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Regulation of tyrosine kinase activity during capacitation in goat sperm

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Abstract

Protein tyrosine phosphorylation is a key event accompanying sperm capacitation. Although this signaling cascade generates an array of tyrosine-phosphorylated polypeptides, their molecular characterization is still limited. It is necessary to differentiate the localization of the tyrosine-phosphorylated proteins in spermatozoa to understand the link between the different phosphorylated proteins and the corresponding regulated sperm function. cAMP plays a pivotal role in the regulation of tyrosine phosphorylation. The intracellular cAMP levels were raised in goat spermatozoa by the addition of the phosphodiesterase inhibitor, IBMX in conjugation with caffeine. Tyrosine phosphorylation was significantly up-regulated following treatment with these two reagents. Treatment of caudal spermatozoa with IBMX and caffeine, time dependent up-regulated phosphorylation of the protein of molecular weights 50 and 200 kDa was observed. Increased phosphorylation was observed with a combination of IBMX and caffeine treatment. Tyrosine phosphorylation in caput spermatozoa was not affected significantly under these conditions. The expression level of tyrosine kinase in sperm was examined with specific inhibitors and with anti-phosphotyrosine antibody. The indirect immunofluorescence staining was carried out on ethanol permeabilized sperm using anti-phosphotyrosine antibody. Western blot analysis was done using two separate PKA antibodies: anti-PKA catalytic and anti-PKA RIα. Almost no difference was found in the intracellular presence of the PKA RIα and RIIα subunits in caput and caudal epididymal spermatozoa. However, the catalytic subunit seemed to be present in higher amount in caudal spermatozoa. The results show that caprine sperm displays an enhancement of phosphorylation in the tyrosine residues of specific proteins under in vitro capacitation conditions.

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Acknowledgments

This study was supported by the financial assistance from Council of Scientific and Industrial Research (37/1088/01/EMR) and from Bose Institute. Authors are thankful to Arindam Basu for technical assistance.

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  1. Division of Molecular Medicine, Bose Institute, P-1/12, C.I.T. Scheme, VII-M, Kolakata, 700054, India

    Madhumouli Chatterjee, Pinki Nandi, Swatilekha Ghosh & Parimal C. Sen

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  1. Madhumouli Chatterjee
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  2. Pinki Nandi
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  4. Parimal C. Sen
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Correspondence to Parimal C. Sen.

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Chatterjee, M., Nandi, P., Ghosh, S. et al. Regulation of tyrosine kinase activity during capacitation in goat sperm. Mol Cell Biochem 336, 39–48 (2010). https://doi.org/10.1007/s11010-009-0261-8

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  • DOI: https://doi.org/10.1007/s11010-009-0261-8

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