Calcium/Calmodulin-Dependent Protein Kinase II and Calmodulin: Regulators of the Meiotic Spindle in Mouse Eggs

Stein

Proc. Natl. Acad. Sci. U.S.A.

et al. 2009

106

110

Fertilization triggers a rise in intracellular Ca 2+ concentration ([Ca 2+ ] i ) in the egg that initiates a series of events known as egg activation. These events include cortical granule exocytosis that establishes a block to polyspermy, resumption of meiosis, and recruitment of maternal mRNAs into polysomes for translation. Several calcium-dependent proteins, including calcium/calmodulin-dependent protein kinase II (CaMKII), have been implicated in egg activation. However, the precise role of CaMKII in mediating specific events of egg activation and the identity of the isoform(s) present in mouse eggs have not been unequivocally established. Through targeted deletion of the γ isoform of CaMKII, we find that CaMKIIγ is the predominant CaMKII isoform in mouse eggs and that it is essential for egg activation. Although CaMKIIγ −/− eggs exhibit a normal pattern of Ca 2+ oscillations after insemination and undergo cortical granule exocytosis, they fail to resume meiosis or to recruit maternal mRNAs. Surprisingly, we find that the recruitment of maternal mRNAs does not directly depend on CaMKII, but requires elevated [Ca 2+ ] i and metaphase II exit. We conclude that CaMKIIγ specifically controls mouse egg activation by regulating cell cycle resumption.T he transition from a fertilization-competent mammalian egg to a developing embryo entails a sequence of events collectively known as egg activation. Early events of egg activation include modifications of the zona pellucida (ZP) that prevent polyspermy, exit from metaphase II arrest, and completion of meiosis, whereas late events include recruitment of maternal mRNAs into polysomes for translation and formation of male and female pronuclei (1). In all animal species studied to date, a rise in intracellular calcium ([Ca 2+ ] i ) is the universal trigger of all of the events of egg activation (EEA) (2). Release of a spermspecific phospholipase C isoform (PLCζ) likely initiates the inositol 1,4,5-trisphosphate (IP 3 )-mediated increase in [Ca 2+ ] i (3). In mammalian eggs, this increase in [Ca 2+ ] i takes the form of repetitive Ca 2+ transients (oscillations) that last several hours (4, 5). Although Ca 2+ oscillations can induce all of the EEA, individual events require different numbers of [Ca 2+ ] i transients to be initiated and completed, with early events requiring fewer oscillations than late events (6).The pathways that connect the rise in [Ca 2+ ] i to the different EEA have only been partially elucidated. Protein kinases, including calcium/calmodulin-dependent protein kinase II (CaM-KII), and the phosphatase calcineurin have been postulated as integrators of the Ca 2+ signal during egg activation (7-9). However, direct genetic evidence for involvement of these enzymes through loss-of-function studies is lacking, leaving open questions as to whether any single enzyme is essential for all of the EEA or whether these Ca 2+ -dependent events are mediated by different effectors.The CaMKII family of serine/threonine kinases mediates many cellular responses to C...

Section: Discussionmentioning

confidence: 99%

The γ isoform of CaM kinase II controls mouse egg activation by regulating cell cycle resumption

Stein

Proc. Natl. Acad. Sci. U.S.A.

et al. 2009

106

110

Journal of Reproduction and Development

“…The activated APC/C, with its E3 ligase activity, leads to subsequent proteolysis of target proteins via the 2 6 S p r o t e a s o m e c o m p l e x ; p 3 4 c d c 2 k i n a s e inactivation via the destruction of cyclin B and securing occur via this mechanism [7][8][9]. In mammals, intracellular Ca 2+ elevation induced by electrical pulse triggers the destruction of cyclin B and meiotic resumption from MII arrest, but the treatment of these oocytes with myristoylatedautocamtide-2-related inhibitory peptide (AIP) prevents decrease of p34 cdc2 kinase activity [10][11][12]. Furthermore, it has been reported that injection of a constitutively active CaMKII construct is sufficient to induce activation in mouse oocytes [13].…”

mentioning

confidence: 99%

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

Ito

Kaneko

Hirabayashi

2006

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.

“…CamKII is a prime example of this premise. Its localization specifically changes spatially and temporally to drive second polar body emission (Winston and Maro, 1995;Johnson et al, 1998). Consequently, based on the kaz data gathered thus far, we propose that kaz is likely a key component that associates with the cytoskeleton to coordinate the mechanism(s) that drive specific developmental transitions, including second polar body, compaction, and possibly postimplantation development.…”

Section: Fig 12 A-fmentioning

confidence: 87%

Dynamics and unexpected localization of the plakin binding protein, kazrin, in mouse eggs and early embryos

Gallicano

Foshay

Pengetnze

et al. 2005

Developmental Dynamics

The cell uses the cytoskeleton in virtually every aspect of cell survival and function. One primary function of the cytoskeleton is to connect to and stabilize intercellular junctions. To accomplish this task, microtubules, actin filaments, and intermediate filaments utilize cytolinker proteins, which physically bind the cytoskeletal filament to the core proteins of the adhesion junction. The plakin family of linker proteins have been in the spotlight recently as critical components for embryo survival and, when mutated, the cause of diseases such as muscular dystrophy and cardiomyopathies. Here, we reveal the dynamics of a recently discovered plakin binding protein, kazrin (kaz), during early mouse development. Kaz was originally found in adult tissues, primarily epidermis, linking periplakin to the plasma membrane and colocalizing with desmoplakin in desmosomes. Using reverse transcriptase-polymerase chain reaction, Western blots, and confocal microscopy, we found kaz in unfertilized eggs associated with the spindle apparatus and cytoskeletal sheets. As quickly as 5 min after egg activation, kaz relocates to a diffuse peri-spindle position, followed 20 -30 min later by clear localization to the presumptive cytokinetic ring. Before the blastocyst stage of development, kaz associates with the nuclear matrix in a cell cycle-dependent manner, and also associates with the cytoplasmic actin cytoskeleton. After blastocyst formation, kaz localization and potential function(s) become highly complex as it is found associating with cell-cell junctions, the cytoskeleton, and nucleus. Postimplantation stages of development reveal that kaz retains a multifunctional, tissue-specific role as it is detected at diverse locations in various embryonic tissue types.