scholarly journals Structural insights into TRPM8 inhibition and desensitization

Science ◽  
2019 ◽  
Vol 365 (6460) ◽  
pp. 1434-1440 ◽  
Author(s):  
Melinda M. Diver ◽  
Yifan Cheng ◽  
David Julius
Keyword(s):  
Conformational Changes ◽  
Calcium Binding ◽  
Transient Receptor Potential ◽  
Trp Channels ◽  
Receptor Potential ◽  
Binding Pocket ◽  
Chemical Structures ◽  
Transient Receptor Potential Melastatin ◽  
Transient Receptor ◽  
Large Rearrangements

The transient receptor potential melastatin 8 (TRPM8) ion channel is the primary detector of environmental cold and an important target for treating pathological cold hypersensitivity. Here, we present cryo–electron microscopy structures of TRPM8 in ligand-free, antagonist-bound, or calcium-bound forms, revealing how robust conformational changes give rise to two nonconducting states, closed and desensitized. We describe a malleable ligand-binding pocket that accommodates drugs of diverse chemical structures, and we delineate the ion permeation pathway, including the contribution of lipids to pore architecture. Furthermore, we show that direct calcium binding mediates stimulus-evoked desensitization, clarifying this important mechanism of sensory adaptation. We observe large rearrangements within the S4-S5 linker that reposition the S1-S4 and pore domains relative to the TRP helix, leading us to propose a distinct model for modulation of TRPM8 and possibly other TRP channels.

scholarly journals Structure of the human TRPM4 ion channel in a lipid nanodisc

Science ◽  
2017 ◽  
Vol 359 (6372) ◽  
pp. 228-232 ◽  
Author(s):  
Henriette E. Autzen ◽  
Alexander G. Myasnikov ◽  
Melody G. Campbell ◽  
Daniel Asarnow ◽  
David Julius ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Calcium Binding ◽  
Transient Receptor Potential ◽  
Trp Channels ◽  
Receptor Potential ◽  
Dependent Manner ◽  
Calcium Binding Site ◽  
Voltage Dependent ◽  
Transient Receptor ◽  
Lipid Nanodiscs

Transient receptor potential (TRP) melastatin 4 (TRPM4) is a widely expressed cation channel associated with a variety of cardiovascular disorders. TRPM4 is activated by increased intracellular calcium in a voltage-dependent manner but, unlike many other TRP channels, is permeable to monovalent cations only. Here we present two structures of full-length human TRPM4 embedded in lipid nanodiscs at ~3-angstrom resolution, as determined by single-particle cryo–electron microscopy. These structures, with and without calcium bound, reveal a general architecture for this major subfamily of TRP channels and a well-defined calcium-binding site within the intracellular side of the S1-S4 domain. The structures correspond to two distinct closed states. Calcium binding induces conformational changes that likely prime the channel for voltage-dependent opening.

scholarly journals Mapping of CaM, S100A1 and PIP2-Binding Epitopes in the Intracellular N- and C-Termini of TRPM4

2020 ◽  
Vol 21 (12) ◽  
pp. 4323
Author(s):  
Kristyna Bousova ◽  
Ivan Barvik ◽  
Petr Herman ◽  
Kateřina Hofbauerová ◽  
Lenka Monincova ◽  
...  
Keyword(s):  
Calcium Binding ◽  
Transient Receptor Potential ◽  
Trp Channels ◽  
Receptor Potential ◽  
Md Simulations ◽  
Protein Docking ◽  
Alanine Scanning Mutagenesis ◽  
Rational Drug ◽  
Transient Receptor ◽  
Endogenous Modulators

