Kinase-dependent Regulation of Inositol 1,4,5-Trisphosphate-dependent Ca2+ Release during Oocyte Maturation

Abstract

Fertilization induces a species-specific Ca2+ transient with specialized spatial and temporal dynamics, which are essential to temporally encode egg activation events such as the block to polyspermy and resumption of meiosis. Eggs acquire the competence to produce the fertilization-specific Ca2+ transient during oocyte maturation, which encompasses dramatic potentiation of inositol 1,4,5-trisphosphate (IP3)-dependent Ca2+ release. Here we show that increased IP3 receptor (IP3R) sensitivity is initiated at the germinal vesicle breakdown stage of maturation, which correlates with maturation promoting factor (MPF) activation. Extensive phosphopeptide mapping of the IP3R resulted in ∼70% coverage and identified three residues, Thr-931, Thr-1136, and Ser-114, which are specifically phos pho ryl a ted during maturation. Phospho-specific antibody analyses show that Thr-1136 phos pho ryl a tion requires MPF activation. Activation of either MPF or the mitogen-activated protein kinase cascade independently, functionally sensitizes IP3-dependent Ca2+ release. Collectively, these data argue that the kinase cascades driving meiotic maturation potentiates IP3-dependent Ca2+ release, possibly trough direct phos pho ryl a tion of the IP3R. Fertilization induces a species-specific Ca2+ transient with specialized spatial and temporal dynamics, which are essential to temporally encode egg activation events such as the block to polyspermy and resumption of meiosis. Eggs acquire the competence to produce the fertilization-specific Ca2+ transient during oocyte maturation, which encompasses dramatic potentiation of inositol 1,4,5-trisphosphate (IP3)-dependent Ca2+ release. Here we show that increased IP3 receptor (IP3R) sensitivity is initiated at the germinal vesicle breakdown stage of maturation, which correlates with maturation promoting factor (MPF) activation. Extensive phosphopeptide mapping of the IP3R resulted in ∼70% coverage and identified three residues, Thr-931, Thr-1136, and Ser-114, which are specifically phos pho ryl a ted during maturation. Phospho-specific antibody analyses show that Thr-1136 phos pho ryl a tion requires MPF activation. Activation of either MPF or the mitogen-activated protein kinase cascade independently, functionally sensitizes IP3-dependent Ca2+ release. Collectively, these data argue that the kinase cascades driving meiotic maturation potentiates IP3-dependent Ca2+ release, possibly trough direct phos pho ryl a tion of the IP3R. Egg activation refers to the cellular and molecular events that take place immediately following fertilization, transitioning the zygote into embryogenesis. In vertebrates, egg activation encompasses the block to polyspermy and the completion of oocyte meiosis, which is coupled to the extrusion of the second polar body. Interestingly, in all sexually reproducing organisms tested to date the cellular events associated with egg activation are Ca2+-dependent (1Stricker S.A. Dev. Biol. 1999; 211: 157-176Crossref PubMed Scopus (583) Google Scholar). Importantly the Ca2+ signal at fertilization encodes the progression of these cellular events in a defined temporal sequence that ensures a functional egg-to-embryo transition (2Ducibella T. Huneau D. Angelichio E. Xu Z. Schultz R.M. Kopf G.S. Fissore R. Madoux S. Ozil J.P. Dev. Biol. 2002; 250: 280-291Crossref PubMed Google Scholar, 3Ozil J.P. Markoulaki S. Toth S. Matson S. Banrezes B. Knott J.G. Schultz R.M. Huneau D. Ducibella T. Dev. Biol. 2005; 282: 39-54Crossref PubMed Scopus (142) Google Scholar). The first order of business for the fertilized egg is to block polyspermy, which could be lethal to the embryo. This presents a particularly difficult problem for the large Xenopus oocyte. Therefore, this species employs a fast and slow blocks to polyspermy, both of which are Ca2+-dependent (4Machaca K. Qu Z. Kuruma A. Hartzell H.C. McCarty N. Fuller C.M. Calcium Activates Chloride Channels. Academic Press, San Diego2001: 3-39Google Scholar). In