Expression and relative abundance of short transient receptor potential channels in the rat renal microcirculation

In the resistance vessels of the renal microcirculation, store- and/or receptor-operated calcium entry contribute to the rise in vascular smooth muscle cell (VSMC) intracellular calcium concentration in response to vasoconstrictor hormones. Short transient receptor potential (TRPC) channels are widely expressed in mammalian tissues and are proposed mediators of voltage-independent cation entry in multiple cell types, including VSMCs. The seven members of the TRPC gene family (TRPC1-7) encode subunit proteins that are thought to form homo- and heterotetrameric channels that are differentially regulated depending on their subunit composition. In the present study, we demonstrate the relative abundance of TRPC mRNA and protein in freshly isolated rat renal resistance vessels, glomeruli, and aorta. TRPC1, 3, 4, 5, and 6 mRNA and protein were detected in both renal resistance vessels and aorta, whereas TRPC2 and TRPC7 mRNA were not expressed. TRPC1, 3, 5, and 6 protein was present in glomeruli. TRPC3 and TRPC6 protein levels were significantly greater in the renal resistance vessels, about six- to eightfold higher than in aorta. These data suggest that TRPC3 and TRPC6 may play a role in mediating voltage-independent calcium entry in renal resistance vessels that is functionally distinct from that in aorta.

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Cloning, expression and subcellular localization of two novel splice variants of mouse transient receptor potential channel 2

Biochemical Journal ◽

10.1042/bj3510115 ◽

2000 ◽

Vol 351 (1) ◽

pp. 115-122 ◽

Cited By ~ 26

Author(s):

Thomas HOFMANN ◽

Michael SCHAEFER ◽

Günter SCHULTZ ◽

Thomas GUDERMANN

Keyword(s):

Membrane Trafficking ◽

Fusion Proteins ◽

Transient Receptor Potential ◽

Splice Variants ◽

Receptor Potential ◽

Calcium Entry ◽

Transient Receptor Potential Channels ◽

Protein Fusion ◽

Hek 293 ◽

Transient Receptor

Transient receptor potential channels (TRPCs) are known as candidate molecular correlates of receptor-activated or store-operated calcium entry. While functional roles for most TRPCs have been suggested, the physiological relevance of TRPC2 remains obscure. Whereas human and bovine TRPC2 are candidate pseudogenes, full-length rodent TRPC2 transcripts have been reported. There is, however, considerable controversy concerning mRNA splicing, tissue distribution and the function of these proteins. We report the molecular cloning of two novel murine TRPC2 splice variants, mTRPC2α and mTRPC2β. mTRPC2α RNA is expressed at low levels in many tissues and cell systems, while mTRPC2β is exclusively and abundantly expressed in the vomeronasal organ (VNO). When expressed in human embryonic kidney (HEK)-293 cells, mTRPC2 did not enhance receptor- or store-activated calcium entry. In order to investigate the basis of such a functional defect, mTRPC2–green fluorescent protein fusion proteins were examined by confocal microscopy. Fusion proteins were retained in endomembranes when expressed in HEK-293 or other cells of epithelial or neuronal origin. Co-expression of TRPC2 with other TRPCs did not restore plasma-membrane trafficking. We conclude that TRPC2 may form functional channels in the cellular context of the VNO, but is unlikely to have a physiological function in other tissues. The sequences of mTRPC2α and mTRPC2β have been submitted to GenBank under the accession numbers AF230802 and AF230803 respectively.

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TRPC Channels in the SOCE Scenario

Cells ◽

10.3390/cells9010126 ◽

2020 ◽

Vol 9 (1) ◽

pp. 126 ◽

Cited By ~ 14

Author(s):

Jose J. Lopez ◽

Isaac Jardin ◽

Jose Sanchez-Collado ◽

Ginés M. Salido ◽

Tarik Smani ◽

...