Molecular determinants of the binding of various endogenous modulators to transient receptor potential (TRP) channels are crucial for the understanding of necessary cellular pathways, as well as new paths for rational drug designs. The aim of this study was to characterise interactions between the TRP cation channel subfamily melastatin member 4 (TRPM4) and endogenous intracellular modulators—calcium-binding proteins (calmodulin (CaM) and S100A1) and phosphatidylinositol 4, 5-bisphosphate (PIP2). We have found binding epitopes at the N- and C-termini of TRPM4 shared by CaM, S100A1 and PIP2. The binding affinities of short peptides representing the binding epitopes of N- and C-termini were measured by means of fluorescence anisotropy (FA). The importance of representative basic amino acids and their combinations from both peptides for the binding of endogenous TRPM4 modulators was proved using point alanine-scanning mutagenesis. In silico protein–protein docking of both peptides to CaM and S100A1 and extensive molecular dynamics (MD) simulations enabled the description of key stabilising interactions at the atomic level. Recently solved cryo-Electron Microscopy (EM) structures made it possible to put our findings into the context of the entire TRPM4 channel and to deduce how the binding of these endogenous modulators could allosterically affect the gating of TRPM4. Moreover, both identified binding epitopes seem to be ideally positioned to mediate the involvement of TRPM4 in higher-order hetero-multimeric complexes with important physiological functions.

scholarly journals Structure of full-length human TRPM4

2018 ◽  
Vol 115 (10) ◽  
pp. 2377-2382 ◽  
Author(s):  
Jingjing Duan ◽  
Zongli Li ◽  
Jian Li ◽  
Ana Santa-Cruz ◽  
Silvia Sanchez-Martinez ◽  
...  
Keyword(s):  
Transient Receptor Potential ◽  
Trp Channels ◽  
Coiled Coil ◽  
Receptor Potential ◽  
Lipid Binding ◽  
Full Length ◽  
Intramolecular Interactions ◽  
Coiled Coil Domain ◽  
Transient Receptor Potential Melastatin ◽  
Transient Receptor

Transient receptor potential melastatin subfamily member 4 (TRPM4) is a widely distributed, calcium-activated, monovalent-selective cation channel. Mutations in human TRPM4 (hTRPM4) result in progressive familial heart block. Here, we report the electron cryomicroscopy structure of hTRPM4 in a closed, Na+-bound, apo state at pH 7.5 to an overall resolution of 3.7 Å. Five partially hydrated sodium ions are proposed to occupy the center of the conduction pore and the entrance to the coiled-coil domain. We identify an upper gate in the selectivity filter and a lower gate at the entrance to the cytoplasmic coiled-coil domain. Intramolecular interactions exist between the TRP domain and the S4–S5 linker, N-terminal domain, and N and C termini. Finally, we identify aromatic interactions via π–π bonds and cation–π bonds, glycosylation at an N-linked extracellular site, a pore-loop disulfide bond, and 24 lipid binding sites. We compare and contrast this structure with other TRP channels and discuss potential mechanisms of regulation and gating of human full-length TRPM4.

scholarly journals TRPM6 N-Terminal CaM- and S100A1-Binding Domains

2019 ◽  
Vol 20 (18) ◽  
pp. 4430 ◽  
Author(s):  
Monika Zouharova ◽  
Petr Herman ◽  
Kateřina Hofbauerová ◽  
Jiri Vondrasek ◽  
Kristyna Bousova
Keyword(s):  
Calcium Binding ◽  
Transient Receptor Potential ◽  
Calcium Signalling ◽  
Receptor Potential ◽  
Basic Amino Acid ◽  
Binding Domain ◽  
Specific Domain ◽  
Binding Domains ◽  
Transient Receptor Potential Melastatin ◽  
Transient Receptor

Transient receptor potential (TRPs) channels are crucial downstream targets of calcium signalling cascades. They can be modulated either by calcium itself and/or by calcium-binding proteins (CBPs). Intracellular messengers usually interact with binding domains present at the most variable TRP regions—N- and C-cytoplasmic termini. Calmodulin (CaM) is a calcium-dependent cytosolic protein serving as a modulator of most transmembrane receptors. Although CaM-binding domains are widespread within intracellular parts of TRPs, no such binding domain has been characterised at the TRP melastatin member—the transient receptor potential melastatin 6 (TRPM6) channel. Another CBP, the S100 calcium-binding protein A1 (S100A1), is also known for its modulatory activities towards receptors. S100A1 commonly shares a CaM-binding domain. Here, we present the first identified CaM and S100A1 binding sites at the N-terminal of TRPM6. We have confirmed the L520-R535 N-terminal TRPM6 domain as a shared binding site for CaM and S100A1 using biophysical and molecular modelling methods. A specific domain of basic amino acid residues (R526/R531/K532/R535) present at this TRPM6 domain has been identified as crucial to maintain non-covalent interactions with the ligands. Our data unambiguously confirm that CaM and S100A1 share the same binding domain at the TRPM6 N-terminus although the ligand-binding mechanism is different.

