Modulation of Ca2+ entry by polypeptides of the inositol 1,4,5-trisphosphate receptor (IP3R) that bind transient receptor potential (TRP): Evidence for roles of TRP and IP3R in store depletion-activated Ca2+ entry (original) (raw)
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Journal of Biological Chemistry, 2001
The mechanism for coupling between Ca 2؉ stores and store-operated channels (SOCs) is an important but unresolved question. Although SOCs have not been molecularly identified, transient receptor potential (TRP) channels share a number of operational parameters with SOCs. The question of whether activation of SOCs and TRP channels is mediated by the inositol 1,4,5-trisphosphate receptor (InsP 3 R) was examined using the permeant InsP 3 R antagonist, 2-aminoethoxydiphenyl borate (2-APB) in both mammalian and invertebrate systems. In HEK293 cells stably transfected with human TRPC3 channels, the actions of 2-APB to block carbachol-induced InsP 3 R-mediated store release and carbachol-induced Sr 2؉ entry through TRPC3 channels were both reversed at high agonist levels, suggesting InsP 3 Rs mediate TRPC3 activation. However, electroretinogram recordings of the lightinduced current in Drosophila revealed that the TRP channel-mediated responses in wild-type as well as trp and trpl mutant flies were all inhibited by 2-APB. This action of 2-APB is likely InsP 3 R-independent since InsP 3 Rs are dispensable for the light response. We used triple InsP 3 R knockout DT40 chicken B-cells to further assess the role of InsP 3 Rs in SOC activation. 45 Ca 2؉ flux analysis revealed that although DT40 wildtype cells retained normal InsP 3 Rs mediating 2-APBsensitive Ca 2؉ release, the DT40InsP 3 R-k/o cells were devoid of functional InsP 3 Rs. Using intact cells, all parameters of Ca 2؉ store function and SOC activation were identical in DT40wt and DT40InsP 3 R-k/o cells. Moreover, in both cell lines SOC activation was completely blocked by 2-APB, and the kinetics of action of 2-APB on SOCs (time dependence and IC 50 ) were identical. The results indicate that (a) the action of 2-APB on Ca 2؉ entry is not mediated by the InsP 3 R and (b) the effects of 2-APB provide evidence for an important similarity in the function of invertebrate TRP channels, mammalian TRP channels, and mammalian storeoperated channels. . The abbreviations used are: ER, endoplasmic reticulum; TRP, transient receptor potential, InsP 3 , inositol 1,4,5-trisphosphate; fura-2/AM, fura-2 acetoxymethylester; 2-APB, 2-aminoethoxydiphenyl borate; SOC, store-operated channel; ERG, electroretinogram; HEK cells, human embryonic kidney cells.
Requirement of the Inositol Trisphosphate Receptor for Activation of Store-Operated Ca2+ Channels
Science, 2000
The coupling mechanism between endoplasmic reticulum (ER) calcium ion (Ca2+) stores and plasma membrane (PM) store-operated channels (SOCs) is crucial to Ca2+ signaling but has eluded detection. SOCs may be functionally related to the TRP family of receptor-operated channels. Direct comparison of endogenous SOCs with stably expressed TRP3 channels in human embryonic kidney (HEK293) cells revealed that TRP3 channels differ in being store independent. However, condensed cortical F-actin prevented activation of both SOC and TRP3 channels, which suggests that ER-PM interactions underlie coupling of both channels. A cell-permeant inhibitor of inositol trisphosphate receptor (InsP3R) function, 2-aminoethoxydiphenyl borate, prevented both receptor-induced TRP3 activation and store-induced SOC activation. It is concluded that InsP3Rs mediate both SOC and TRP channel opening and that the InsP3R is essential for maintaining coupling between store emptying and physiological activation of SOCs.
