Inositol 1,4,5-Trisphosphate Receptors Are Downregulated in Mouse Oocytes in Response to Sperm or Adenophostin A but Not to Increases in Intracellular Ca2+ or Egg Activation

Abstract

Fertilization in mammals stimulates a series of Ca(2+) oscillations that continue for 3-4 h. Cell-cycle-dependent changes in the ability to release Ca(2+) are one mechanism that leads to the inhibition of Ca(2+) transients after fertilization. The downregulation of InsP(3)Rs at fertilization may be an additional mechanism for inhibiting Ca(2+) transients. In the present study we examine the mechanism of this InsP(3)R downregulation. We find that neither egg activation nor Ca(2+) transients are necessary or sufficient for the stimulation of InsP(3)R downregulation. First, parthenogenetic activation fails to stimulate downregulation. Second, downregulation persists when fertilization-induced Ca(2+) transients and egg activation are inhibited using BAPTA. Third, downregulation can be induced in immature oocytes that do not undergo egg activation. Other than fertilization, the only stimulus that downregulated InsP(3)Rs was microinjection of the potent InsP(3)R agonist adenophostin A. InsP(3)R downregulation was inhibited by the cysteine protease inhibitor ALLN but MG132 and lactacystin were not effective. Finally, we have injected maturing oocytes with adenophostin A and produced MII eggs depleted of InsP(3)Rs. We show that sperm-induced Ca(2+) signaling is inhibited in such InsP(3)R-depleted eggs. These data show that InsP(3)R binding is sufficient for downregulation and that Ca(2+) signaling at fertilization is mediated via the InsP(3)R.

Mammalian Fertilization: From Sperm Factor to Phospholipase Cζ

“…In addition, inhibition of IP 3 R type 1 by injection of a function blocking antibody, or by injection of heparin, a competitive IP 3 R inhibitor, prevents [Ca 2+ ] i oscillations induced by sperm or sperm extracts in hamster and mouse eggs (Miyazaki et al, 1992;Wu et al, 1997). Moreover, the degradation of IP 3 R type 1 that is observed in eggs following sperm entry is an indication of IP 3 production (Brind et al, 2000;Jellerette et al, 2000), since it is well documented in somatic cells that IP 3 R type 1 degradation only occurs as a consequence of IP 3 binding (Bokkala and Joseph, 1997;Oberdorf et al, 1999). In this context, it is worth noting that monitoring of IP 3 production in single mammalian zygotes is still not technically possible, although increases in intracellular IP 3 levels have been detected after fertilization in sea urchin and Xenopus laevis eggs (Stith et al, 1993;Snow et al, 1996;Lee and Shen, 1998;Sato et al, 2000;Kuroda et al, 2001) and after injection of porcine sperm extracts into Xenopus eggs (Wu et al, 2001b (Swann, 1992;Ayabe et al, 1995;Yue et al, 1995;Machaty et al, 1997).…”

Calcium wave pacemakers in eggs

“…Among the three known isoforms of IP3R found in somatic cells (Taylor et al, 1999), IP3R1 is the most prevalent and functionally important isoform in the egg [Xenopus (Runft et al, 1999); mammals (Miyazaki et al, 1993;Fissore et al, 1999;Brind et al, 2000); ascidian (Kyozuka et al, 1998)]. Low levels of IP3R2 and IP3R3 have been reported in mouse eggs (Fissore et al, 1999), but their physiological roles remain unclear (Brind et al, 2000;Jellerette et al, 2000).…”

“…Low levels of IP3R2 and IP3R3 have been reported in mouse eggs (Fissore et al, 1999), but their physiological roles remain unclear (Brind et al, 2000;Jellerette et al, 2000). The other family of Ca 2+ release channels (RyRs) is present on the cortical ER of sea urchin eggs (McPherson et al, 1992) and in ascidian (Albrieux et al, 2000) and mouse eggs (Ayabe et al, 1995).…”

Molecular cloning, testicular postnatal expression, and oocyte-activating potential of porcine phospholipase Cζ

“…Oocytes activated by Ca 2C oscillation resume the second meiosis and extrude a second polar body, and form male and female pronuclei. It is known that this Ca 2C oscillation is derived from the inositol 1,4,5-triphosphate (IP 3 )-signaling pathway in oocytes, which stimulates IP 3 receptors of the endoplasmic reticulum as the intracellular store of Ca 2C (Miyazaki et al 1993, Brind et al 2000, Jellerette et al 2000. However, the molecular mechanism that results in the activation of the IP 3 -signaling pathway followed by sperm penetration has not been elucidated.…”