scholarly journals Structure–function analyses of the ion channel TRPC3 reveal that its cytoplasmic domain allosterically modulates channel gating

2018 ◽  
Vol 293 (41) ◽  
pp. 16102-16114 ◽  
Author(s):  
Francisco Sierra-Valdez ◽  
Caleigh M. Azumaya ◽  
Luis O. Romero ◽  
Terunaga Nakagawa ◽  
Julio F. Cordero-Morales
Keyword(s):  
Conformational Changes ◽  
Transient Receptor Potential ◽  
Transmembrane Domain ◽  
Trp Channels ◽  
Channel Gating ◽  
Memory Loss ◽  
Receptor Potential ◽  
Cytoplasmic Domain ◽  
Terminal Loop ◽  
Transient Receptor

The transient receptor potential ion channels support Ca2+ permeation in many organs, including the heart, brain, and kidney. Genetic mutations in transient receptor potential cation channel subfamily C member 3 (TRPC3) are associated with neurodegenerative diseases, memory loss, and hypertension. To better understand the conformational changes that regulate TRPC3 function, we solved the cryo-EM structures for the full-length human TRPC3 and its cytoplasmic domain (CPD) in the apo state at 5.8- and 4.0-Å resolution, respectively. These structures revealed that the TRPC3 transmembrane domain resembles those of other TRP channels and that the CPD is a stable module involved in channel assembly and gating. We observed the presence of a C-terminal domain swap at the center of the CPD where horizontal helices (HHs) transition into a coiled-coil bundle. Comparison of TRPC3 structures revealed that the HHs can reside in two distinct positions. Electrophysiological analyses disclosed that shortening the length of the C-terminal loop connecting the HH with the TRP helices increases TRPC3 activity and that elongating the length of the loop has the opposite effect. Our findings indicate that the C-terminal loop affects channel gating by altering the allosteric coupling between the cytoplasmic and transmembrane domains. We propose that molecules that target the HH may represent a promising strategy for controlling TRPC3-associated neurological disorders and hypertension.

scholarly journals Drosophila menthol sensitivity and the Precambrian origins of transient receptor potential-dependent chemosensation

2019 ◽  
Vol 374 (1785) ◽  
pp. 20190369 ◽  
Author(s):  
Nathaniel J. Himmel ◽  
Jamin M. Letcher ◽  
Akira Sakurai ◽  
Thomas R. Gray ◽  
Maggie N. Benson ◽  
...  
Keyword(s):  
Drosophila Melanogaster ◽  
Transient Receptor Potential ◽  
Trp Channels ◽  
Receptor Potential ◽  
Cation Channels ◽  
Transient Receptor Potential Melastatin ◽  
Growing Body ◽  
Sensitivity Studies ◽  
Transient Receptor ◽  
Class Iv

Transient receptor potential (TRP) cation channels are highly conserved, polymodal sensors which respond to a wide variety of stimuli. Perhaps most notably, TRP channels serve critical functions in nociception and pain. A growing body of evidence suggests that transient receptor potential melastatin (TRPM) and transient receptor potential ankyrin (TRPA) thermal and electrophile sensitivities predate the protostome–deuterostome split (greater than 550 Ma). However, TRPM and TRPA channels are also thought to detect modified terpenes (e.g. menthol). Although terpenoids like menthol are thought to be aversive and/or harmful to insects, mechanistic sensitivity studies have been largely restricted to chordates. Furthermore, it is unknown if TRP-menthol sensing is as ancient as thermal and/or electrophile sensitivity. Combining genetic, optical, electrophysiological, behavioural and phylogenetic approaches, we tested the hypothesis that insect TRP channels play a conserved role in menthol sensing. We found that topical application of menthol to Drosophila melanogaster larvae elicits a Trpm - and TrpA1 -dependent nocifensive rolling behaviour, which requires activation of Class IV nociceptor neurons. Further, in characterizing the evolution of TRP channels, we put forth the hypotheses that three previously undescribed TRPM channel clades (basal, αTRPM and βTRPM), as well as TRPs with residues critical for menthol sensing, were present in ancestral bilaterians. This article is part of the Theo Murphy meeting issue ‘Evolution of mechanisms and behaviour important for pain’.

