Cell cycle-coupled [Ca 2+] i oscillations in mouse zygotes and function of the inositol 1,4,5-trisphosphate receptor-1

Abstract

Sperm entry in mammalian eggs initiates oscillations in the concentration of free calcium ([Ca 2+] i). In mouse eggs, oscillations start at metaphase II (MII) and conclude as the zygotes progress into interphase and commence pronuclear (PN) formation. The inositol 1,4,5-trisphosphate receptor (IP 3R-1), which underlies the oscillations, undergoes degradation during this transition, suggesting that one or more of the eggs' Ca 2+-releasing machinery components may be regulated in a cell cycle-dependent manner, thereby coordinating [Ca 2+] i responses with the cell cycle. To ascertain the site(s) of interaction, we initiated oscillations at different stages of the cell cycle in zygotes with different IP 3R-1 mass. In addition to sperm, we used two other agonists: porcine sperm factor (pSF), which stimulates production of IP 3, and adenophostin A, a non-hydrolyzable analogue of IP 3. None of the agonists tested induced oscillations at interphase, suggesting that neither decreased IP 3R-1 mass nor lack of production or excessive IP 3 degradation can account for the insensitivity to IP 3 at this stage. Moreover, the releasable Ca 2+ content of the stores did not change by interphase, but it did decrease by first mitosis. More importantly, experiments revealed that IP 3R-1 sensitivity and possibly IP 3 binding were altered at interphase, and our data demonstrate stage-specific IP 3R-1 phosphorylation by M-phase kinases. Accordingly, increasing the activity of M-phase kinases restored the oscillatory-permissive state in zygotes. We therefore propose that the restriction of oscillations in mouse zygotes to the metaphase stage may be coordinated at the level of IP 3R-1 and that this involves cell cycle stage-specific receptor phosphorylation.