The role of actin filaments in the organization of the endoplasmic reticulum in honeybee photoreceptor cells

Close to the bases of the photoreceptive microvilli, arthropod photoreceptors contain a dense network of endoplasmic reticulum that is involved in the regulation of the intracellular calcium concentration, and in the biogenesis of the photoreceptive membrane. Here, we examine the role of the cytoskeleton in organizing this submicrovillar endoplasmic reticulum in honeybee photoreceptors. Immunofluorescence microscopy of taxol-stabilized specimens, and electron-microscopic examination of high-pressure frozen, freeze-substituted retinae demonstrate that the submicrovillar cytoplasm lacks microtubules. The submicrovillar region contains a conspicuous F-actin system that codistributes with the submicrovillar endoplasmic reticulum. Incubation of retinal tissue with cytochalasin B leads to depolymerization of the submicrovillar F-actin system, and to disorganization and disintegration of the submicrovillar endoplasmic reticulum, indicating that an intact F-actin cytoskeleton is required to maintain the architecture of this domain of the endoplasmic reticulum. We have also developed a permeabilized cell model in order to study the physiological requirements for the interaction of the endoplasmic reticulum with actin filaments. The association of submicrovillar endoplasmic reticulum with actin filaments appears to be independent of ATP, Ca²⁺ and Mg²⁺, suggesting a tight static anchorage.     

Structural interactions of actin filaments and endoplasmic reticulum in honeybee photoreceptor cells

Summary Fluorescent phallotoxins and heavy meromyosin were used to reveal the organization of the actin cytoskeleton in honeybee photoreceptor cells, and the relationship of actin filaments to the submicrovillar, palisade-like cisternae of the endoplasmic reticulum (ER). Bundles of unipolar actin filaments (pointed end towards the cell center) protrude from the microvillar bases and extend through cytoplasmic bridges that traverse the submicrovillar ER. Within the cytoplasmic bridges, the filaments are regularly spaced and tightly apposed to the ER membrane. In addition, actin filaments are deployed close to the microvillar bases to form a loose web. Actin filaments are scarce in cell areas remote from the rhabdom; these areas contain microtubule-associated ER domains. The results suggest that the actin system of the submicrovillar cytoplasm shapes the submicrovillar ER cisternae, and that the distinct ER domains interact with different cytoskeletal elements.     

Orai3 channel is the 2-APB-induced endoplasmic reticulum calcium leak

We have studied in HeLa cells the molecular nature of the 2-APB induced ER Ca²⁺ leak using synthetic Ca²⁺ indicators that report changes in both the cytoplasmic ([Ca²⁺]_i) and the luminal ER ([Ca²⁺]_ER) Ca²⁺ concentrations. We have tested the hypothesis that Orai channels participate in the 2-APB-induced ER Ca²⁺ leak that was characterized in the companion paper. The expression of the dominant negative Orai1 E106A mutant, which has been reported to block the activity of all three types of Orai channels, inhibited the effect of 2-APB on the [Ca²⁺]_ER but did not decrease the ER Ca²⁺ leak after thapsigargin (TG). Orai3 channel, but neither Orai1 nor Orai2, colocalizes with expressed IP₃R and only Orai3 channel supported the 2-APB-induced ER Ca²⁺ leak, while Orai1 and Orai2 inhibited this type of ER Ca²⁺ leak. Decreasing the expression of Orai3 inhibited the 2-APB-induced ER Ca²⁺ leak but did not modify the ER Ca²⁺ leak revealed by inhibition of SERCA pumps with TG. However, reducing the expression of Orai3 channel resulted in larger [Ca²⁺]_i response after TG but only when the ER store had been overloaded with Ca²⁺ by eliminating the acidic internal Ca²⁺ store with bafilomycin. These data suggest that Orai3 channel does not participate in the TG-revealed ER Ca²⁺ leak but forms an ER Ca²⁺ leak channel that is limiting the overloading with Ca²⁺ of the ER store.     

