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Role of phospholipase C-zeta domains in Ca2+-dependent phosphatidylinositol 4,5-bisphosphate hydrolysis and cytoplasmic Ca2+ oscillations

Nomikos, Michail, Blayney, Lynda Mary, Larman, Mark Graham, Campbell, Karen, Rossbach, Andreas, Saunders, Christopher Matthew, Swann, Karl and Lai, Francis Anthony 2005. Role of phospholipase C-zeta domains in Ca2+-dependent phosphatidylinositol 4,5-bisphosphate hydrolysis and cytoplasmic Ca2+ oscillations. Journal of biological chemistry 280 (35) , pp. 31011-31018. 10.1074/jbc.M500629200

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The sperm-specific phospholipase C- (PLC) elicits fertilization-like Ca2+ oscillations and activation of embryo development when microinjected into mammalian eggs (Saunders, C. M., Larman, M. G., Parrington, J., Cox, L. J., Royse, J., Blayney, L. M., Swann, K., and Lai, F. A. (2002) Development (Camb.) 129, 3533-3544; Cox, L. J., Larman, M. G., Saunders, C. M., Hashimoto, K., Swann, K., and Lai, F. A. (2002) Reproduction 124, 611-623). PLC may represent the physiological stimulus for egg activation and development at mammalian fertilization. PLC is the smallest known mammalian PLC isozyme, comprising two EF hand domains, a C2 domain, and the catalytic X and Y core domains. To gain insight into PLC structure-function, we assessed the ability of PLC and a series of domain-deletion constructs to cause phosphatidylinositol 4,5-bisphosphate hydrolysis in vitro and also to generate cytoplasmic Ca2+ changes in intact mouse eggs. PLC and the closely related PLC1 had similar Km values for phosphatidylinositol 4,5-bisphosphate, but PLC was around 100 times more sensitive to Ca2+ than was PLC1. Notably, specific phosphatidylinositol 4,5-bisphosphate hydrolysis activity was retained in PLC constructs that had either EF hand domains or the C2 domain removed, or both. In contrast, Ca2+ sensitivity was greatly reduced when either one, or both, of the EF hand domains were absent, and the Hill coefficient was reduced upon deletion of the C2 domain. Microinjection into intact mouse eggs revealed that all domain-deletion constructs were ineffective at initiating Ca2+ oscillations. These data suggest that the exquisite Ca2+-dependent features of PLC regulation are essential for it to generate inositol 1,4,5-trisphosphate and Ca2+ oscillations in intact mouse eggs.

Item Type: Article
Date Type: Publication
Status: Published
Schools: Medicine
ISSN: 1083-351X
Last Modified: 27 Mar 2020 02:38

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