scholarly journals Major contribution of the 3/6/7 class of TRPC channels to myocardial ischemia/reperfusion and cellular hypoxia/reoxygenation injuries

2017 ◽  
Vol 114 (23) ◽  
pp. E4582-E4591 ◽  
Author(s):  
Xiju He ◽  
Shoutian Li ◽  
Benju Liu ◽  
Sebastian Susperreguy ◽  
Karina Formoso ◽  
...  
Keyword(s):  
Calcium Release ◽  
Transient Receptor Potential ◽  
Mitochondrial Permeability Transition ◽  
Ischemia Reperfusion ◽  
Receptor Potential ◽  
Permeability Transition ◽  
Transient Receptor Potential Channels ◽  
Trpc Channels ◽  
Potential Channels

The injury phase after myocardial infarcts occurs during reperfusion and is a consequence of calcium release from internal stores combined with calcium entry, leading to cell death by apoptopic and necrotic processes. The mechanism(s) by which calcium enters cells has(ve) not been identified. Here, we identify canonical transient receptor potential channels (TRPC) 3 and 6 as the cation channels through which most of the damaging calcium enters cells to trigger their death, and we describe mechanisms activated during the injury phase. Working in vitro with H9c2 cardiomyoblasts subjected to 9-h hypoxia followed by 6-h reoxygenation (H/R), and analyzing changes occurring in areas-at-risk (AARs) of murine hearts subjected to a 30-min ischemia followed by 24-h reperfusion (I/R) protocol, we found: (i) that blocking TRPC with SKF96365 significantly ameliorated damage induced by H/R, including development of the mitochondrial permeability transition and proapoptotic changes in Bcl2/BAX ratios; and (ii) that AAR tissues had increased TUNEL+ cells, augmented Bcl2/BAX ratios, and increased p(S240)NFATc3, p(S473)AKT, p(S9)GSK3β, and TRPC3 and -6 proteins, consistent with activation of a positive-feedback loop in which calcium entering through TRPCs activates calcineurin-mediated NFATc3-directed transcription of TRPC genes, leading to more Ca2+ entry. All these changes were markedly reduced in mice lacking TRPC3, -6, and -7. The changes caused by I/R in AAR tissues were matched by those seen after H/R in cardiomyoblasts in all aspects except for p-AKT and p-GSK3β, which were decreased after H/R in cardiomyoblasts instead of increased. TRPC should be promising targets for pharmacologic intervention after cardiac infarcts.

scholarly journals Genetic variants affecting human TRPA1 or TRPM8 structure can be classified in vitro as ‘well expressed’, ‘poorly expressed’ or ‘salvageable’

2015 ◽  
Vol 35 (5) ◽  
Author(s):  
Kevin Morgan ◽  
Laura Rachel Sadofsky ◽  
Alyn Hugh Morice
Keyword(s):  
Genetic Variants ◽  
Transient Receptor Potential ◽  
Kinase Inhibitor ◽  
Receptor Potential ◽  
Transient Receptor Potential Channels ◽  
Human Embryonic Kidney ◽  
Potential Channels ◽  
Pre Treatment ◽  
Transient Receptor

Genetic variants of human transient receptor potential channels A1 and M8 expressed in human embryonic kidney HEK293 and SH-SY5Y cells were assayed using Ca2+ signalling. TRPA1 Y69C responded well. Poorly expressed variant signalling was enhanced by pre-treatment with tyrosine kinase inhibitor PP2 or Zn2+.

scholarly journals The Membrane Proximal Domain of TRPV1 and TRPV2 Channels Mediates Protein–Protein Interactions and Lipid Binding In Vitro

2019 ◽  
Vol 20 (3) ◽  
pp. 682 ◽  
Author(s):  
Pau Doñate-Macián ◽  
Elena Álvarez-Marimon ◽  
Francesc Sepulcre ◽  
José Vázquez-Ibar ◽  
Alex Perálvarez-Marín
Keyword(s):  
Protein Interactions ◽  
Transient Receptor Potential ◽  
Receptor Potential ◽  
Lipid Binding ◽  
Transient Receptor Potential Channels ◽  
Trpv Channels ◽  
Potential Channels ◽  
Transient Receptor ◽  
Lipid Protein Interactions