Membrane traffic from the endoplasmic reticulum to the Golgi apparatus is disturbed by an inhibitor of the Ca2+-ATPase in the ER

Summary An electron microscopic study of cress (Lepidium sativum L.) roots treated with cyclopiazonic acid (CPA), an inhibitor of the Ca²⁺-ATPase in the endoplasmic reticulum (ER) has been carried out. Drastic changes in the endomembrane system of the secretory root cap cells were observed. After treatment with CPA dense spherical or elliptoidal aggregates of ER (diameter 2–4 m) were formed in addition to the randomly distributed ER cisternae characteristic for control cells. The formation of ER aggregates indicates that in spite of an inhibition of the Ca²⁺ -ATPase in the ER by CPA, membrane synthesis in the ER continued. The ER aggregates are interpreted as a reservoir of ER membrane material newly synthesized during the 2 h CPA-treatment. Hypertrophied Golgi cisternae and secretory vesicles, which are characteristic for secretory cells under control conditions, were completely absent. Additionally the shape of the Golgi stacks was flat and the diameter of the cisternae was shortened by about one third. These phenomena are indicative of an inactive state of the Golgi apparatus. The cellular organization of both other cell types of the root cap, meristematic cells and statocytes, was not visibly affected by CPA, both having a relatively low secretory activity. The formation of ER aggregates as well as the reduction of Golgi compartments are indications for the existence of a unidirectional transport of membrane material from the ER to the Golgi. It is suggested that the membrane traffic from the ER to the Golgi apparatus is regulated by the cytosolic and/or luminal calcium concentration in secretory cells of the root cap.Abbreviations CPA cyclopiazonic acid - ER endoplasmic reticulum     

Reliable measurement of free Ca2+ concentrations in the ER lumen using Mag-Fluo-4

Synthetic Ca²⁺ indicators are widely used to report changes in free [Ca²⁺], usually in the cytosol but also within organelles. Mag-Fluo-4, loaded into the endoplasmic reticulum (ER) by incubating cells with Mag-Fluo-4 AM, has been used to measure changes in free [Ca²⁺] within the ER, where the free [Ca²⁺] is estimated to be between 100 μM and 1 mM. Many results are consistent with Mag-Fluo-4 reliably reporting changes in free [Ca²⁺] within the ER, but the results are difficult to reconcile with the affinity of Mag-Fluo-4 for Ca²⁺ measured in vitro (K_D^Ca ∼22 μM). Using an antibody to quench the fluorescence of indicator that leaked from the ER, we established that the affinity of Mag-Fluo-4 within the ER is much lower (K_D^Ca ∼1 mM) than that measured in vitro. We show that partially de-esterified Mag-Fluo-4 has reduced affinity for Ca²⁺, suggesting that incomplete de-esterification of Mag-Fluo-4 AM within the ER provides indicators with affinities for Ca²⁺ that are both appropriate for the ER lumen and capable of reporting a wide range of free [Ca²⁺].     

Ouabain enhances exocytosis through the regulation of calcium handling by the endoplasmic reticulum of chromaffin cells

The augmentation of neurotransmitter and hormone release produced by ouabain inhibition of plasmalemmal Na⁺/K⁺-ATPase (NKA) is well established. However, the mechanism underlying this action is still controversial. Here we have shown that in bovine adrenal chromaffin cells ouabain diminished the mobility of chromaffin vesicles, an indication of greater number of docked vesicles at subplasmalemmal exocytotic sites. On the other hand, ouabain augmented the number of vesicles undergoing exocytosis in response to a K⁺ pulse, rather than the quantal size of single vesicles. Furthermore, ouabain produced a tiny and slow Ca²⁺ release from the endoplasmic reticulum (ER) and gradually augmented the transient elevations of the cytosolic Ca²⁺ concentrations ([Ca²⁺]_c) triggered by K⁺ pulses. These effects were paralleled by gradual increments of the transient catecholamine release responses triggered by sequential K⁺ pulses applied to chromaffin cell populations treated with ouabain. Both, the increases of K⁺-elicited [Ca²⁺]_c and secretion in ouabain-treated cells were blocked by thapsigargin (THAPSI), 2-aminoethoxydiphenyl borate (2-APB) and caffeine. These results are compatible with the view that ouabain may enhance the ER Ca²⁺ load and facilitate the Ca²⁺-induced-Ca²⁺ release (CICR) component of the [Ca²⁺]_c signal generated during K⁺ depolarisation. This could explain the potentiating effects of ouabain on exocytosis.     