scholarly journals Transient receptor potential melastatin 3 is a phosphoinositide-dependent ion channel

2015 ◽  
Vol 146 (1) ◽  
pp. 65-77 ◽  
Author(s):  
Doreen Badheka ◽  
Istvan Borbiro ◽  
Tibor Rohacs
Keyword(s):  
Ion Channel ◽  
Transient Receptor Potential ◽  
Kinase Inhibitors ◽  
Trp Channels ◽  
Receptor Potential ◽  
Channel Activity ◽  
Intact Cells ◽  
Transient Receptor Potential Melastatin ◽  
Transient Receptor ◽  
Phosphatidylinositol 4

Phosphoinositides are emerging as general regulators of the functionally diverse transient receptor potential (TRP) ion channel family. Phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2) has been reported to positively regulate many TRP channels, but in several cases phosphoinositide regulation is controversial. TRP melastatin 3 (TRPM3) is a heat-activated ion channel that is also stimulated by chemical agonists, such as pregnenolone sulfate. Here, we used a wide array of approaches to determine the effects of phosphoinositides on TRPM3. We found that channel activity in excised inside-out patches decreased over time (rundown), an attribute of PI(4,5)P2-dependent ion channels. Channel activity could be restored by application of either synthetic dioctanoyl (diC8) or natural arachidonyl stearyl (AASt) PI(4,5)P2. The PI(4,5)P2 precursor phosphatidylinositol 4-phosphate (PI(4)P) was less effective at restoring channel activity. TRPM3 currents were also restored by MgATP, an effect which was inhibited by two different phosphatidylinositol 4-kinase inhibitors, or by pretreatment with a phosphatidylinositol-specific phospholipase C (PI-PLC) enzyme, indicating that MgATP acted by generating phosphoinositides. In intact cells, reduction of PI(4,5)P2 levels by chemically inducible phosphoinositide phosphatases or a voltage-sensitive 5′-phosphatase inhibited channel activity. Activation of PLC via muscarinic receptors also inhibited TRPM3 channel activity. Overall, our data indicate that TRPM3 is a phosphoinositide-dependent ion channel and that decreasing PI(4,5)P2 abundance limits its activity. As all other members of the TRPM family have also been shown to require PI(4,5)P2 for activity, our data establish PI(4,5)P2 as a general positive cofactor of this ion channel subfamily.

scholarly journals Mechanism of praziquantel action at a parasitic flatworm ion channel

2021 ◽  
Author(s):  
Sang-Kyu Park ◽  
Lukas Friedrich ◽  
Nawal A. Yahya ◽  
Claudia Rohr ◽  
Evgeny G. Chulkov ◽  
...  
Keyword(s):  
Ion Channel ◽  
Transient Receptor Potential ◽  
Essential Medicine ◽  
Receptor Potential ◽  
Binding Pocket ◽  
Single Amino Acid ◽  
Transient Receptor Potential Melastatin ◽  
Single Amino Acid Change ◽  
Parasitic Flatworm ◽  
Transient Receptor

Praziquantel (PZQ) is an essential medicine for treating parasitic flatworm infections such as schistosomiasis, which afflicts over 250 million people. However, PZQ is not universally effective, lacking activity against the liver fluke Fasciola. The reason for this insensitivity is unclear, as the mechanism of PZQ action is unknown. Here, we show PZQ activates a transient receptor potential melastatin ion channel (TRPMPZQ) in schistosomes by engaging a hydrophobic ligand binding pocket within the voltage-sensor like domain to cause Ca2+ entry and worm paralysis. PZQ activates TRPMPZQ homologues in other PZQ-sensitive flukes, but not Fasciola. However, a single amino acid change in the Fasciola TRPMPZQ binding pocket, to mimic schistosome TRPMPZQ, confers PZQ sensitivity. After decades of clinical use, the basis of PZQ action at a druggable TRP channel is resolved.