Biochimica et Biophysica Acta (BBA) - Molecular Cell Research, 2018
Inositol 1,4,5-trisphosphate receptors (IP 3 R) are the most widely expressed intracellular Ca 2+ release channels. Their activation by IP 3 and Ca 2+ allows Ca 2+ to pass rapidly from the ER lumen to the cytosol. The resulting increase in cytosolic [Ca 2+ ] may directly regulate cytosolic effectors or fuel Ca 2+ uptake by other organelles, while the decrease in ER luminal [Ca 2+ ] stimulates store-operated Ca 2+ entry (SOCE). We are close to understanding the structural basis of both IP 3 R activation, and the interactions between the ER Ca 2+-sensor, STIM, and the plasma membrane Ca 2+ channel, Orai, that lead to SOCE. IP 3 Rs are the usual means through which extracellular stimuli, through ER Ca 2+ release, stimulate SOCE. Here, we review evidence that the IP 3 Rs most likely to respond to IP 3 are optimally placed to allow regulation of SOCE. We also consider evidence that IP 3 Rs may regulate SOCE downstream of their ability to deplete ER Ca 2+ stores. Finally, we review evidence that IP 3 Rs in the plasma membrane can also directly mediate Ca 2+ entry in some cells.
The Journal of General Physiology, 2012
generated in the cytoplasm in response to extracellular stimuli binds to and activates InsP 3 R channels to release Ca 2+ stored in Correspondence to Don-On Daniel Mak: d m a k @ m a i l . m e d . u p e n n . e d u Abbreviations used in this paper: HEDTA, hydroxyethylethylenediaminetriacetic acid; InsP 3 , inositol 1,4,5-trisphosphate; InsP 3 R, InsP 3 receptor; lum-out, luminal-side-out; P o , open probability; r-InsP 3 R-3, rat type 3 InsP 3 R. the ER lumen into the cytoplasm, generating diverse local and global [Ca 2+ ] i signals . Whereas much is known regarding the intricate regulation of InsP 3 R channel gating by multiple processes-binding of cytoplasmic ligands (Ca 2+ , InsP 3 , and ATP 4 ), posttranslational modifications, interactions with proteins, clustering, dif-The ubiquitous inositol 1,4,5-trisphosphate (InsP 3 ) receptor (InsP 3 R) Ca 2+ release channel plays a central role in the generation and modulation of intracellular Ca 2+ signals, and is intricately regulated by multiple mechanisms including cytoplasmic ligand (InsP 3 , free Ca 2+ , free ATP 4 ) binding, posttranslational modifications, and interactions with cytoplasmic and endoplasmic reticulum (ER) luminal proteins. However, regulation of InsP 3 R channel activity by free Ca 2+ in the ER lumen ([Ca 2+ ] ER ) remains poorly understood because of limitations of Ca 2+ flux measurements and imaging techniques. Here, we used nuclear patch-clamp experiments in excised luminal-side-out configuration with perfusion solution exchange to study the effects of [Ca 2+ ] ER on homotetrameric rat type 3 InsP 3 R channel activity. In optimal [Ca 2+ ] i and subsaturating [InsP 3 ], jumps of [Ca 2+ ] ER from 70 nM to 300 µM reduced channel activity significantly. This inhibition was abrogated by saturating InsP 3 but restored when [Ca 2+ ] ER was raised to 1.1 mM. In suboptimal [Ca 2+ ] i , jumps of [Ca 2+ ] ER (70 nM to 300 µM) enhanced channel activity. Thus, [Ca 2+ ] ER effects on channel activity exhibited a biphasic dependence on [Ca 2+ ] i . In addition, the effect of high [Ca 2+ ] ER was attenuated when a voltage was applied to oppose Ca 2+ flux through the channel. These observations can be accounted for by Ca 2+ flux driven through the open InsP 3 R channel by [Ca 2+ ] ER , raising local [Ca 2+
Proceedings of the National Academy of Sciences, 2005
Virtually all functions of a cell are influenced by cytoplasmic [Ca 2؉ ] increases. Inositol 1,4,5-trisphosphate receptor (IP3R) channels, located in the endoplasmic reticulum (ER), release Ca 2؉ in response to binding of the second messenger, IP3. IP3Rs thus are part of the information chain interpreting external signals and transforming them into cytoplasmic Ca 2؉ transients. IP3Rs function as tetramers, each unit comprising an N-terminal ligand-binding domain (LBD) and a C-terminal channel domain linked by a long regulatory region. It is not yet understood how the binding of IP3 to the LBD regulates the gating properties of the channel. Here, we use the expression of IP3 binding protein domains tethered to the surface of the endoplasmic reticulum (ER) to show that the all-helical domain of the IP3R LBD is capable of depleting the ER Ca 2؉ pools by opening the endogenous IP3Rs, even without IP3 binding. This effect requires the domain to be within 50 Å of the ER membrane and is impaired by the presence of the N-terminal inhibitory segment on the LBD. These findings raise the possibility that the helical domain of the LBD functions as an effector module possibly interacting with the channel domain, thereby being part of the gating mechanisms by which the IP3-induced conformational change within the LBD regulates Ca 2؉ release.
