Structure of the ancestral TRPY1 channel from Saccharomyces cerevisiae reveals mechanisms of modulation by lipids and calcium

ABSTRACTTransient Receptor Potential (TRP) channels have evolved in eukaryotes to control various cellular functions in response to a wide variety of chemical and physical stimuli. This large and diverse family of channels emerged in fungi as mechanosensitive osmoregulators. The Saccharomyces cerevisiae vacuolar TRP yeast 1 (TRPY1) is the most studied TRP channel from fungi, but the molecular details of channel modulation remain elusive so far. Here, we describe the full-length cryo-electron microscopy structure of TRPY1 at 3.1 Å resolution. The structure reveals a distinctive architecture for TRPY1 among all eukaryotic TRP channels with an evolutionarily conserved and archetypical transmembrane domain, but distinct structural folds for the cytosolic N- and C-termini. We identified the inhibitory phosphatidylinositol 3-phosphate (PI(3)P) lipid binding site, which sheds light into the lipid modulation of TRPY1 in the vacuolar membrane. The structure also exhibited two Ca2+-binding sites: one in the cytosolic side, implicated in channel activation, and the other in the vacuolar lumen side, involved in channel inhibition. These findings, together with data from molecular dynamics simulations, provide structural insights into the basis of TRPY1 channel modulation by lipids and Ca2+.

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Structure of full-length human TRPM4

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1722038115 ◽

2018 ◽

Vol 115 (10) ◽

pp. 2377-2382 ◽

Cited By ~ 47

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.

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Molecular mechanism of TRPV2 channel modulation by cannabidiol

10.1101/521880 ◽

2019 ◽

Cited By ~ 2

Author(s):

Ruth A. Pumroy ◽

Amrita Samanta ◽

Yuhang Liu ◽

Taylor E.T. Hughes ◽

Siyuan Zhao ◽

...

Keyword(s):

Transient Receptor Potential ◽

Cannabis Sativa ◽

Trp Channels ◽

Critical Role ◽

Channel Gating ◽

Receptor Potential ◽

Lipid Binding ◽

Transient Receptor Potential Vanilloid ◽

Structure Based Drug Design ◽

Transient Receptor

SUMMARYTransient receptor potential vanilloid 2 (TRPV2) plays a critical role in neuronal development, cardiac function, immunity, and cancer. Cannabidiol (CBD), the non-psychotropic therapeutically active ingredient of Cannabis sativa, is a potent activator of TRPV2 and also modulates other transient receptor potential (TRP) channels. Here, we determined structures of the full-length TRPV2 channel in a CBD-bound state in detergent and in PI(4,5)P2 enriched nanodiscs by cryo-electron microscopy. CBD interacts with TRPV2 through a hydrophobic pocket located between S5 and S6 helices of adjacent subunits, which differs from known ligand and lipid binding sites in other TRP channels. Comparison between apo- and two CBD-bound TRPV2 structures reveals that the S4-S5 linker plays a critical role in channel gating upon CBD binding. The TRPV2 “vanilloid” pocket, which is critical for ligand-dependent gating in other TRPV channels, stays unoccupied by annular lipids, PI(4,5)P2, or CBD. Together these results provide a foundation to further understand TRPV channel gating properties and their divergent physiological functions and to accelerate structure-based drug design.

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The push-to-open mechanism of the tethered mechanosensitive ion channel NompC

eLife ◽

10.7554/elife.58388 ◽

2021 ◽

Vol 10 ◽

Author(s):

Yang Wang ◽

Yifeng Guo ◽

Guanluan Li ◽

Chunhong Liu ◽

Lei Wang ◽

...

Keyword(s):

Ion Channel ◽

Transient Receptor Potential ◽

Trp Channels ◽

Receptor Potential ◽

Ankyrin Repeat ◽

Gating Mechanism ◽

Repeat Domain ◽

Mechanosensitive Ion Channel ◽

Transient Receptor ◽

Dynamics Simulations

NompC is a mechanosensitive ion channel responsible for the sensation of touch and balance in Drosophila melanogaster. Based on a resolved cryo-EM structure, we performed all-atom molecular dynamics simulations and electrophysiological experiments to study the atomistic details of NompC gating. Our results showed that NompC could be opened by compression of the intracellular ankyrin repeat domain but not by a stretch, and a number of hydrogen bonds along the force convey pathway are important for the mechanosensitivity. Under intracellular compression, the bundled ankyrin repeat region acts like a spring with a spring constant of ~13 pN nm−1 by transferring forces at a rate of ~1.8 nm ps−1. The linker helix region acts as a bridge between the ankyrin repeats and the transient receptor potential (TRP) domain, which passes on the pushing force to the TRP domain to undergo a clockwise rotation, resulting in the opening of the channel. This could be the universal gating mechanism of similar tethered mechanosensitive TRP channels, which enable cells to feel compression and shrinkage.

