scholarly journals Tyrosine phosphorylation–dependent activation of TRPC6 regulated by PLC-γ1 and nephrin: effect of mutations associated with focal segmental glomerulosclerosis

2011 ◽  
Vol 22 (11) ◽  
pp. 1824-1835 ◽  
Author(s):  
Shoichiro Kanda ◽  
Yutaka Harita ◽  
Yoshio Shibagaki ◽  
Takashi Sekine ◽  
Takashi Igarashi ◽  
...  
Keyword(s):  
Complex Formation ◽  
Tyrosine Phosphorylation ◽  
Focal Segmental Glomerulosclerosis ◽  
Transient Receptor Potential ◽  
Receptor Potential ◽  
Surface Expression ◽  
Activation Mechanism ◽  
Channel Activity ◽  
Segmental Glomerulosclerosis ◽  
Specific Regulation

Transient receptor potential canonicals (TRPCs) play important roles in the regulation of intracellular calcium concentration. Mutations in the TRPC6 gene are found in patients with focal segmental glomerulosclerosis (FSGS), a proteinuric disease characterized by dysregulated function of renal glomerular epithelial cells (podocytes). There is as yet no clear picture for the activation mechanism of TRPC6 at the molecular basis, however, and the association between its channel activity and pathogenesis remains unclear. We demonstrate here that tyrosine phosphorylation of TRPC6 induces a complex formation with phospholipase C (PLC)-γ1, which is prerequisite for TRPC6 surface expression. Furthermore, nephrin, an adhesion protein between the foot processes of podocytes, binds to phosphorylated TRPC6 via its cytoplasmic domain, competitively inhibiting TRPC6–PLC-γ1 complex formation, TRPC6 surface localization, and TRPC6 activation. Importantly, FSGS-associated mutations render the mutated TRPC6s insensitive to nephrin suppression, thereby promoting their surface expression and channel activation. These results delineate the mechanism of TRPC6 activation regulated by tyrosine phosphorylation, and imply the cell type–specific regulation, which correlates the FSGS mutations with deregulated TRPC6 channel activity.

scholarly journals TRPC6 mutations associated with focal segmental glomerulosclerosis cause constitutive activation of NFAT-dependent transcription

2009 ◽  
Vol 296 (3) ◽  
pp. C558-C569 ◽  
Author(s):  
Johannes Schlöndorff ◽  
Donato del Camino ◽  
Robert Carrasquillo ◽  
Vanessa Lacey ◽  
Martin R. Pollak
Keyword(s):  
Focal Segmental Glomerulosclerosis ◽  
Transient Receptor Potential ◽  
Receptor Potential ◽  
Human Kidney ◽  
Transient Receptor Potential Channel ◽  
Surface Expression ◽  
Channel Activity ◽  
Dominant Form ◽  
Segmental Glomerulosclerosis ◽  
Nfat Activation

Mutations in the canonical transient receptor potential channel TRPC6 lead to an autosomal dominant form of human kidney disease characterized histologically by focal and segmental glomerulosclerosis. Several of these mutations enhance the amplitude and duration of the channel current. However, the effect of these mutations on the downstream target of TRPC6, the nuclear factor of activated T cell (NFAT) transcription factors, has not been previously examined. Here we demonstrate that all three TRPC6 mutations previously shown to enhance channel activity lead to enhanced basal NFAT-mediated transcription in several cell lines, including cultured podocytes. These effects are dependent on channel activity and are dominant when mutants are coexpressed with wild-type TRPC6. While TRPC6 mutants do not demonstrate an increase in basal channel currents, a subset of cells expressing the R895C and E897K mutants have elevated basal calcium levels as measured by Fura-2 imaging. Activation of NFAT by TRPC6 mutants is blocked by inhibitors of calcineurin, calmodulin-dependent kinase II, and phosphatidylinositol 3-kinase. PP2 partially inhibits NFAT activation by mutant TRPC6 independently of Src, Yes, or Fyn. Differences in channel glycosylation and surface expression do not explain the ability of mutants to enhance NFAT activation. Taken together, these results identify the activation of the calcineurin-NFAT pathway as a potential mediator of focal segmental glomerulosclerosis.

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

2016 ◽  
Vol 114 (1) ◽  
pp. E37-E46 ◽  
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.