Calreticulin is an upstream regulator of calcineurin

Ca²⁺ is a signalling molecule involved in virtually every aspect of cell function. The endoplasmic reticulum (ER) is an important and dynamic organelle responsible for storage of the majority of intracellular Ca²⁺. Within the ER lumen are proteins that function as Ca²⁺ buffers and/or molecular chaperones including calreticulin, a multifunctional Ca²⁺-binding protein. Calreticulin-deficiency is lethal in utero due to impaired cardiac development. In the absence of calreticulin Ca²⁺ storage capacity in the ER and InsP₃ receptor mediated Ca²⁺ release from ER are compromised. Remarkably, over-expression of constitutively active calcineurin in the hearts of calreticulin deficient mice rescues them from embryonic lethality and produces live calreticulin deficient animals. These observations provide first evidence that calreticulin is a key upstream regulator of calcineurin in the Ca²⁺-signalling cascade and they highlight the importance of ER during early stages of cellular commitment and tissue development during organogenesis.     

Activation of galanin receptor 2 stimulates large conductance Ca-dependent K (BK) channels through the IP3 pathway in human embryonic kidney (HEK293) cells

The large conductance Ca²⁺-activated K⁺ (BK) channels are widely distributed in the brain, and act as intracellular calcium sensors in neurons. They play an important feedback role in controlling Ca²⁺ flux and Ca²⁺-dependent processes, including neurotransmitter release and cellular excitability. In this study, the effects of the neuropeptide galanin on BK channels were examined by determining the whole-cell currents and single-channel activities in human embryonic kidney (HEK293) cells co-expressing GalR2 and the BK alpha subunit. Galanin enhanced the currents of BK channels, in a concentration-dependent and PTX-independent manner, with an ED50 value of 71.8 ± 16.9 nM. This activation was mediated by GalR2, since its agonist AR-M1896 mimicked the effect of galanin, and since galanin did not facilitate BK currents in cells co-expressing cDNAs of BK and GalR1 or GalR3. The galanin-induced BK current persisted after replacement with Ca²⁺-free solution, suggesting that extracellular Ca²⁺ is not essential. Chelating intracellular Ca²⁺ by either the slow Ca²⁺ buffer EGTA or the fast Ca²⁺ buffer BAPTA abolished galanin-mediated activation of BK channels, indicating the important role of intracellular Ca²⁺. The role of Ca²⁺ efflux from the sarcoplasmic reticulum/endoplasmic reticulum (SR/ER) was confirmed by application of thapsigargin, an irreversible inhibitor that depletes Ca²⁺ from SR/ER. Moreover, the inositol-1,4,5-triphosphate receptor (IP3R) was identified as the mediator responsible for increased intracellular Ca²⁺ activating BK channels. Taken together, activation of GalR2 leads to elevation of intracellular Ca²⁺ is due to Ca²⁺ efflux from ER through IP3R sequentially opening BK channels.     

Calcium stores in drosophila retina revisited: An electron-cytochemical localization of calcium

According to the commonly accepted model of phototransduction in insects, the endoplasmic submicrovillar cisternae (SMC) is the main element of Ca²⁺ homeostasis in the photoreceptor cell. It is generally believed that the light-induced inositol 1,4,5-trisphosphate-mediated Ca²⁺ release from these putative Ca²⁺ stores is an obligatory intermediate event in the cascade of phototransduction resulting in activation of the light-sensitive channels. However, it appears that this model fits well mainly the insects with a fused rhabdom and large SMC. In this study it has been found electron-cytochemically that in Drosophila that has an open rhabdom and miniature SMC, the reaction product for Ca²⁺ (calcium oxalate) accumulates not in SMC, but in the specialized extracellular compartment formed by the distended lacunae at the bases of the rhabdomeral microvilli, which sometimes deeply invaginate into the submicrovillar cytoplasm. It is suggested that in Drosophila and probably in other insects with an open rhabdom, it is this extracellular compartment, not SMC, that together with the calphotin area of the cytoplasm functions as a key element of the Ca²⁺ homeostasis in the photoreceptor cell.     

