Inactivation of histone H1 kinase by Ca2+ in rabbit oocytes

Abstract

The present study investigated the role of intracellular Ca2+ (Ca2+i) elevation on the inactivation of maturation promoting factor (MPF) in rabbit oocytes. The effects of the number of Ca2+ stimulations and of the amplitude of Ca2+i elevation on the profile of histone H1 kinase activity were determined. A Ca2+ stimulation consisted of transferring mature oocytes from culture medium to 0.3 M mannitol containing 0.1-1.0 mM CaCl2, and pulsing them at 1.25 kV/cm for 10 microseconds, or microinjecting 2-8 mM CaCl2 into the oocyte cytoplasm. The number of electrically-induced Ca2+ stimulations was varied, and amplitude of the Ca2+i rise was controlled by altering Ca2+ concentration in the pulsing medium or the injection pipette. Ca2+i concentration was determined with fura-2 dextran; oocytes were snap-frozen at indicated time points and assayed for H1 kinase activity. The activity was quantified by densitometry and expressed as a fraction of activity in nonstimulated oocytes. Electrically-mediated Ca2+i rises inactivated H1 kinase in a manner dependent on the number of Ca2+ stimulations. A single Ca2+ stimulation inactivated H1 kinase to 30-40% of its initial activity. However, H1 kinase inactivation was only transient, regardless of the amplitude of the electrically- or injection-mediated Ca2+i elevation. Increasing the number of Ca2+ stimulations helped to maintain H1 kinase activity at basal (pronuclear) levels. The results show the necessity of a threshold of Ca2+i concentration to trigger MPF inactivation, and suggest a role for the extended period of time over which Ca2+i oscillates at fertilization.