A cell-based impedance assay for monitoring transient receptor potential (TRP) ion channel activity

2011 ◽  
Vol 26 (5) ◽  
pp. 2376-2382 ◽  
Author(s):  
Oliver Pänke ◽  
Winnie Weigel ◽  
Sabine Schmidt ◽  
Anja Steude ◽  
Andrea A. Robitzki
Keyword(s):  
Ion Channel ◽  
Transient Receptor Potential ◽  
Receptor Potential ◽  
Channel Activity ◽  
A Cell ◽  
Transient Receptor ◽  
Trp Ion Channel

scholarly journals TRPM8 in prostate cancer cells: a potential diagnostic and prognostic marker with a secretory function?

2006 ◽  
Vol 13 (1) ◽  
pp. 27-38 ◽  
Author(s):  
Lei Zhang ◽  
Gregory J Barritt
Keyword(s):  
Prostate Cancer ◽  
Cancer Cells ◽  
Prognostic Marker ◽  
Cell Biology ◽  
Transient Receptor Potential ◽  
Sensory Nerve ◽  
Receptor Potential ◽  
Activation Mechanism ◽  
Prostate Cancer Cells ◽  
Genetic Interventions

During the past 5 years it has emerged that the transient receptor potential (TRP) family of Ca2+-and Na+-permeable channels plays a diverse and important role in cell biology and in pathology. One member of this family, TRPM8, is highly expressed in prostate cancer cells but the physiological and pathological functions of TRPM8 in these cells are not known. Here we address these questions, and the issue of whether or not TRPM8 is an effective diagnostic and prognostic marker in prostate cancer. TRPM8 is known to be activated by cool stimuli (17–25 °C) and cooling compounds such as menthol. The activation mechanism(s) involves voltage sensing of membrane potential, phosphatidylinositol 4,5-bisphosphate and Ca2+. In addition to prostate cancer cells, TRPM8 is expressed in sensory neurons where it acts as a sensor of cold. In prostate epithelial cells, expression of TRPM8 is regulated by androgen and is elevated in androgen-sensitive cancerous cells compared with normal cells. While there is some evidence that in prostate cancer cells Ca2+ and Na+ inflow through TRPM8 is necessary for survival and function, including secretion at the apical membrane, the function of TRPM8 in these cells is not really known. It may well differ from the role of TRPM8 as a cool sensor in sensory nerve cells. Androgen unresponsive prostate cancer is difficult to treat effectively and there are limited diagnostic and prognostic markers available. TRPM8 is a potential tissue marker in differential diagnosis and a potential prognostic marker for androgen-unresponsive and metastatic prostate cancer. As a consequence of its ability to convey Ca2+ and Na+ and its expression in only a limited number of cell types, TRPM8 is considered to be a promising target for pharmaceutical, immunological and genetic interventions for the treatment of prostate cancer.

scholarly journals Transient receptor potential melastatin 2 channels are overexpressed in myalgic encephalomyelitis/chronic fatigue syndrome patients

2019 ◽  
Vol 17 (1) ◽  
Author(s):  
Cassandra Balinas ◽  
Helene Cabanas ◽  
Donald Staines ◽  
Sonya Marshall-Gradisnik
Keyword(s):  
Nk Cells ◽  
Transient Receptor Potential ◽  
Nk Cell ◽  
Receptor Potential ◽  
Surface Expression ◽  
Chronic Fatigue ◽  
Cell Cytotoxicity ◽  
Fatigue Syndrome ◽  
Transient Receptor Potential Melastatin ◽  
Nk Cell Cytotoxicity