Ca2+-induced Ca2+ Release in Chromaffin Cells Seen from inside the ER with Targeted Aequorin

The presence and physiological role of Ca²⁺-induced Ca²⁺ release (CICR) in nonmuscle excitable cells has been investigated only indirectly through measurements of cytosolic [Ca²⁺] ([Ca²⁺]_c). Using targeted aequorin, we have directly monitored [Ca²⁺] changes inside the ER ([Ca²⁺]_ER) in bovine adrenal chromaffin cells. Ca²⁺ entry induced by cell depolarization triggered a transient Ca²⁺ release from the ER that was highly dependent on [Ca²⁺]_ER and sensitized by low concentrations of caffeine. Caffeine-induced Ca²⁺ release was quantal in nature due to modulation by [Ca²⁺]_ER. Whereas caffeine released essentially all the Ca²⁺ from the ER, inositol 1,4,5-trisphosphate (InsP₃)- producing agonists released only 60–80%. Both InsP₃ and caffeine emptied completely the ER in digitonin-permeabilized cells whereas cyclic ADP-ribose had no effect. Ryanodine induced permanent emptying of the Ca²⁺ stores in a use-dependent manner after activation by caffeine. Fast confocal [Ca²⁺]_c measurements showed that the wave of [Ca²⁺]_c induced by 100-ms depolarizing pulses in voltage-clamped cells was delayed and reduced in intensity in ryanodine-treated cells. Our results indicate that the ER of chromaffin cells behaves mostly as a single homogeneous thapsigargin-sensitive Ca²⁺ pool that can release Ca²⁺ both via InsP₃ receptors or CICR.     

Calcium stores in drosophila retina revisited: An electron-cytochemical localization of calcium

According to the commonly accepted model of phototransduction in insects, the endoplasmic submicrovillar cisternae (SMC) is the main element of Ca²⁺ homeostasis in the photoreceptor cell. It is generally believed that the light-induced inositol 1,4,5-trisphosphate-mediated Ca²⁺ release from these putative Ca²⁺ stores is an obligatory intermediate event in the cascade of phototransduction resulting in activation of the light-sensitive channels. However, it appears that this model fits well mainly the insects with a fused rhabdom and large SMC. In this study it has been found electron-cytochemically that in Drosophila that has an open rhabdom and miniature SMC, the reaction product for Ca²⁺ (calcium oxalate) accumulates not in SMC, but in the specialized extracellular compartment formed by the distended lacunae at the bases of the rhabdomeral microvilli, which sometimes deeply invaginate into the submicrovillar cytoplasm. It is suggested that in Drosophila and probably in other insects with an open rhabdom, it is this extracellular compartment, not SMC, that together with the calphotin area of the cytoplasm functions as a key element of the Ca²⁺ homeostasis in the photoreceptor cell.     

Visualization and quantification of endoplasmic reticulum Ca in renal cells using confocal microscopy and Fluo5F

