Myristoylated alanine-rich C kinase substrate, but not Ca2+/calmodulin-dependent protein kinase II, is the mediator in cortical granules exocytosis

Abstract

Sperm-egg fusion induces cortical granules exocytosis (CGE), a process that ensures the block to polyspermy. CGE can be induced independently by either a rise in intracellular calcium concentration or protein kinase C (PKC) activation. We have previously shown that myristoylated alanine-rich C kinase substrate (MARCKS) cross-links filamentous actin (F-actin) and regulates its reorganization. This activity is reduced either by PKC-induced MARCKS phosphorylation (PKC pathway) or by its direct binding to calmodulin (CaM; CaM pathway), both inducing MARCKS translocation, F-actin reorganization, and CGE. Currently, we examine the involvement of Ca 2C /CaM-dependent protein kinase II (CaMKII) and MARCKS in promoting CGE and show that PKC pathway can compensate for lack of Ca 2C /CaM pathway. Microinjecting eggs with either overexpressed protein or complementary RNA of constitutively active aCaMKII triggered resumption of second meiotic division, but induced CGE of an insignificant magnitude compared with CGE induced by wt aCaMKII. Microinjecting eggs with mutant-unphosphorylatable MARCKS reduced the intensity of 12-O-tetradecanoylphorbol 13-acetate or ionomycin-induced CGE by 50%, indicating that phosphorylation of MARCKS by novel and/or conventional PKCs (n/cPKCs) is a pivotal event associated with CGE. Moreover, we were able to demonstrate cPKCs involvement in ionomycin-induced MARCKS translocation and CGE. These results led us to propose that MARCKS, rather than CaMKII, as a key mediator of CGE. Reproduction (2008) 135 613-624

The γ isoform of CaM kinase II controls mouse egg activation by regulating cell cycle resumption

“…Nevertheless, expressing an artificially truncated, constitutively active form of CaMKIIα in mouse eggs results in ab- normal CG exocytosis, even though other EEA are normal (28). Both myosin light-chain kinase and protein kinase C have been implicated in CG exocytosis (30,31). In these studies, inhibition of CG release appeared incomplete, raising the possibility that more than one effector is involved in CG exocytosis.…”

Endoplasmic reticulum distribution during bovine oocyte activation is regulated by protein kinase C via actin filaments

“…However, it has been shown that PKC induces F‐actin depolymerization, allowing cortical granules (CG) exocytosis in rodent's oocyte (Eliyahu et al, 2005; Tsaadon, Kaplan‐Kraicer, & Shalgi, 2008). PKC regulates actin function by phosphorylating several actin regulatory proteins such as MARCKS, Adducin, Fascin and ERM proteins (Larsson, 2006).…”

“…F‐actin is dynamically assembled into higher‐order structures at specified locations to regulate diverse functions and PKC actively participates in this process. F‐actin depolymerization by PKC results in CG exocytosis in rat oocytes through PKC activity on MARCKS substrate (Eliyahu et al, 2005; Tsaadon et al, 2008). In this way, it is reasonable to infer that whereas PKC regulates CG exocytosis by F‐actin depolymerization through its action on MARCKS, PKC could regulate ER organization by F‐actin polymerization through another actin regulatory protein.…”

PKC Regulation of Gametogenesis and Early Development

“…MARCKS proteins are substrates for PKC, and MARCKS have the ability to crosslink actin filaments into a network at the cell cortex (Eliyahu et al, 2005(Eliyahu et al, , 2006. However, once MARCKS are phosphorylated by PKC the actin network can then disassemble (Tsaadon et al, 2008). Disassembly of the actin network at the egg cortex could release cortical granules from the cortical actin network and allow them to reach the plasma membrane to exocytose (Tsaadon et al, 2008).…”

“…However, once MARCKS are phosphorylated by PKC the actin network can then disassemble (Tsaadon et al, 2008). Disassembly of the actin network at the egg cortex could release cortical granules from the cortical actin network and allow them to reach the plasma membrane to exocytose (Tsaadon et al, 2008). Once cPKCs are activated, as part of their down regulation, the kinase is cleaved between its membrane-binding domain and catalytic subunit to form PKM (Jaken 1990;Hashimoto et al, 1991).…”