Journal of Biological Chemistry, 1999
Inositol 1,4,5-trisphosphate (InsP 3) mobilizes intracellular Ca 2؉ by binding to its receptor (InsP 3 R), an endoplasmic reticulum-localized Ca 2؉ release channel. Patch clamp electrophysiology of Xenopus oocyte nuclei was used to study the effects of cytoplasmic ATP concentration on the cytoplasmic Ca 2؉ ([Ca 2؉ ] i) dependence of single type 1 InsP 3 R channels in native endoplasmic reticulum membrane. Cytoplasmic ATP free-acid ([ATP] i), but not the MgATP complex, activated gating of the InsP 3-liganded InsP 3 R, by stabilizing open channel state(s) and destabilizing the closed state(s). Activation was associated with a reduction of the half-maximal activating [Ca 2؉ ] i from 500 ؎ 50 nM in 0 [ATP] i to 29 ؎ 4 nM in 9.5 mM [ATP] i , with apparent ATP affinity ؍ 0.27 ؎ 0.04 mM, similar to in vivo concentrations. In contrast, ATP was without effect on maximum open probability or the Hill coefficient for Ca 2؉ activation. Thus, ATP enhances gating of the InsP 3 R by allosteric regulation of the Ca 2؉ sensitivity of the Ca 2؉ activation sites of the channel. By regulating the Ca 2؉-induced Ca 2؉ release properties of the InsP 3 R, ATP may play an important role in shaping cytoplasmic Ca 2؉ signals, possibly linking cell metabolic state to important Ca 2؉-dependent processes.
Biochemical Journal, 2000
In the present study we have investigated the role of inositol 1,4,5-trisphosphate (IP $ ), functional IP $ receptors (IP $ Rs) and the human homologue of the Drosophila transient receptor potential (Trp) channel, human Trp1 (hTrp1), in store-mediated Ca# + entry (SMCE) in human platelets. Inhibition of IP $ recycling using Li + , or the inhibition of IP $ Rs using xestospongin C, both resulted in the inhibition of SMCE activation following Ca# + store depletion using thapsigargin. Co-immunoprecipitation experiments indicated that endogenously expressed hTrp1 couples with
The Journal of biological chemistry, 2010
There is a body of evidence suggesting that Ca 2+ -handling proteins assemble into signaling complexes required for a fine regulation of Ca 2+ signals, events that regulate a variety of critical cellular processes. Canonical transient receptor potential (TRPC) and Orai proteins have both been proposed to form Ca 2+permeable channels mediating Ca 2+ entry upon agonist stimulation. A number of studies have demonstrated that inositol 1,4,5-trisphosphate receptors (IP 3 Rs) interact with plasma membrane TRPC channels; however at present there is no evidence supporting the interaction between Orai proteins and IP 3 Rs. Here we report that treatment with thapsigargin or cellular agonists results in association of Orai1 with the types I and II IP 3 Rs. In addition, we have found that TRPC3, RACK1 (receptor for activated Ckinase-1) and STIM1 (stromal interaction molecule 1) interact with Orai1 upon stimulation with agonists. TRPC3 expression silencing prevented both the interaction of Orai1 with TRPC3 and, more interestingly, the association of Orai1 with the type I IP 3 R, but not with the type II IP 3 R, thus suggesting that TRPC3 selectively mediates the interaction between Orai1 and the type I IP 3 R. In addition, TRPC3 expression silencing attenuated ATPand CCh-stimulated interaction between RACK1 and the type I IP 3 R, as well as Ca 2+ release and entry. In conclusion, our results indicate that agonist stimulation results in the formation of an Orai1-STIM1-TRPC3-RACK1-type I IP 3 R complex, where TRPC3 plays a central role. This Ca 2+ signaling complex might be important for agonist induced both Ca 2+ release and entry.
