The dynamics of PKC-induced phosphorylation triggered by Ca2+oscillations in mouse eggs

Gonzalez Garcia, Jose, Machaty, Zoltan, Lai, Francis Anthony ORCID: https://orcid.org/0000-0003-2852-8547 and Swann, Karl ORCID: https://orcid.org/0000-0002-4355-1449 2013. The dynamics of PKC-induced phosphorylation triggered by Ca2+oscillations in mouse eggs. Journal of Cellular Physiology 228 (1) , pp. 110-119. 10.1002/jcp.24110

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Abstract

Fertilization of mammalian eggs is characterized by a series of Ca2+ oscillations triggered by a phospholipase C activity. These Ca2+ increases and the parallel generation of diacylglycerol (DAG) stimulate protein kinase C (PKC). However, the dynamics of PKC activity have not been directly measured in living eggs. Here, we have monitored the dynamics of PKC-induced phosphorylation in mouse eggs, alongside Ca2+ oscillations, using fluorescent C-kinase activity reporter (CKAR) probes. Ca2+ oscillations triggered either by sperm, phospholipase C zeta (PLCζ) or Sr2+ all caused repetitive increases in PKC-induced phosphorylation, as detected by CKAR in the cytoplasm or plasma membrane. The CKAR responses lasted for several minutes in both the cytoplasm and plasma membrane then returned to baseline values before subsequent Ca2+ transients. High frequency oscillations caused by PLCζ led to an integration of PKC-induced phosphorylation. The conventional PKC inhibitor, Gö6976, could inhibit CKAR increases in response to thapsigargin or ionomycin, but not the repetitive responses seen at fertilization. Repetitive increases in PKCδ activity were also detected during Ca2+ oscillations using an isoform-specific δCKAR. However, PKCδ may already be mostly active in unfertilized eggs, since phorbol esters were effective at stimulating δCKAR only after fertilization, and the PKCδ-specific inhibitor, rottlerin, decreased the CKAR signals in unfertilized eggs. These data show that PKC-induced phosphorylation outlasts each Ca2+ increase in mouse eggs but that signal integration only occurs at a non-physiological, high Ca2+ oscillation frequency. The results also suggest that Ca2+-induced DAG formation on intracellular membranes may stimulate PKC activity oscillations at fertilization.

Item Type:	Article
Date Type:	Publication
Status:	Published
Schools:	Medicine
Subjects:	R Medicine > R Medicine (General)
Publisher:	Wiley
ISSN:	0021-9541
Last Modified:	21 Oct 2022 10:48
URI:	https://orca.cardiff.ac.uk/id/eprint/41404

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