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The dynamics of MAPK inactivation at fertilization in mouse eggs.

ABSTRACT: Egg activation at fertilization in mammals is initiated by prolonged Ca(2+) oscillations that trigger the completion of meiosis and formation of pronuclei. A fall in mitogen-activated protein kinase (MAPK) activity is essential for pronuclear formation, but the precise timing and mechanism of decline are unknown. Here, we have measured the dynamics of MAPK pathway inactivation during fertilization of mouse eggs using novel chemiluminescent MAPK activity reporters. This reveals that the MAPK activity decrease begins during the Ca(2+) oscillations, but MAPK does not completely inactivate until after pronuclear formation. The MAPKs present in eggs are Mos, MAP2K1 and MAP2K2 (MEK1 and MEK2, respectively) and MAPK3 and MAPK1 (ERK1 and ERK2, respectively). Notably, the MAPK activity decline at fertilization is not explained by upstream destruction of Mos, because a decrease in the signal from a Mos-luciferase reporter is not associated with egg activation. Furthermore, Mos overexpression does not affect the timing of MAPK inactivation or pronuclear formation. However, the late decrease in MAPK could be rapidly reversed by the protein phosphatase inhibitor, okadaic acid. These data suggest that the completion of meiosis in mouse zygotes is driven by an increased phosphatase activity and not by a decline in Mos levels or MEK activity.

SUBMITTER: Gonzalez-Garcia JR

PROVIDER: S-EPMC4058113 | biostudies-literature | 2014 Jun

REPOSITORIES: biostudies-literature

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The dynamics of MAPK inactivation at fertilization in mouse eggs.

Gonzalez-Garcia Jose Raul JR Bradley Josephine J Nomikos Michail M Paul Laboni L Machaty Zoltan Z Lai F Anthony FA Swann Karl K

Journal of cell science 20140416 Pt 12

Egg activation at fertilization in mammals is initiated by prolonged Ca(2+) oscillations that trigger the completion of meiosis and formation of pronuclei. A fall in mitogen-activated protein kinase (MAPK) activity is essential for pronuclear formation, but the precise timing and mechanism of decline are unknown. Here, we have measured the dynamics of MAPK pathway inactivation during fertilization of mouse eggs using novel chemiluminescent MAPK activity reporters. This reveals that the MAPK acti ...[more]

PMID: 24741069

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Project description:BACKGROUND:Germinal Vesicle (GV) stage mouse oocytes in first meiotic prophase exhibit highly active HCO(3)(-)/Cl(-) exchange--a class of transport nearly ubiquitously involved in regulation of intracellular pH and cell volume. During meiosis, however, oocyte HCO(3)(-)/Cl(-) exchange becomes inactivated during first metaphase (MI), remains inactive in second metaphase (MII), and is reactivated only after egg activation. Previous work using pharmacological manipulations had indicated that activity of the MEK/MAPK signaling pathway was negatively correlated with HCO(3)(-)/Cl(-) exchange activity during meiosis. However, the mechanism by which the exchanger is inactivated during meiotic progression had not been determined, nor had the role of MEK/MAPK been directly established. METHODOLOGY/PRINCIPAL FINDINGS:Expression of a constitutively active form of MEK (MAP kinase kinase), which prevented the normal downregulation of MAPK after egg activation, also prevented reactivation of HCO(3)(-)/Cl(-) exchange. Conversely, suppression of endogenous MAPK activity with dominant negative MEK activated the normally quiescent HCO(3)(-)/Cl(-) exchange in mature MII eggs. A GFP-tagged form of the HCO(3)(-)/Cl(-) exchanger isoform Ae2 (Slc4a2) was strongly expressed at the GV oocyte plasma membrane, but membrane localization decreased markedly during meiotic progression. A similar pattern for endogenous Ae2 was confirmed by immunocytochemistry. The loss of membrane-localized Ae2 appeared selective, since membrane localization of a GFP-tagged human dopamine D1 receptor did not change during meiotic maturation. CONCLUSIONS:Direct manipulation of MAPK activity indicated that GFP-tagged Ae2 localization depended upon MAPK activity. Inactivation of HCO(3)(-)/Cl(-) exchange during the meiotic cell cycle may therefore reflect the loss of Ae2 from the oocyte plasma membrane, downstream of MEK/MAPK signaling. This identifies a novel role for MEK/MAPK-mediated cytostatic factor (CSF) activity during meiosis in membrane protein trafficking in mouse oocytes, and shows for the first time that selective retrieval of membrane proteins is a feature of meiosis in mammalian oocytes.

Dataset Information

The dynamics of MAPK inactivation at fertilization in mouse eggs.

Publications

The dynamics of MAPK inactivation at fertilization in mouse eggs.

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