Sarcoplasmic/endoplasmic reticulum (ER) Ca²⁺ is the most abundant store of intracellular Ca²⁺, and its release is an important trigger of physiological and cell death pathways. Previous work in our laboratory revealed the importance of ER Ca²⁺ in toxicant-induced renal proximal tubular cell (RPTC) death. The purpose of this study was to evaluate the use of confocal microscopy and Fluo5F, a low affinity Ca²⁺ indicator, to directly monitor changes in RPTC ER Ca²⁺. Fluo5F staining reflected ER Ca²⁺, resolved ER structure, and showed no colocalization with tetramethyl rhodamine methyl ester (TMRM), a marker of mitochondrial membrane potential. Thapsigargin, an ER Ca²⁺ pump inhibitor, decreased ER fluorescence by 30% and 55% at 5 and 15 min, respectively, whereas A23187, a Ca²⁺ ionophore caused more rapid ER Ca²⁺ release (55% and 75% decrease in fluorescence at 5 and 15 min).Carbonylcyanide-p-trifluoromethoxyphenylhydrazone (FCCP), a mitochondrial uncoupler, added at the end of the experiment, further decreased ER fluorescence after thapsigargin treatment, revealing that thapsigargin did not release all ER Ca²⁺. In contrast, FCCP did not decrease ER fluorescence after A23187 treatment, suggesting complete ER Ca²⁺ release. ER Ca²⁺ release in response to A23187 or thapsigargin resulted in a modest but significant decrease in mitochondrial membrane potential. These data provide evidence that confocal microscopy and Fluo5F are useful and effective tools for directly monitoring ER Ca²⁺ in live cells.     

Ca transfer from the ER to mitochondria: When, how and why

The heterogenous subcellular distribution of a wide array of channels, pumps and exchangers allows extracellular stimuli to induce increases in cytoplasmic Ca²⁺ concentration ([Ca²⁺]_c) with highly defined spatial and temporal patterns, that in turn induce specific cellular responses (e.g. contraction, secretion, proliferation or cell death). In this extreme complexity, the role of mitochondria was considered marginal, till the direct measurement with targeted indicators allowed to appreciate that rapid and large increases of the [Ca²⁺] in the mitochondrial matrix ([Ca²⁺]_m) invariably follow the cytosolic rises. Given the low affinity of the mitochondrial Ca²⁺ transporters, the close proximity to the endoplasmic reticulum (ER) Ca²⁺-releasing channels was shown to be responsible for the prompt responsiveness of mitochondria. In this review, we will summarize the current knowledge of: i) the mitochondrial and ER Ca²⁺ channels mediating the ion transfer, ii) the structural and molecular foundations of the signaling contacts between the two organelles, iii) the functional consequences of the [Ca²⁺]_m increases, and iv) the effects of oncogene-mediated signals on mitochondrial Ca²⁺ homeostasis. Despite the rapid progress carried out in the latest years, a deeper molecular understanding is still needed to unlock the secrets of Ca²⁺ signaling machinery.     

Bax Inhibitor-1-mediated Ca leak is decreased by cytosolic acidosis

Bax Inhibitor-1 (BI-1) is an evolutionarily conserved six-transmembrane domain endoplasmic reticulum (ER)-localized protein that protects against ER stress-induced apoptotic cell death. This function is closely connected to its ability to lower steady-state ER Ca²⁺ levels. Recently, we elucidated BI-1's Ca²⁺-channel pore in the C-terminal part of the protein and identified the critical amino acids of its pore. Based on these insights, a Ca²⁺-channel pore-dead mutant BI-1 (BI-1^D213R) was developed. We determined whether BI-1 behaves as a bona fide H⁺/Ca²⁺ antiporter or as an ER Ca²⁺-leak channel by investigating the effect of pH on unidirectional Ca²⁺-efflux rates. At pH 6.8, wild-type BI-1 expression in BI-1^−/− cells increased the ER Ca²⁺-leak rate, correlating with its localization in the ER compartment. In contrast, BI-1^D231R expression in BI-1^−/−, despite its ER localization, did not increase the ER Ca²⁺-leak rate. However, at pH < 6.8, the BI-1-mediated ER Ca²⁺ leak was blocked. Finally, a peptide representing the Ca²⁺-channel pore of BI-1 promoting Ca²⁺ flux from the ER was used. Lowering the pH from 6.8 to 6.0 completely abolished the ability of the BI-1 peptide to mediate Ca²⁺ flux from the ER. We propose that this pH dependence is due to two aspartic acid residues critical for the function of the Ca²⁺-channel pore and located in the ER membrane-dipping domain, which facilitates the protonation of these residues.     