Keyword(s):

Transient Receptor Potential ◽

Receptor Potential ◽

Cell Types ◽

Trpc Channels ◽

Crac Channels ◽

Relevant Role ◽

Transient Receptor ◽

Trpc Proteins ◽

Soc Channels

Transient receptor potential (TRP) proteins form non-selective Ca2+ permeable channels that contribute to the modulation of a number of physiological functions in a variety of cell types. Since the identification of TRP proteins in Drosophila, it is well known that these channels are activated by stimuli that induce PIP2 hydrolysis. The canonical TRP (TRPC) channels have long been suggested to be constituents of the store-operated Ca2+ (SOC) channels; however, none of the TRPC channels generate Ca2+ currents that resemble ICRAC. STIM1 and Orai1 have been identified as the components of the Ca2+ release-activated Ca2+ (CRAC) channels and there is a body of evidence supporting that STIM1 is able to gate Orai1 and TRPC1 in order to mediate non-selective cation currents named ISOC. STIM1 has been found to interact to and activate Orai1 and TRPC1 by different mechanisms and the involvement of TRPC1 in store-operated Ca2+ entry requires both STIM1 and Orai1. In addition to the participation of TRPC1 in the ISOC currents, TRPC1 and other TRPC proteins might play a relevant role modulating Orai1 channel function. This review summarizes the functional role of TRPC channels in the STIM1–Orai1 scenario.

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Pregnancy-enhanced store-operated Ca2+ channel function in uterine artery endothelial cells is associated with enhanced agonist-specific transient receptor potential channel 3-inositol 1,4,5-trisphosphate receptor 2 interaction

Journal of Endocrinology ◽

10.1677/joe.1.06773 ◽

2006 ◽

Vol 190 (2) ◽

pp. 385-395 ◽

Cited By ~ 24

Author(s):

Shannon M Gifford ◽

Fu-Xian Yi ◽

Ian M Bird

Keyword(s):

Endothelial Cells ◽

Uterine Artery ◽

Transient Receptor Potential ◽

Purinergic Receptor ◽

Receptor Potential ◽

Cell Types ◽

Transient Receptor Potential Channels ◽

Significant Difference ◽

Transient Receptor ◽

Trisphosphate Receptor

We have previously shown that endothelial cells (EC) derived from the uterine artery (UA) of both pregnant (P-UAEC) and nonpregnant (NP-UAEC) ewes show a biphasic intracellular free Ca2+ ([Ca2+]i) response after ATP stimulation. In each case, the initial transient peak, caused by the release of Ca2+ from the intracellular Ca2+ stores, is mediated by purinergic receptor-Y2 and is very similar in both cell types. However, the sustained phase in particular, caused by the influx of extracellular Ca2+, is heightened in the P-UAEC, and associates with an increased ability of the cells to demonstrate enhanced capacitative Ca2+ entry (CCE) via store-operated channels (SOCs). Herein we demonstrated that the difference in the sustained [Ca2+]i response is maintained for at least 30 min. When 2-aminoethoxydiphenyl borate (2APB) (an inhibitor of the inosital 1,4,5-trisphosphate receptor (IP3R) and possibly SOC) was used in conjunction with ATP, it was capable of completely inhibiting CCE. Since 2APB can inhibit SOC in some cell types and 2APB was capable of inhibiting CCE in the UAEC model, the role of SOC in CCE was first evaluated using the classical inhibitor La3+. The ATP-induced sustained phase was inhibited by 10 μM La3+, implying a role for SOC in the [Ca2+]i response. Since canonical transient receptor potential channels (TRPCs) have recently been identified as putative SOCs in many cell types, including EC, the expression levels of several isoforms were evaluated in UAEC. Expression of TRPC3 and TRPC6 channels in particular was detected, but no significant difference in expression level was found between NP- and P-UAEC. Nonetheless, we were able to show that IP3R2 interacts with TRPC3 in UAEC, forming a protein complex, and that this interaction is considerably enhanced in an agonist sensitive manner by pregnancy. Thus, while IP3R and TRPC isoforms are not altered in their expression by pregnancy, enhanced functional interaction of TRPC3 with IP3R2 may underlie pregnancy-enhanced CCE in the UAEC model and so explain the prolonged [Ca2+]i sustained phase seen in response to ATP.

