incubated under conditions that support capacitation in vitro, display a reproducible pattern of protein tyrosine phosphorylations that are regulated by a cAMP-dependent pathway. The appearance of these tyrosine phosphorylated proteins correlated temporally with the time course of capacitation induced by heparin, and these phosphorylations displayed a similar heparin concentration dependence. Glucose, which inhibits capacitation, inhibited these protein tyrosine phosphorylations in media containing heparin. The biologically active cAMP analogues (dibutyryl cAMP [db-cAMP], 8-bromo cAMP, sp-cAMPS) and the phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine (IBMX) induced the same protein tyrosine phosphorylation patterns as seen with heparin. Moreover, these cAMP agonists could overcome the inhibition of the heparin-induced tyrosine phosphorylations by glucose. In contrast, Rp-adenosine-3',5'-cyclic monophosphorothioate (Rp-cAMPS), a protein kinase A (PK-A) antagonist, blocked the capacitation-associated increases in protein tyrosine phosphorylation. This cAMP regulation of the protein tyrosine phosphorylation pattern is mediated by PK-A since N-[2-(p-bromocinnamylamino) ethyl]-5-isoquinolinesulfonamide-dihydrochloride (H89), another inhibitor of PK-A, inhibited the heparin-induced protein tyrosine phosphorylation pattern in a concentration-dependent manner in either the absence or presence of db-cAMP, IBMX, and glucose. These data support a model for sperm capacitation that includes protein tyrosine phosphorylation as an important regulatory pathway, and a role for cAMP/PK-A in the regulation of this pathway leading to capacitation. These studies are the first to report a unique interrelationship between tyrosine kinase/phosphatase and cAMP signaling pathways at the level of PK-A in bovine sperm capacitation.
Galantino-Homer
H L
HL
Center for Research on Reproduction and Women's Health, University of Pennsylvania School of Medicine, Philadelphia 19104-6080, USA.
Visconti
P E
PE
Kopf
G S
GS
D015056:1-Methyl-3-isobutylxanthine / Q000494:pharmacology; D000177:Acrosome / Q000502:physiology; D000818:Animals; D015153:Blotting, Western; D002417:Cattle; D003470:Culture Media; D000242:Cyclic AMP / Q000502:physiology*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors; D004591:Electrophoresis, Polyacrylamide Gel; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005947:Glucose / Q000494:pharmacology; D006493:Heparin / Q000494:pharmacology; D066298:In Vitro Techniques; D007546:Isoquinolines / Q000494:pharmacology; D008297:Male; D010726:Phosphodiesterase Inhibitors / Q000494:pharmacology; D010766:Phosphorylation; D017027:Protein Tyrosine Phosphatases / Q000378:metabolism*; D013075:Sperm Capacitation / Q000502:physiology*; D013449:Sulfonamides
D003470:Culture Media; D004791:Enzyme Inhibitors; D007546:Isoquinolines; D010726:Phosphodiesterase Inhibitors; D013449:Sulfonamides; D006493:Heparin; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D017027:Protein Tyrosine Phosphatases; D005947:Glucose; C063509:N-(2-(4-bromocinnamylamino)ethyl)-5-isoquinolinesulfonamide; D015056:1-Methyl-3-isobutylxanthine