The role of MATER in endoplasmic reticulum distribution and calcium homeostasis in mouse oocytes

Ca²⁺ oscillations are a hallmark of mammalian fertilization and play a central role in the activation of development. The calcium required for these oscillations is primarily derived from the endoplasmic reticulum (ER), which accumulates in clusters at the microvillar subcortex during oocyte maturation. The migration of the ER to the cortex during maturation is thought to play an important role in rendering the ER competent to generate the calcium transients, and the redistribution of ER is believed to be primarily mediated by microtubules and microfilaments. We have previously shown that the oocyte- and early embryo-restricted maternal effect gene Mater (Nlrp5) localizes to, and is required for, formation of the oocyte cytoplasmic lattices, a tubulin-containing structure that appears to play an important role in organelle positioning and distribution during oocyte maturation. Given these observations, we hypothesized that Mater may also be required for ER redistribution and Ca²⁺ homeostasis in oocytes. To test this hypothesis, we first investigated ER localization in metaphase-II Mater^tm/tm (hypomorph) oocytes and found ER clusters to be less abundant at the microvillar cortex when compared to wild type oocytes. To examine the potential mechanisms by which MATER mediates ER redistribution, we tested whether tubulin expression levels and localization were affected in the mutant oocytes and found that the Triton-insoluble fraction of tubulin was significantly decreased in Mater^tm/tm oocytes. To identify potential functional defects associated with these ER abnormalities, we next set out to investigate if the pattern of Ca²⁺ oscillations was altered in Mater^tm/tm oocytes after fertilization in vitro. Intriguingly, Ca²⁺ oscillations in Mater^tm/tm oocytes exhibited a significantly lower first peak amplitude and a higher frequency when compared to wild type oocytes. We then found that the Ca²⁺ oscillation defect in Mater^tm/tm oocytes was likely caused by a reduced amount of Ca²⁺ in the ER stores. Taken together, these observations support the hypothesis that MATER is required for ER distribution and Ca²⁺ homeostasis in oocytes, likely due to defects in lattice-mediated ER positioning and/or redistribution.     

Endoplasmic reticulum stress in insulin resistance and diabetes

The endoplasmic reticulum is the main intracellular Ca²⁺ store for Ca²⁺ release during cell signaling. There are different strategies to avoid ER Ca²⁺ depletion. Release channels utilize first Ca²⁺-bound to proteins and this minimizes the reduction of the free luminal [Ca²⁺]. However, if release channels stay open after exhaustion of Ca²⁺-bound to proteins, then the reduction of the free luminal ER [Ca²⁺] (via STIM proteins) activates Ca²⁺ entry at the plasma membrane to restore the ER Ca²⁺ load, which will work provided that SERCA pump is active. Nevertheless, there are several noxious conditions that result in decreased activity of the SERCA pump such as oxidative stress, inflammatory cytokines, and saturated fatty acids, among others. These conditions result in a deficient restoration of the ER [Ca²⁺] and lead to the ER stress response that should facilitate recovery of the ER. However, if the stressful condition persists then ER stress ends up triggering cell death and the ensuing degenerative process leads to diverse pathologies; particularly insulin resistance, diabetes and several of the complications associated with diabetes. This scenario suggests that limiting ER stress should decrease the incidence of diabetes and the mobility and mortality associated with this illness.     

Mitochondrial Ca uptake with and without the formation of high-Ca microdomains

The mitochondrial Ca²⁺ uniporter has low affinity for Ca²⁺, therefore it has been assumed that submicromolar Ca²⁺ signals cannot induce mitochondrial Ca²⁺ uptake. The close apposition of the plasma membrane or the endoplamic reticulum (ER) to the mitochondria and the limited Ca²⁺ diffusion in the cytoplasm result in the formation of perimitochondrial high-Ca²⁺ microdomains (HCMDs) capable of activating mitochondrial Ca²⁺ uptake. The possibility of mitochondrial Ca²⁺ uptake at low submicromolar [Ca²⁺]_c has not yet been generally accepted.Earlier we found in permeabilized glomerulosa, luteal and pancreatic β cells that [Ca²⁺]_m increased when [Ca²⁺]_c was raised from 60 nM to less than 200 nM. Here we report data obtained from H295R (adrenocortical) cells transfected with ER-targeted GFP. Cytoplasmic Ca²⁺ response to angiotensin II was different in mitochondrion-rich and mitochondrion-free domains. The mitochondrial Ca²⁺ response to angiotensin II correlated with GFP fluorescence indicating the vicinity of ER. When the cells were exposed to K⁺ (inducing Ca²⁺ influx), no correlation was found between the mitochondrial Ca²⁺ signal and the vicinity of the plasma membrane or the ER. The results presented here provide evidence that mitochondrial Ca²⁺ uptake may occur both with and without the formation of HCMDs within the same cell.     

