Cloning and Expression of a Novel Mammalian Homolog ofDrosophila Transient Receptor Potential (Trp) Involved in Calcium Entry Secondary to Activation of Receptors Coupled by the Gq Class of G Protein

Boulay, Guylain; Zhu, Xi; Peyton, Mike; Jiang, Meisheng; Hurst, Raymond S; Stefani, Enrico; Birnbaumer, Lutz

doi:10.1074/jbc.272.47.29672

Cited by 318 publications

(272 citation statements)

References 69 publications

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“…Our results show that the externalization of TRPC6 in stably transfected HEK 293 cells preceded Ca 2ϩ entry and occurred at an agonist concentration lower than that required for Ca 2ϩ entry. Also, Ca 2ϩ store depletion by thapsigargin caused an externalization of TRPC6 but did not activate TRPC6 (14,15), suggesting that another component is necessary to activate TRPC6. An obvious component is DAG (15), since we have shown that OAG can promote the externalization and the activation of TRPC6.…”

Section: Discussionmentioning

confidence: 99%

“…Cell Culture and Preparation of Caveolae-related MicrodomainsStably transfected mTRPC6 HEK 293 cells, T6.11 (14), were maintained in Dulbecco's modified Eagle's medium supplemented with 10% fetal bovine serum and 400 g/ml G418 at 37°C in a humidified 5% CO 2 atmosphere. All steps in the preparation of caveolae-related microdomains (CRM) were carried out at 4°C.…”

Section: Methodsmentioning

confidence: 99%

“…Overexpression studies showed that the homotetramer channel consisting of TRPC1, TRPC4, or TRPC5 can be activated either by the depletion of the intracellular Ca 2ϩ pool (8 -10) or by the activation of G q protein-coupled receptor stimulation (11,12). TRPC3, TRPC6, and TRPC7 form a non-selective cationic channel that is activated by the stimulation of a G q protein-coupled receptor or by the exogenous application of diacylglycerol analogs (13)(14)(15)(16). The activity of TRPC channels can be modulated by Ca 2ϩ -calmodulin, which has been shown to bind directly to TRPC (17)(18)(19)(20)(21).…”

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confidence: 99%

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Exocytotic Insertion of TRPC6 Channel into the Plasma Membrane upon Gq Protein-coupled Receptor Activation

Cayouette¹,

Lussier

Mathieu

et al. 2004

Journal of Biological Chemistry

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172

126

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TRPC proteins are the mammalian homologues of the Drosophila transient receptor potential channel and are involved in calcium entry after agonist stimulation of non-excitable cells. Seven mammalian TRPCs have been cloned, and their mechanisms of activation and regulation are still the subject of intense research. TRPC proteins interact with the inositol 1,4,5-trisphosphate receptor, and the conformational coupling plays a critical role in the activation of calcium entry. Some evidence also supports an exocytotic mechanism as part of the activation of calcium entry. To investigate the possible involvement of exocytosis in TRPC6 activation, we evaluated the location of TRPC6 at the plasma membrane by biotinylation labeling of cell surface proteins and by indirect immunofluorescence marking of TRPC6 in stably transfected HEK 293 cells. We showed that when the muscarinic receptor was stimulated or the thapsigargin-induced intracellular calcium pool was depleted the level of TRPC6 at the plasma membrane increased. The carbachol concentration at which TRPC6 externalization occurred was lower than the concentration required to activate TRPC6. Externalization occurred within the first 30 s of stimulation, and TRPC6 remained at the plasma membrane as long as the stimulus was present. These results indicate that an exocytotic mechanism is involved in the activation of TRPC6.Increases in intracellular calcium ([Ca 2ϩ ] i ) regulate important cellular functions, including cell growth, differentiation, contraction, and secretion (1, 2). The increase in [Ca 2ϩ ] i is initiated by the activation of phospholipase C␤ or -␥ by a G q protein-coupled receptor or tyrosine kinase receptor, respectively, causing the hydrolysis of phosphatidylinositol 4,5-bisphosphate and generating two-second messengers, inositol 1,4,5-trisphosphate (IP 3 ) 1 and diacylglycerol. IP 3 binds to its specific receptor on the endoplasmic reticulum and causes the first phase of Ca 2ϩ mobilization by releasing Ca 2ϩ from the intracellular pool. The second phase involves Ca 2ϩ entry through the plasma membrane, which maintains [Ca 2ϩ ] i above basal levels. No cellular activity has ever been observed in the absence of this second phase of Ca 2ϩ mobilization.The TRPC protein family is involved in Ca 2ϩ entry (3-5). Seven genes have been identified in the mammalian genome by homology with Drosophila TRP and TRPL, two proteins that are responsible for the light-induced current in insect phototransduction. TRPC channels are probably composed of four subunits, each containing three to four ankyrin-like repeats, two predicted coiled-coil regions (one in the amino terminus and another one in the carboxyl terminus), and a membranespanning region. The amino acids that link the fifth and sixth transmembrane segments form the putative pore region loop. Recent studies on the subunit arrangement of native TRPC in rat brain synaptosomes and the overexpression of TRPC in various systems suggest that the TRPC protein family can be subdivided in two distinct groups and that...

