Sperm-triggered calcium oscillations during meiosis in ascidian oocytes first pause, restart, then stop: correlations with cell cycle kinase activity.

Abstract

We have investigated the relationship between the sperm-triggered Ca2+ oscillations and the activities of two cell cycle kinases (MPF activity and MAP kinase activity) at fertilisation of Ascidiella aspersa oocytes. Maturation Promoting Factor (MPF) activity is elevated in the metaphase I (MI)-arrested unfertilised oocyte (as measured by phosphorylation of exogenous histone H1) and falls 5 minutes after fertilisation to remain at low levels for 5 minutes. The first polar body (pb1) is extruded when the MPF activity is low. The MPF activity is elevated again 15 minutes after fertilisation and finally becomes inactivated 25 minutes after fertilisation when the pb2 is extruded. MAP kinase activity increases from an initially elevated level to reach maximal activity 10 minutes after fertilisation and subsequently falls to reach low levels 25 minutes after fertilisation. Sperm trigger a series of Ca2+ oscillations that pause for 5 minutes while only the MPF activity is low and are present when both MPF and MAP kinase activity are elevated. We next attempted to determine whether the second phase of calcium oscillations is required to reactivate the MPF activity that precedes extrusion of the second polar body. To do this, we triggered a monotonic Ca2+ signal. This leads to the inactivation of MPF followed by MPF reactivation. The MPF activity then remains elevated for an extended period of time. During this period, the chromatin remains condensed and a metaphase II (MII) spindle forms. Fertilisation of these MII oocytes triggers extrusion of pb2 in 7 minutes. Interestingly, the second phase of Ca2+ oscillations is completely absent when MII oocytes are fertilised. Thus, in both MI and MII oocytes, the sperm-triggered Ca2+ oscillations follow the MPF activity. Finally we discuss our finding that the Ca2+ release system remains sensitive during the metaphase-like state (including the period when the Ca2+ oscillations pause).