Ca(2+) (cyt) negatively regulates the initiation of oocyte maturation

Ca(2+) is a ubiquitous intracellular messenger that is important for cell cycle progression. Genetic and biochemical evidence support a role for Ca(2+) in mitosis. In contrast, there has been a long-standing debate as to whether Ca(2+) signals are required for oocyte meiosis. Here, we show that cytoplasmic Ca(2+) (Ca(2+) (cyt)) plays a dual role during Xenopus oocyte maturation. Ca(2+) signals are dispensable for meiosis entry (germinal vesicle breakdown and chromosome condensation), but are required for the completion of meiosis I. Interestingly, in the absence of Ca(2+) (cyt) signals oocytes enter meiosis more rapidly due to faster activation of the MAPK-maturation promoting factor (MPF) kinase cascade. This Ca(2+)-dependent negative regulation of the cell cycle machinery (MAPK-MPF cascade) is due to Ca(2+) (cyt) acting downstream of protein kinase A but upstream of Mos (a MAPK kinase kinase). Therefore, high Ca(2+) (cyt) delays meiosis entry by negatively regulating the initiation of the MAPK-MPF cascade. These results show that Ca(2+) modulates both the cell cycle machinery and nuclear maturation during meiosis

Induction of maturation-promoting factor during Xenopus oocyte maturation uncouples Ca(2+) store depletion from store-operated Ca(2+) entry

Department of Physiology and Biophysics, University of Arkansas Medical Science, Little Rock, AR 72205 During oocyte maturation, eggs acquire the ability to generate specialized Ca(2+) signals in response to sperm entry. Such Ca(2+) signals are crucial for egg activation and the initiation of embryonic development. We examined the regulation during Xenopus oocyte maturation of store-operated Ca(2+) entry (SOCE), an important Ca(2+) influx pathway in oocytes and other nonexcitable cells. We have previously shown that SOCE inactivates during Xenopus oocyte meiosis. SOCE inactivation may be important in preventing premature egg activation. In this study, we investigated the correlation between SOCE inactivation and the Mos–mitogen-activated protein kinase (MAPK)–maturation-promoting factor (MPF) kinase cascade, which drives Xenopus oocyte maturation. SOCE inactivation at germinal vesicle breakdown coincides with an increase in the levels of MAPK and MPF. By differentially inducing Mos, MAPK, and MPF, we demonstrate that the activation of MPF is necessary for SOCE inactivation during oocyte maturation. In contrast, sustained high levels of Mos kinase and the MAPK cascade have no effect on SOCE activation. We further show that preactivated SOCE is not inactivated by MPF, suggesting that MPF does not block Ca(2+) influx through SOCE channels, but rather inhibits coupling between store depletion and SOCE activation

IP3 receptors and store-operated Ca2+ entry: a license to fill.

Inositol 1,4,5-trisphosphate receptors (IP3Rs) are widely expressed intracellular Ca2+ channels that evoke large local increases in cytosolic Ca2+ concentration. By depleting the ER of Ca2+, IP3Rs also activate store-operated Ca2+ entry (SOCE). Immobile IP3Rs close to the plasma membrane (PM) are the only IP3Rs that respond to physiological stimuli. The association of these 'licensed' IP3Rs with the ER-PM junctions where STIM interacts with Orai PM Ca2+ channels may define the autonomous functional unit for SOCE. Ca2+ entering cells through SOCE can be delivered directly to specific effectors, or it may reach them only after the Ca2+ has been sequestered by the ER and then released through IP3Rs. This 'tunnelling' of Ca2+ through the ER to IP3Rs selectively delivers Ca2+ to different effectors.Wellcome Trust BBSR

Ca2+ tunnelling through the ER lumen as a mechanism for delivering Ca2+ entering via store-operated Ca2+ channels to specific target sites

Ca2+ signaling is perhaps the most universal and versatile mechanism regulating a wide range of cellular processes. Because of the many different calcium-binding proteins distributed throughout cells, signaling precision requires localized rises in the cytosolic Ca2+ concentration. In electrically non-excitable cells, for example epithelial cells, this is achieved by primary release of Ca2+ from the endoplasmic reticulum via Ca2+ release channels placed close to the physiological target. Because any rise in the cytosolic Ca2+ concentration activates Ca2+ extrusion, and in order for cells not to run out of Ca2+, there is a need for compensatory Ca2+ uptake from the extracellular fluid. This Ca2+ uptake occurs through a process known as store-operated Ca2+ entry. Ideally Ca2+ entering the cell should not diffuse to the target site through the cytosol, as this would potentially activate undesirable processes. Ca2+ tunneling through the lumen of the endoplasmic reticulum is a mechanism for delivering Ca2+ entering via store-operated Ca2+ channels to specific target sites, and this process has been described in considerable detail in pancreatic acinar cells and oocytes. Here we review the most important evidence and present a generalized concept