Calcium- and inositol polyphosphate-sensitivity of the calcium-sequestering endoplasmic reticulum in the photoreceptor cells of the honeybee drone

Summary The photoreceptor cells in the honeybee drone contain an elaborate Ca²⁺-sequestering endoplasmic reticulum (ER). We measured Ca-oxalate formation within the ER of permeabilized retinal slices with a microphotometer and studied the kinetics of Ca²⁺-uptake into the ER and the properties of Ins(1,4,5)P₃-induced Ca²⁺-release.The ATP-dependent Ca²⁺-uptake mechanism has a high affinity for Ca²⁺: Uptake rate was half maximal at Ca²⁺ _free 0.6 M.Addition of Ins(1,4,5)P₃ caused a persistent depression of Ca-oxalate formation due to Ca²⁺ -release from the ER. The Ins(1,4,5)P₃-dependent Ca²⁺-release mechanism has a high affinity (half maximal rate with 0.2 M Ins(1,4,5)P₃) and a high specificity for Ins(1,4,5)P₃: Ins(2,4,5)P₃ was 6 times, Ins(1,3,4,5)P₄ was 15 times less potent in inducing Ca²⁺-release. 3 M Ins(1,4)P₂ had no detectable effect. The sensitivity for Ins(1,4,5)P₃ was maximal between 280 nM and 1.6 M Ca²⁺ _free and decreased at higher and lower Ca²⁺-concentrations.Our data show that the ER in invertebrate photoreceptor cells is an effective Ca²⁺ -sink and an Ins(1,4,5)P₃-sensitive Ca²⁺-source. We support the idea (Payne et al. 1988) that the ER-network close to the photoreceptive membrane, the submicrovillar cisternae (SMC), are the light- and Ins(1,4,5)P₃-sensitive Ca²⁺-stores.Abbreviations ER endoplasmic reticulum - Ins(1,4,5)P ₃ D-inositol 1,4,5-trisphosphate - Ins(1,3,4)P ₃ D-inositol 1,3,4-trisphosphate - Ins(2,4,5)P ₃ D-inositol 2,4,5-trisphosphate - Ins(1,4)P ₂ D-inositol 1,4-bisphosphate - Ins(1,3,4,5)P ₄ D-inositol 1,3,4,5-tetrakisphosphate - SMC submicrovillar cisternae - [Ca ²⁺]_i intracellular free Ca²⁺-concentration     

Darier disease : A disease model of impaired calcium homeostasis in the skin

The importance of extracellular calcium in epidermal differentiation and intra-epidermal cohesion has been recognized for many years. Darier disease (DD) was the first genetic skin disease caused by abnormal epidermal calcium homeostasis to be identified. DD is characterized by loss of cell-to-cell adhesion and abnormal keratinization. DD is caused by genetic defects in ATP2A2 encoding the sarco/endoplasmic reticulum Ca²⁺-ATPase isoform 2 (SERCA2). SERCA2 is a calcium pump of the endoplasmic reticulum (ER) transporting Ca²⁺ from the cytosol to the lumen of ER. ATP2A2 mutations lead to loss of Ca²⁺ transport by SERCA2 resulting in decreased ER Ca²⁺ concentration in Darier keratinocytes. Here, we review the role of SERCA2 pumps and calcium in normal epidermis, and we discuss the consequences of ATP2A2 mutations on Ca²⁺ signaling in DD. This article is part of a Special Issue entitled: 11th European Symposium on Calcium.