Agonist-induced Ca2+ entry determined by inositol 1,4,5-trisphosphate recognition
Proceedings of the National Academy of Sciences, 2004
It has been considered that Ca 2؉ release is the causal trigger for Ca 2؉ entry after receptor activation. In DT40 B cells devoid of inositol 1,4,5-trisphosphate receptors (IP3R), the lack of Ca 2؉ entry in response to receptor activation is attributed to the absence of Ca 2؉ release. We reveal in this article that IP 3 R recognition of IP 3 determines agonist-induced Ca 2؉ entry (ACE), independent of its Ca 2؉ release activity. In DT40 IP3R ؊/؊ cells, endogenous ACE can be rescued with type 1 IP3R mutants (both a ⌬C-terminal truncation mutant and a D2550A pore mutant), which are defective in Ca 2؉ release channel activity. Thus, in response to B cell receptor activation, ACE is restored in an IP3R-dependent manner without Ca 2؉ store release. Conversely, ACE cannot be rescued with mutant IP3Rs lacking IP3 binding (both the ⌬90 -110 and R265Q IP3-binding site mutants). We conclude that an IP3-dependent conformational change in the IP3R, not endoplasmic reticulum Ca 2؉ pool release, triggers ACE. C a 2ϩ transients elicited in response to cell surface receptor activation by neurotransmitters, hormones, and other molecular messengers are major messengers of intracellular communication (1). Stimulation of G protein-coupled receptors, tyrosine kinase receptors, and nonreceptor tyrosine kinases activate phospholipase C (PLC), catalyzing the breakdown of phosphatidylinositol 4,5-bisphosphate (PIP 2 ) into the secondmessenger molecules: inositol 1,4,5-trisphosphate (IP 3 ) and diacylglycerol (DAG). IP 3 mediates rapid Ca 2ϩ store release by activating IP 3 receptors (IP 3 Rs) in the endoplasmic reticulum (ER), whereas DAG activates protein kinase C (PKC) (2). After this initial Ca 2ϩ release phase, external Ca 2ϩ enters through plasma membrane channels, providing a secondary and more prolonged Ca 2ϩ signal (1), a phenomenon designated here as agonist-induced Ca 2ϩ entry (ACE) (3).
The Journal of biological chemistry, 2000
The mechanism for coupling between Ca(2+) stores and store-operated channels (SOCs) is an important but unresolved question. SOC-mediated Ca(2+) entry is complex and may reflect more than one type of channel and coupling mechanism. To assess such possible divergence the function and coupling of SOCs was compared with two other distinct yet related Ca(2+) entry mechanisms. SOC coupling in DDT(1)MF-2 smooth muscle cells was prevented by the permeant inositol 1,4,5-trisphosphate (InsP(3)) receptor blockers, 2-aminoethoxydiphenyl borate (2-APB) and xestospongin C. In contrast, Ca(2+) entry induced by S-nitrosylation and potentiated by store depletion (Ma, H-T., Favre, C. J., Patterson, R. L., Stone, M. R., and Gill, D. L. (1999) J. Biol. Chem. 274, 35318-35324) was unaffected by 2-APB, suggesting that this entry mechanism is independent of InsP(3) receptors. The cycloalkyl lactamimide, MDL-12, 330A (MDL), prevented SOC activation (IC(50) 10 micrometer) and similarly completely blocked S...