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8 October 2021 Effects of Calyculin a on the Motility and Protein Phosphorylation in Frozen-Thawed Bull Spermatozoa
Honami Ogata, Mariko Tsukamoto, Kenichi Yamashita, Tokuko Iwamori, Hideyuki Takahashi, Takane Kaneko, Naoki Iwamori, Tetsuichiro Inai, Hiroshi Iida
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Abstract

In this study, we examined the effects of calyculin A, a phosphatase inhibitor, on motility, protein phosphorylation, and the distribution of phospho-(Ser/Thr) PKA substrates in frozen-thawed bull spermatozoa that are actually used by most farmers for breeding. The data showed that calyculin A, which has been reported to have a positive effect on the motility of ejaculated fresh spermatozoa, distinctly decreased the motility of frozen-thawed bull spermatozoa even if a cell activator, such as caffeine, was present in the incubation medium and that the suppressive effect of calyculin A was dose-dependent and continued for at least 200 min. Immunoblot analyses revealed that de novo protein phosphorylation was not detected in spermatozoa exposed to caffeine or dbcAMP (a cell-permeable cAMP analog), while the addition of calyculin A to the medium brought about the appearance of several phosphorylated proteins at 50 kDa and 75 kDa, suggesting that 50 kDa and 75 kDa proteins, which were phosphorylated by activation of cAMP-dependent PKA, were not dephosphorylated and were accumulated in spermatozoa due to the suppression of calyculin A-sensitive protein phosphatases. Immunofluorescence microscopy revealed that calyculin A caused, alone or in conjunction with caffeine or dbcAMP, the accumulation of phospho-PKA substrates at the annulus, although caffeine or dbcAMP alone did not. This study suggested that calyculin A decreases the motility of frozen-thawed bull spermatozoa concomitant with the accumulation of phospho-(Ser/Thr) PKA substrates at the annulus of flagella.

© 2021 Zoological Society of Japan
Honami Ogata, Mariko Tsukamoto, Kenichi Yamashita, Tokuko Iwamori, Hideyuki Takahashi, Takane Kaneko, Naoki Iwamori, Tetsuichiro Inai, and Hiroshi Iida "Effects of Calyculin a on the Motility and Protein Phosphorylation in Frozen-Thawed Bull Spermatozoa," Zoological Science 38(6), 531-543, (8 October 2021). https://doi.org/10.2108/zs210046
Received: 10 May 2021; Accepted: 6 August 2021; Published: 8 October 2021
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