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Abstract 164: Microdomain Specific Effects of Transient Receptor Potential Channels on Pathological Cardiac Hypertrophy and Myocyte Contractility

Circulation Research ◽

10.1161/res.113.suppl_1.a164 ◽

2013 ◽

Vol 113 (suppl_1) ◽

Author(s):

Catherine A Makarewich ◽

Hongyu Zhang ◽

Hui Gao ◽

Robert N Correll ◽

Jason M Duran ◽

...

Keyword(s):

Cardiac Hypertrophy ◽

Transient Receptor Potential ◽

Receptor Potential ◽

Transient Receptor Potential Channels ◽

Trpc Channels ◽

Ec Coupling ◽

Transient Receptor ◽

Myocyte Contractility ◽

Signaling Domains ◽

Ca2 Channels

Hypothesis: Ca2+ influx through transient receptor potential canonical (TRPC) channels and L-type Ca2+ channels (LTCCs) within caveolin-3 (Cav3) stabilized signaling microdomains provide a unique source of Ca2+ to activate pathologic cardiac hypertrophy through calcineurin (Cn)-mediated nuclear factor of activated T-cells (NFAT) signaling. We suggest that a distinct and separate population of TRPC channels localized in excitation-contraction (EC) coupling microdomains may have potent effects on myocyte contractility independent of Cav3 signaling domains. Methods and Results: Membrane localization studies and immunohistochemistry show that TRPC channels and LTCCs co-localize to Cav3 signaling microdomains. To explore a role for these caveolae based Ca2+ channels in the initiation of Cn-NFAT signaling we used an adenoviral NFAT-GFP reporter in cultured adult feline myocytes (AFMs). Infecting AFMs with ad-TRPC3 dramatically increased NFAT translocation, which was inhibited with dominant negative ad-dnTRPC6. Expression of a Cav3 targeted LTCC blocker (ad-Cav-Rem) reduced NFAT translocation while a targeted LTCC activator (ad-Cav-β2a) significantly increased NFAT activation. Neither LTCC modulator had significant effects on Ca2+ current or contractility in AFMs but we found that the expression of TRPC3 reduced myocyte contractility and induced spontaneous Ca2+ spark activity that was exacerbated by the DAG activator OAG. Moreover, dnTRPC6 blocked spontaneous Ca2+ sparks even in the presence of OAG. Immunohistochemistry analysis revealed the presence of TRPC channels in transverse tubules, consistent with the idea that they could have direct effects on EC coupling microdomains. Conclusions: Our data show that TRPC channels and LTCCs co-localize to Cav3 signaling domains where they generate a unique Ca2+ microenvironment that directly regulates Cn-NFAT signaling. Our findings also suggest that a separate and distinct population of TRPC channels within EC coupling microdomains cause reduced myocyte contractility by inducing SR Ca2+ leak and Ca2+ spark activity.

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Dynamic NHERF interaction with TRPC4/5 proteins is required for channel gating by diacylglycerol

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1612263114 ◽

2016 ◽

Vol 114 (1) ◽

pp. E37-E46 ◽

Cited By ~ 52

Author(s):

Ursula Storch ◽

Anna-Lena Forst ◽

Franziska Pardatscher ◽

Serap Erdogmus ◽

Maximilian Philipp ◽

...