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Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

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Exocytotic Insertion of TRPC6 Channel into the Plasma Membrane upon Gq Protein-coupled Receptor Activation

Cayouette¹,

Lussier

Mathieu

et al. 2004

Journal of Biological Chemistry

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172

126

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“…T3-9, T3-65, and T3a-10 are HEK-293 cell-derived cell clones that stably overexpress TRPC3 or TRPC3a tagged at their N termini with the HA epitope (MYPYDVPDYA) (12,19). T6-11 is a HEK-293 cell-derived clone that overexpresses the murine HA-tagged (N terminus) TRPC6 in stable form (56).…”

Section: Methodsmentioning

confidence: 99%

Orai proteins interact with TRPC channels and confer responsiveness to store depletion

Liao¹,

Erxleben

Yildirim³

et al. 2007

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

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The TRPC (C-type transient receptor potential) class of ion channels has been hypothesized to participate in store-operated Ca 2؉ entry (SOCE). Recently, however, STIM1 and Orai1 proteins have been proposed to form SOCE channels. Whether TRPCs participate in SOCE that is dependent on or regulated by Orai has not been explored. Here we show that Orai1 physically interacts with the N and C termini of TRPC3 and TRPC6, and that in cells overexpressing either TRPC3 or TRPC6 in a store-depletion insensitive manner, these TRPCs become sensitive to store depletion upon expression of an exogenous Orai. Thus, Orai-1, -2, and -3 enhanced thapsigargin-induced calcium entry by 50 -150% in cells stably overexpressing either TRPC3 or TRPC6. Orai1 expression had no significant effect on endogenous, thapsigargin-induced calcium entry in wildtype cells (HEK-293, COS1), in HEK cells expressing a thapsigarginsensitive variant of TRPC3 (TRPC3a), or in HEK cells overexpressing another membrane protein, V1aR. Single-channel cation currents present in membrane patches of TRPC3-overexpressing cells were suppressed by expression of Orai1. We propose that Orai proteins by interacting with TRPCs act as regulatory subunits that confer STIM1-mediated store depletion sensitivity to these channels.capacitative calcium entry ͉ ion channels ͉ thapsigargin ͉ transmembrane signaling

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“…It had long been speculated that, in mammalian cells, capacitative Ca# + entry (CCE), which occurs as a consequence of Ca# + store depletion following the activation of phospholipase C [4,5], might be mediated by channels that share sequence similarity with the Drosophila Trp proteins. Indeed, seven Trp homologues have been isolated from mammalian species in recent years, and heterologous expression has revealed a great degree of complexity as to the mode of activation, single-channel conductance and ion selectivity of different Trp homologues [6][7][8][9][10][11][12][13][14].…”

Section: Introductionmentioning

confidence: 99%

Increased inwardly rectifying potassium currents in HEK-293 cells expressing murine transient receptor potential 4

2001

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Drosophila transient receptor potential (Trp) and its mammalian homologues are postulated to form capacitative Ca# + entry or store-operated channels. Here we show that expression of murine Trp4 in HEK 293 cells also leads to an increase in inwardly rectifying K + currents. No similar increase was found in cell lines expressing Trp1, Trp3 or Trp6. Consistent with typical characteristics of inward rectifiers, the K + currents in Trp4-expressing cells were blocked by low millimolar concentrations of Cs + and Ba# + , but not by 1.2 mM Ca# + , and were only slightly inhibited by 5 mM tetraethylammonium. Single channel recordings of excised inside-out patches revealed the presence of two con-

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Cloning and Expression of a Novel Mammalian Homolog ofDrosophila Transient Receptor Potential (Trp) Involved in Calcium Entry Secondary to Activation of Receptors Coupled by the Gq Class of G Protein

Cited by 318 publications

References 69 publications

Exocytotic Insertion of TRPC6 Channel into the Plasma Membrane upon Gq Protein-coupled Receptor Activation

Exocytotic Insertion of TRPC6 Channel into the Plasma Membrane upon Gq Protein-coupled Receptor Activation

Orai proteins interact with TRPC channels and confer responsiveness to store depletion

Increased inwardly rectifying potassium currents in HEK-293 cells expressing murine transient receptor potential 4

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