Rabphilin localizes with the cell actin cytoskeleton and stimulates association of granules with F-actin cross-linked by α-actinin

In endocrine cell, granules accumulate within an F-actin-rich region below the plasma membrane. The mechanisms involved in this process are largely unknown. Rabphilin is a cytosolic protein that is expressed in neurons and neuroendocrine cells and binds with high affinity to members of the Rab3 family of GTPases localized to synaptic vesicles and dense core granules. Rabphilin also interacts with alpha-actinin, a protein that cross-links F-actin into bundles and networks and associates with the granule membrane. Here we asked whether rabphilin, in addition to its granule localization, also interacts with the cell actin cytoskeleton. Immunofluorescence and immunoelectron microscopy show that rabphilin localizes to the sub-plasmalemmal actin cytoskeleton both in neuroendocrine and unspecialized cells. By using purified components, it is found that association of rabphilin with F-actin is dependent on added alpha-actinin. In an in vitro assay, granules, unlike endosomes or mitochondria, associate with F-actin cross-linked by alpha-actinin. Rabphilin is shown to stimulate this process. Rabphilin enhances by approximately 8-fold the granule ability to localize within regions of elevated concentration of cross-linked F-actin. These results suggest that rabphilin, by interacting with alpha-actinin, organizes the cell cytoskeleton to facilitate granule localization within F-actin-rich regions

Constitutive recycling of the store-operated Ca2+ channel Orai1 and its internalization during meiosis

Orai1 trafficking between the cell membrane and endosomes during meiosis requires caveolin and is important for tuning calcium signaling during oocyte maturation

A longer isoform of Stim1 is a negative SOCE regulator but increases cAMP-modulated NFAT signaling

Alternative splicing is a potent modifier of protein function. Stro mal interaction molecule 1 (Stim1) is the essential activator of store-operated Ca2+ entry (SOCE) triggering activation of transcrip tion factors. Here, we characterize Stim1A, a splice variant with an additional 31 amino acid domain inserted in frame within its cytosolic domain. Prominent expression of exon A is found in astro cytes, heart, kidney, and testes. Full-length Stim1A functions as a dominant-negative regulator of SOCE and ICRAC, facilitating sequence-specific fast calcium-dependent inactivation and desta bilizing gating of Orai channels. Downregulation or absence of native Stim1A results in increased SOCE. Despite reducing SOCE, Stim1A leads to increased NFAT translocation. Differential proteo mics revealed an interference of Stim1A with the cAMP-SOCE crosstalk by altered modulation of phosphodiesterase 8 (PDE8), resulting in reduced cAMP degradation and increased PIP5K activ ity, facilitating NFAT activation. Our study uncovers a hitherto unknown mechanism regulating NFAT activation and indicates that cell-type-specific splicing of Stim1 is a potent means to regu late the NFAT signalosome and cAMP-SOCE crosstalk

Endoplasmic Reticulum Remodeling Tunes IP3-Dependent Ca2+ Release Sensitivity

The activation of vertebrate development at fertilization relies on IP3-dependent Ca2+ release, a pathway that is sensitized during oocyte maturation. This sensitization has been shown to correlate with the remodeling of the endoplasmic reticulum into large ER patches, however the mechanisms involved are not clear. Here we show that IP3 receptors within ER patches have a higher sensitivity to IP3 than those in the neighboring reticular ER. The lateral diffusion rate of IP3 receptors in both ER domains is similar, and ER patches dynamically fuse with reticular ER, arguing that IP3 receptors exchange freely between the two ER compartments. These results suggest that increasing the density of IP3 receptors through ER remodeling is sufficient to sensitize IP3-dependent Ca2+ release. Mathematical modeling supports this concept of ‘geometric sensitization’ of IP3 receptors as a population, and argues that it depends on enhanced Ca2+-dependent cooperativity at sub-threshold IP3 concentrations. This represents a novel mechanism of tuning the sensitivity of IP3 receptors through ER remodeling during meiosis