Constitutive or regulated membrane protein trafficking is a key cell biology process. Transient receptor potential channels are somatosensory proteins in charge of detecting several physical and chemical stimuli, thus requiring fine vesicular trafficking. The membrane proximal or pre-S1 domain (MPD) is a highly conserved domain in transient receptor potential channels from the vanilloid (TRPV) subfamily. MPD shows traits corresponding to protein-protein and lipid-protein interactions, and protein regulatory regions. We have expressed MPD of TRPV1 and TRPV2 as green fluorescente protein (GFP)-fusion proteins to perform an in vitro biochemical and biophysical characterization. Pull-down experiments indicate that MPD recognizes and binds Soluble N-ethylmaleimide-sensitive factor Attachment Protein Receptors (SNARE). Synchrotron radiation scattering experiments show that this domain does not self-oligomerize. MPD interacts with phosphatidic acid (PA), a metabolite of the phospholipase D (PLD) pathway, in a specific manner as shown by lipid strips and Trp fluorescence quenching experiments. We show for the first time, to the best of our knowledge, the binding to PA of an N-terminus domain in TRPV channels. The presence of a PA binding domain in TRPV channels argues for putative PLD regulation. Findings in this study open new perspectives to understand the regulated and constitutive trafficking of TRPV channels exerted by protein-protein and lipid-protein interactions.

scholarly journals FeRIC-based magnetogenetics: evaluation of methods and protocols in in vitro models

2020 ◽  
Author(s):  
Miriam Hernández-Morales ◽  
Victor Han ◽  
Richard H Kramer ◽  
Chunlei Liu
Keyword(s):  
Transient Receptor Potential ◽  
Receptor Potential ◽  
Functional Expression ◽  
In Vitro Models ◽  
Transient Receptor Potential Channels ◽  
Iron Level ◽  
Ferritin Iron ◽  
Potential Channels ◽  
Transient Receptor

AbstractFeRIC (Ferritin iron Redistribution to Ion Channels) is a magnetogenetic technique that uses radiofrequency (RF) waves to activate the transient receptor potential channels, such as TRPV1 and TRPV4, coupled to cellular ferritins. In cells expressing ferritin-tagged TRPV, RF stimulation increases the cytosolic Ca2+ levels via a biochemical pathway. The interaction between RF and ferritin increases the free cytosolic iron level that in turn, triggers chemical reactions producing reactive oxygen species and oxidized lipids that activate the ferritin-tagged TRPV. In this pathway, it is expected that experimental factors that disturb the ferritin expression, the ferritin iron load, the TRPV functional expression, or the cellular redox state will impact the RF efficacy to activate ferritin-tagged TRPV. Here, three in vitro protocols were compared for using FeRIC to remotely activate ferritin-tagged TRPV. Further, several experimental factors were examined that either enhance or abolish the RF control of ferritin-tagged TRPV. The findings may help establish reproducible magnetogenetic experimental protocols.

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.

scholarly journals Does Erythropoietin Regulate TRPC Channels in Red Blood Cells?

2017 ◽  
Vol 41 (3) ◽  
pp. 1219-1228 ◽  
Author(s):  
Jens Danielczok ◽  
Laura Hertz ◽  
Sandra Ruppenthal ◽  
Elisabeth Kaiser ◽  
Polina Petkova-Kirova ◽  
...  
Keyword(s):  
Red Blood Cells ◽  
Blood Cells ◽  
Transient Receptor Potential ◽  
Ion Homeostasis ◽  
Receptor Potential ◽  
Transient Receptor Potential Channels ◽  
Erythroid Cell ◽  
Trpc Channels ◽  
Potential Channels ◽  
Human Rbcs

Background: Cation channels play an essential role in red blood cells (RBCs) ion homeostasis. One set of ion channels are the transient receptor potential channels of canonical type (TRPC channels). The abundance of these channels in primary erythroblasts, erythroid cell lines and RBCs was associated with an increase in intracellular Ca2+ upon stimulation with Erythropoietin (Epo). In contrast two independent studies on Epo-treated patients revealed diminished basal Ca2+ concentration or reduced phosphatidylserine exposure to the outer membrane leaflet. Methods: To resolve the seemingly conflicting reports we challenged mature human and mouse RBCs of several genotypes with Epo and Prostaglandin E2 (PGE2) and recorded the intracellular Ca2+ content. Next Generation Sequencing was utilised to approach a molecular analysis of reticulocytes. Results/Conclusions: Our results allow concluding that Epo and PGE2 regulation of the Ca2+ homeostasis is distinctly different between murine and human RBCs and that changes in intracellular Ca2+ upon Epo treatment is a primary rather than a compensatory effect. In human RBCs, Epo itself has no effect on Ca2+ fluxes but inhibits the PGE2-induced Ca2+ entry. In murine mature RBCs functional evidence indicates TRPC4/C5 mediated Ca2+ entry activated by Epo whereas PGE2 leads to a TRPC independent Ca2+ entry.

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