Abstract Background Myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) is hallmarked by a significant reduction in natural killer (NK) cell cytotoxicity, a mechanism tightly regulated by calcium (Ca2+). Interestingly, interleukin-2 (IL-2) increases NK cell cytotoxicity. Transient receptor potential melastatin 2 (TRPM2) ion channels are fundamental for Ca2+ signalling in NK cells. This pilot investigation aimed to characterise TRPM2 and CD38 surface expression in vitro on NK cells in ME/CFS patients. This investigation furthermore examined the pharmaceutical effect of 8-bromoadenosine phosphoribose (8-Br-ADPR) and N6-Benzoyladenosine-3′,5′-cyclic monophosphate (N6-Bnz-cAMP) on TRPM2 and CD38 surface expression and NK cell cytotoxicity between ME/CFS and healthy control (HC) participants. Methods Ten ME/CFS patients (43.45 ± 12.36) and 10 HCs (43 ± 12.27) were age and sex-matched. Isolated NK cells were labelled with fluorescent antibodies to determine baseline and drug-treated TRPM2 and CD38 surface expression on NK cell subsets. Following IL-2 stimulation, NK cell cytotoxicity was measured following 8-Br-ADPR and N6-Bnz-cAMP drug treatments by flow cytometry. Results Baseline TRPM2 and CD38 surface expression was significantly higher on NK cell subsets in ME/CFS patients compared with HCs. Post IL-2 stimulation, TRPM2 and CD38 surface expression solely decreased on the CD56DimCD16+ subset. 8-Br-ADPR treatment significantly reduced TRPM2 surface expression on the CD56BrightCD16Dim/− subset within the ME/CFS group. Baseline cell cytotoxicity was significantly reduced in ME/CFS patients, however no changes were observed post drug treatment in either group. Conclusion Overexpression of TRPM2 on NK cells may function as a compensatory mechanism to alert a dysregulation in Ca2+ homeostasis to enhance NK cell function in ME/CFS, such as NK cell cytotoxicity. As no improvement in NK cell cytotoxicity was observed within the ME/CFS group, an impairment in the TRPM2 ion channel may be present in ME/CFS patients, resulting in alterations in [Ca2+]i mobilisation and influx, which is fundamental in driving NK cell cytotoxicity. Differential expression of TRPM2 between NK cell subtypes may provide evidence for their role in the pathomechanism involving NK cell cytotoxicity activity in ME/CFS.

scholarly journals Upregulation of Transient Receptor Potential Vanilloid Type-1 Channel Activity and Ca2+Influx Dysfunction in Human Pterygial Cells

2016 ◽  
Vol 57 (6) ◽  
pp. 2564 ◽  
Author(s):  
Fabian Garreis ◽  
Antje Schröder ◽  
Peter S. Reinach ◽  
Stefanie Zoll ◽  
Noushafarin Khajavi ◽  
...  
Keyword(s):  
Transient Receptor Potential ◽  
Receptor Potential ◽  
Transient Receptor Potential Vanilloid ◽  
Channel Activity ◽  
Transient Receptor

scholarly journals Cellular cholesterol controls TRPC3 function: evidence from a novel dominant-negative knockdown strategy

2006 ◽  
Vol 396 (1) ◽  
pp. 147-155 ◽  
Author(s):  
Annarita Graziani ◽  
Christian Rosker ◽  
Sepp D. Kohlwein ◽  
Michael X. Zhu ◽  
Christoph Romanin ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Transient Receptor Potential ◽  
Lipid Membrane ◽  
Membrane Conductance ◽  
Receptor Potential ◽  
Surface Expression ◽  
Dominant Negative ◽  
Membrane Microdomains ◽  
Membrane Expression ◽  
Hek 293

TRPC3 (canonical transient receptor potential protein 3) has been suggested to be a component of cation channel complexes that are targeted to cholesterol-rich lipid membrane microdomains. In the present study, we investigated the potential role of membrane cholesterol as a regulator of cellular TRPC3 conductances. Functional experiments demonstrated that cholesterol loading activates a non-selective cation conductance and a Ca2+ entry pathway in TRPC3-overexpressing cells but not in wild-type HEK-293 (human embryonic kidney 293) cells. The cholesterol-induced membrane conductance exhibited a current-to-voltage relationship similar to that observed upon PLC (phospholipase C)-dependent activation of TRPC3 channels. Nonetheless, the cholesterol-activated conductance lacked negative modulation by extracellular Ca2+, a typical feature of agonist-activated TRPC3 currents. Involvement of TRPC3 in the cholesterol-dependent membrane conductance was further corroborated by a novel dominant-negative strategy for selective blockade of TRPC3 channel activity. Expression of a TRPC3 mutant, which contained a haemagglutinin epitope tag in the second extracellular loop, conferred antibody sensitivity to both the classical PLC-activated as well as the cholesterol-activated conductance in TRPC3-expressing cells. Moreover, cholesterol loading as well as PLC stimulation was found to increase surface expression of TRPC3. Promotion of TRPC3 membrane expression by cholesterol was persistent over 30 min, while PLC-mediated enhancement of plasma membrane expression of TRPC3 was transient in nature. We suggest the cholesterol content of the plasma membrane as a determinant of cellular TRPC3 activity and provide evidence for cholesterol dependence of TRPC3 surface expression.

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