The Regulation of Calcium/Calmodulin-dependent Protein Kinase II during Oocyte Activation in the Rat

Abstract

Abstract. Increases in intracellular Ca2+ are required for oocyte activation and subsequent development. Calmodulin-dependent protein kinase II (CaMKII) plays a crucial role in oocyte activation. However, how CaMKII is regulated during this process is not well characterized. We show here for the first time in rat oocytes that CaMKII is phosphorylated during oocyte activation. CaMKII phosphorylation was suppressed by KN93, a CaMKII inhibitor, but not KN92, which is the inactive analogue of KN93. Electrical stimulation of rat oocytes resulted in degradation of both cyclin B and Mos, presumably due a rise in Ca 2+ induced by the electrical pulse. KN93 blocked the degradation of both proteins induced by the electrical pulse. Addition of a protein phosphatase inhibitor, okadaic acid (OA), further increased the amount of CaMKII and also increased the amount of phosphorylated enzyme. Importantly, in oocytes undergoing spontaneous activation, accumulation and phosphorylation of CaMKII also occurs in a time-dependent manner. Consistent with this, addition of KN93 inhibited spontaneous activation. Collectively, our results show that CaMKII is phosphorylated during oocyte activation and that this phosphorylation is involved in inactivation of p34 cdc2 kinase and somewhat involved in degradation of Mos. Furthermore, CaMKII phosphorylation is negatively regulated by a protein phosphatase.

Myristoylated alanine-rich C kinase substrate, but not Ca2+/calmodulin-dependent protein kinase II, is the mediator in cortical granules exocytosis

“…Two articles supporting our findings have been published lately; Ito et al (2006) have shown that the CaMKII phosphorylation occurring during egg activation can be suppressed by KN93, a CaMKII inhibitor. They claimed that this phosphorylation is involved in inactivation of p34 cdc2 kinase and degradation of Mos.…”

“…The involvement of CaMKII in egg activation events has been the topic of many studies conducted lately. While CaMKII involvement in promoting RMII is well established (Markoulaki et al 2003, Sun 2003, Madgwich et al 2005, Ito et al 2006, Knott et al 2006, its involvement in CGE is still controversial.…”

Superovulatory Response, Oocyte Spontaneous Activation, and Embryo Development in WMN/Nrs Inbred Rats

“…Ito et al [13,14] showed that ovulated rat oocytes, especially those from the Wistar strain, cannot be arrested at metaphase II due to the loss of activity of p34 cdc2 kinase maintained by high activity of Mos/MeK/MAPK pathways which in turn are negatively regulated by Ca 2+ -dependent proteasome. Therefore, incubation of rat oocytes in Ca 2+ -free conditions or addition of BAPTA-AM, a Ca 2+ chelator, to the incubation medium effectively inhibited OSA [10,11,14].…”

“…Under the conditions employed in this study, most mature WMN/Nrs females showed 4-day estrous cycles. An average of 12.2 embryos (range 6-17 embryos) and an average litter size of 10.2 (n=31, range [5][6][7][8][9][10][11][12][13][14] were observed during natural ovulation and mating (Ueno M., personal communication). Body weights of WMN/Nrs females were 30.2 ± 0.5 g at 3 weeks of age, 50.7 ± 0.8 g at 4 weeks, 76.2 ± 1.2 g at 5 weeks, 97.9 ± 1.1 g at 6 weeks, 117.0 ± 1.4 g at 7 weeks, 134.5 ± 1.6 g at 8 weeks and 147.0 ± 2.0 g at 9 weeks of age (n=15) (Yamada Y., personal communication).…”

Kinetics of Nuclear Status and Kinase Activities During In Vitro Maturation of Canine Oocytes

“…In general, the nuclear status of in vitro matured canine oocytes is evaluated using [27][28][29][30]. It has also been demonstrated that p34 cdc2 kinase activity decreases depending on the length of the culture period, a process designated 'aging' after the progression to MII [31][32][33][34]. This aging is involved in abnormal formation of pronuclei after in vitro fertilization or artificial activation in pigs [35,36].…”