Keyword(s):

Transient Receptor Potential ◽

Receptor Potential ◽

Transient Receptor Potential Channels ◽

Activation Mechanism ◽

Binding Motif ◽

Trpc Channels ◽

Channel Activity ◽

C Terminus ◽

Potential Channels ◽

Transient Receptor

The activation mechanism of the classical transient receptor potential channels TRPC4 and -5 via the Gq/11 protein-phospholipase C (PLC) signaling pathway has remained elusive so far. In contrast to all other TRPC channels, the PLC product diacylglycerol (DAG) is not sufficient for channel activation, whereas TRPC4/5 channel activity is potentiated by phosphatidylinositol 4,5-bisphosphate (PIP2) depletion. As a characteristic structural feature, TRPC4/5 channels contain a C-terminal PDZ-binding motif allowing for binding of the scaffolding proteins Na+/H+ exchanger regulatory factor (NHERF) 1 and 2. PKC inhibition or the exchange of threonine for alanine in the C-terminal PDZ-binding motif conferred DAG sensitivity to the channel. Altogether, we present a DAG-mediated activation mechanism for TRPC4/5 channels tightly regulated by NHERF1/2 interaction. PIP2 depletion evokes a C-terminal conformational change of TRPC5 proteins leading to dynamic dissociation of NHERF1/2 from the C terminus of TRPC5 as a prerequisite for DAG sensitivity. We show that NHERF proteins are direct regulators of ion channel activity and that DAG sensitivity is a distinctive hallmark of TRPC channels.

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Abstract 39: The Transient Receptor-Potential Channel 6 Regulates Thromboxane A2 Receptor-Operated Calcium Entry and Plays a Critical Role in Platelet Function

Arteriosclerosis Thrombosis and Vascular Biology ◽

10.1161/atvb.33.suppl_1.a39 ◽

2013 ◽

Vol 33 (suppl_1) ◽

Author(s):

Fadi T Khasawneh ◽

Enma V Espinosa ◽

Olivia A Lin ◽

John P Murad

Keyword(s):

Platelet Aggregation ◽

Platelet Function ◽

Transient Receptor Potential ◽

Critical Role ◽

Receptor Potential ◽

Underlying Mechanism ◽

Calcium Entry ◽

Transient Receptor Potential Channels ◽

Dependent Manner ◽

Transient Receptor

Although changes in the intracellular levels of calcium is a central step in platelet activation, the underlying mechanism of changes that are dependent on receptor-operated calcium entry (ROCE) remains to be defined. Furthermore, it was proposed, though never proven, that the Transient Receptor-Potential Channels (TRPCs) may play a key role in this process. In this connection, we have previously shown that TRPC6 plays a vital role in physiological hemostasis and thrombogenesis. However, the underlying mechanism by which TRPC6 modulates these processes also remains to be determined. Based on these considerations, we hypothesized that TRPC6 plays an essential role in ROCE and hence platelet function. Using a (genetic) TRPC6 knockout (KO) mouse approach, our preliminary studies revealed that these platelets exhibited a defect in platelet aggregation induced by the thromboxane A 2 receptor (TPR) agonist U46619. Conversely, the aggregation response triggered by ADP or the thrombin receptor activating peptide 4 (TRAP4) was found to be comparable to that of the wild-type (WT) platelets. Separate studies revealed that the TRPC6 deficient platelets were also found to exhibit a defect in TPR-mediated dense granule (ATP) secretion, whereas that of ADP and TRAP4 was normal. Moreover, we observed a defect in integrin GP IIb-IIIa activation that was again specific to TPR (normal activation in response to ADP and TRAP4), suggesting a defect in inside-out signaling. Finally, TPR-dependent CE was also found to be deficient in the TRPC6 KO platelets, unlike that stimulated by ADP or TRAP4. Future studies will further investigate the molecular mechanism of TRPC6-regulated platelet function and CE. Taken together, these findings demonstrate for the first time that TRPC6 regulates CE in a TPR-dependent manner and that this regulation consequently modulates platelet aggregation, secretion, as well as GP IIb-IIIa activation. These studies may define a new therapeutic target for managing multiple thrombosis-based disorders.

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