Role of tyrosine phosphorylation of flagellar proteins in hamster sperm hyperactivation

Despite extensive study of sperm motility, little is known of the mechanism of mammalian sperm hyperactivation. Here we describe a novel method for pr eparation of rodent sperm flagella and use it to show a correlation between tyrosine phosphorylation of flagellar proteins and hyperactivation of hams ter sperm. When hyperactivation was produced by a 3.5-h incubation in a med ium supporting capacitation, four major tyrosine-phosphorylated peptides of 90-, 80-, 62-,and 48-kDa mass were detected in flagellar extracts. Incubat ion with calyculin A, an inhibitor of protein phosphatases 1 and 2A, produc ed hyperactivation within 40 min but only a single 80-kDa phosphotyrosine-c ontaining flagellar component. Conversely, incubation with inhibitors of ei ther protein kinase A (H8) or protein tyrosine kinase (tyrphostin 47) preve nted both hyperactivation and the production of tyrosine-phosphorylated fla gellar peptides. These results indicate a strong correlation of hyperactiva tion with the tyrosine phosphorylation of sperm flagellar peptides, and the y strongly implicate an 80-kDa component as a major mediator of the mechani sm that produces hyperactivated motility of hamster sperm.