title: Calcium/calmodulin and calmodulin kinase II stimulate hyperactivation in demembranated bovine sperm.
abstract: Hyperactivated motility is observed among sperm in the mammalian oviduct near the time of ovulation. It is characterized by high-amplitude, asymmetrical flagellar beating and assists sperm in penetrating the cumulus oophorus and zona pellucida. Elevated intracellular Ca2+ is required for the initiation of hyperactivated motility, suggesting that calmodulin (CALM) and Ca2+/CALM-stimulated pathways are involved. A demembranated sperm model was used to investigate the role of CALM in promoting hyperactivation. Ejaculated bovine sperm were demembranated and immobilized by brief exposure to Triton X-100. Motility was restored by addition of reactivation medium containing MgATP and Ca2+, and hyperactivation was observed as free Ca2+ was increased from 50 nM to 1 microM. However, when 2.5 mM Ca2+ was added to the demembranation medium to extract flagellar CALM, motility was not reactivated unless exogenous CALM was readded. The inclusion of anti-CALM IgG in the reactivation medium reduced the proportion hyperactivated in 1 microM Ca2+ to 5%. Neither control IgG, the CALM antagonist W-7, nor a peptide directed against the CALM-binding domain of myosin light chain kinase (MYLK2) inhibited hyperactivation. However, when sperm were reactivated in the presence of CALM kinase II (CAMK2) inhibiting peptides, hyperactivation was reduced by 75%. Furthermore, an inhibitor of CAMK2, KN-93, inhibited hyperactivation without impairing normal motility of intact sperm. CALM and CAMK2 were immunolocalized to the acrosomal region and flagellum. These results indicate that hyperactivation is stimulated by a Ca2+/CALM pathway involving CAMK2.
authors.affiliation: Department of Biomedical Sciences, Cornell University, Ithaca, New York 14853, USA.
authors.forename: George G
authors.initials: GG
authors.lastname: Ignotz
authors.forename: Susan S
authors.initials: SS
authors.lastname: Suarez
mesh_terms: D000818:Animals; D002118:Calcium / Q000502:physiology*; D020013:Calcium Signaling; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D002147:Calmodulin / Q000502:physiology*; D002417:Cattle; D002462:Cell Membrane / Q000502:physiology; D008297:Male; D009219:Myosin-Light-Chain Kinase / Q000037:antagonists & inhibitors; D013081:Sperm Motility; D013094:Spermatozoa / Q000502:physiology*
chemical_list: D002147:Calmodulin; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases; C080215:MYLK protein, Bos taurus; D009219:Myosin-Light-Chain Kinase; D002118:Calcium