scholarly journals Structures and gating mechanism of human TRPM2

Science ◽  
2018 ◽  
Vol 362 (6421) ◽  
pp. eaav4809 ◽  
Author(s):  
Longfei Wang ◽  
Tian-Min Fu ◽  
Yiming Zhou ◽  
Shiyu Xia ◽  
Anna Greka ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Transient Receptor Potential ◽  
Trp Channels ◽  
Receptor Potential ◽  
Adenosine Diphosphate ◽  
Transmembrane Helices ◽  
Gating Mechanism ◽  
Channel Opening ◽  
Transient Receptor ◽  
Intersubunit Interactions

Transient receptor potential (TRP) melastatin 2 (TRPM2) is a cation channel associated with numerous diseases. It has a C-terminal NUDT9 homology (NUDT9H) domain responsible for binding adenosine diphosphate (ADP)–ribose (ADPR), and both ADPR and calcium (Ca2+) are required for TRPM2 activation. Here we report cryo–electron microscopy structures of human TRPM2 alone, with ADPR, and with ADPR and Ca2+. NUDT9H forms both intra- and intersubunit interactions with the N-terminal TRPM homology region (MHR1/2/3) in the apo state but undergoes conformational changes upon ADPR binding, resulting in rotation of MHR1/2 and disruption of the intersubunit interaction. The binding of Ca2+ further engages transmembrane helices and the conserved TRP helix to cause conformational changes at the MHR arm and the lower gating pore to potentiate channel opening. These findings explain the molecular mechanism of concerted TRPM2 gating by ADPR and Ca2+ and provide insights into the gating mechanism of other TRP channels.

scholarly journals Identification of the ADPR binding pocket in the NUDT9 homology domain of TRPM2

2017 ◽  
Vol 149 (2) ◽  
pp. 219-235 ◽  
Author(s):  
Peilin Yu ◽  
Xiwen Xue ◽  
Jianmin Zhang ◽  
Xupang Hu ◽  
Yan Wu ◽  
...  
Keyword(s):  
Transient Receptor Potential ◽  
Structural Model ◽  
Receptor Potential ◽  
Adenosine Diphosphate ◽  
Binding Pocket ◽  
Dynamics Simulation ◽  
Structure Based Drug Design ◽  
Transient Receptor Potential Melastatin ◽  
Trpm2 Channel ◽  
Transient Receptor

Activation of the transient receptor potential melastatin 2 (TRPM2) channel occurs during the response to oxidative stress under physiological conditions as well as in pathological processes such as ischemia and diabetes. Accumulating evidence indicates that adenosine diphosphate ribose (ADPR) is the most important endogenous ligand of TRPM2. However, although it is known that ADPR binds to the NUDT9 homology (NUDT9-H) domain in the intracellular C-terminal region, the molecular mechanism underlying ADPR binding and activation of TRPM2 remains unknown. In this study, we generate a structural model of the NUDT9-H domain and identify the binding pocket for ADPR using induced docking and molecular dynamics simulation. We find a subset of 11 residues—H1346, T1347, T1349, L1379, G1389, S1391, E1409, D1431, R1433, L1484, and H1488—that are most likely to directly interact with ADPR. Results from mutagenesis and electrophysiology approaches support the predicted binding mechanism, indicating that ADPR binds tightly to the NUDT9-H domain, and suggest that the most significant interactions are the van der Waals forces with S1391 and L1484, polar solvation interaction with E1409, and electronic interactions (including π–π interactions) with H1346, T1347, Y1349, D1431, and H1488. These findings not only clarify the roles of a range of newly identified residues involved in ADPR binding in the TRPM2 channel, but also reveal the binding pocket for ADPR in the NUDT9-H domain, which should facilitate structure-based drug design for the TRPM2 channel.

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