Distinct segment-specific functions of calyculin A-sensitive protein phosphatases in the regulation of cAMP-triggered events in ejaculated bull spermatozoa.
Livestock spermatozoa possess more tenacious suppressors of cAMP-triggered events-including capacitation-associated changes-than laboratory animal spermatozoa, leading to flagellar hyperactivation. In order to identify the suppressors, we examined effects of an inhibitor of serine/threonine protein phosphatases (calyculin A) on cAMP-triggered changes in the protein phosphorylation state, and subsequent occurrence of hyperactivation and acrosome reaction in ejaculated bull spermatozoa. Ejaculated spermatozoa were incubated in cAMP-supplemented medium, then assessed for motility, acrosome morphology, and phosphorylated protein localization. The addition of calyculin A greatly enhanced cAMP-triggered protein phosphorylation at serine/threonine and tyrosine residues in the connecting piece and induction of flagellar hyperactivation. Most hyperactivated spermatozoa exhibited extremely asymmetrical bends at the middle piece, which produced intensive twisting or figure-eight movements. In the sperm head, however, cAMP-triggered dephosphorylation of serine/threonine-phosphorylated proteins and subsequent acrosome reaction were abolished by the addition of calyculin A. Based on these results, we suggest that calyculin A-sensitive protein phosphatases in the connecting piece are suppressors of cAMP-triggered events leading to hyperactivation. By contrast, similar protein phosphatases in the sperm head accelerate cAMP-triggered events leading to the acrosome reaction. These findings are consistent with the indication that calyculin A-sensitive protein phosphatases have distinct functions in the regulation of cAMP-triggered events in different regions of ejaculated bull spermatozoa.
Laboratory of Reproductive Biology, Division of Animal Science, Department of Bioresource Science, Graduate School of Agricultural Science, Kobe University, Kobe, 657-8501, Japan.
Yohei
Y
Mizuno
Ayane
A
Isono
Aya
A
Kojima
Miyuki M
MM
Arai
Taichi
T
Noda
Mitsuhiro
M
Sakase
Moriyuki
M
Fukushima
Hiroshi
H
Harayama
D020101:Acrosome Reaction / Q000187:drug effects*; D000818:Animals; D015153:Blotting, Western; D002417:Cattle; D002465:Cell Movement / Q000187:drug effects; D000242:Cyclic AMP / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D046149:Embryo Culture Techniques; D005407:Flagella / Q000187:drug effects / Q000502:physiology*; D019084:Fluorescent Antibody Technique, Indirect; D066298:In Vitro Techniques; D008297:Male; D008387:Marine Toxins; D010080:Oxazoles / Q000037:antagonists & inhibitors / Q000494:pharmacology*; D010749:Phosphoprotein Phosphatases / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D013094:Spermatozoa / Q000187:drug effects / Q000378:metabolism*
D008387:Marine Toxins; D010080:Oxazoles; C059041:calyculin A; D000242:Cyclic AMP; D010749:Phosphoprotein Phosphatases