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

Journal of Biological Chemistry ◽

10.1074/jbc.ra118.005066 ◽

2018 ◽

Vol 293 (41) ◽

pp. 16102-16114 ◽

Cited By ~ 14

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.

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Glutathione depletion activates the yeast vacuolar transient receptor potential channel, Yvc1p, by reversible glutathionylation of specific cysteines

Molecular Biology of the Cell ◽

10.1091/mbc.e16-05-0281 ◽

2016 ◽

Vol 27 (24) ◽

pp. 3913-3925 ◽

Cited By ~ 11

Author(s):

Avinash Chandel ◽

Krishna K. Das ◽

Anand K. Bachhawat

Keyword(s):

Calcium Channel ◽

Transient Receptor Potential ◽

Transmembrane Domain ◽

Trp Channels ◽

Calcium Influx ◽

Receptor Potential ◽

Transient Receptor Potential Channel ◽

Glutathione Depletion ◽

Glutathione S Transferase ◽

Transient Receptor

Glutathione depletion and calcium influx into the cytoplasm are two hallmarks of apoptosis. We have been investigating how glutathione depletion leads to apoptosis in yeast. We show here that glutathione depletion in yeast leads to the activation of two cytoplasmically inward-facing channels: the plasma membrane, Cch1p, and the vacuolar calcium channel, Yvc1p. Deletion of these channels partially rescues cells from glutathione depletion–induced cell death. Subsequent investigations on the Yvc1p channel, a homologue of the mammalian TRP channels, revealed that the channel is activated by glutathionylation. Yvc1p has nine cysteine residues, of which eight are located in the cytoplasmic regions and one on the transmembrane domain. We show that three of these cysteines, Cys-17, Cys-79, and Cys-191, are specifically glutathionylated. Mutation of these cysteines to alanine leads to a loss in glutathionylation and a concomitant loss in calcium channel activity. We further investigated the mechanism of glutathionylation and demonstrate a role for the yeast glutathione S-transferase Gtt1p in glutathionylation. Yvc1p is also deglutathionylated, and this was found to be mediated by the yeast thioredoxin, Trx2p. A model for redox activation and deactivation of the yeast Yvc1p channel is presented.

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Molecular mechanism of TRPV2 channel modulation by cannabidiol

eLife ◽

10.7554/elife.48792 ◽

2019 ◽

Vol 8 ◽

Cited By ~ 20

Author(s):

Ruth A Pumroy ◽

Amrita Samanta ◽

Yuhang Liu ◽

Taylor ET Hughes ◽

Siyuan Zhao ◽

...

Keyword(s):

Bound States ◽

Transient Receptor Potential ◽

Cannabis Sativa ◽

Trp Channels ◽

Critical Role ◽

Channel Gating ◽

Receptor Potential ◽

Lipid Binding ◽

Transient Receptor Potential Vanilloid ◽

Transient Receptor

Transient receptor potential vanilloid 2 (TRPV2) plays a critical role in neuronal development, cardiac function, immunity, and cancer. Cannabidiol (CBD), the non-psychotropic therapeutically active ingredient of Cannabis sativa, is an activator of TRPV2 and also modulates other transient receptor potential (TRP) channels. Here, we determined structures of the full-length rat TRPV2 channel in apo and CBD-bound states in nanodiscs by cryo-electron microscopy. We show that CBD interacts with TRPV2 through a hydrophobic pocket located between S5 and S6 helices of adjacent subunits, which differs from known ligand and lipid binding sites in other TRP channels. CBD-bound TRPV2 structures revealed that the S4-S5 linker plays a critical role in channel gating upon CBD binding. Additionally, nanodiscs permitted us to visualize two distinct TRPV2 apo states in a lipid environment. Together these results provide a foundation to further understand TRPV channel gating, their divergent physiological functions, and to accelerate structure-based drug design.

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Microbe Engineering of Guaia-6,10(14)-diene as a Building Block for the Semisynthetic Production of Plant-Derived (−)-Englerin A

10.26434/chemrxiv.10333625.v1 ◽

2019 ◽

Author(s):

Thomas Siemon ◽

Zhangqian Wang ◽

Guangkai Bian ◽

Tobias Seitz ◽

Ziling Ye ◽

...

Keyword(s):

Escherichia Coli ◽

Saccharomyces Cerevisiae ◽

Synthetic Biology ◽

Chemical Synthesis ◽

Building Block ◽

Transient Receptor Potential ◽

Receptor Potential ◽

Economical Method ◽

Englerin A ◽

Transient Receptor

Herein, we report the semisynthetic production of the potent transient receptor potential canonical (TRPC) channel agonist (−)-englerin A (EA), using guaia-6,10(14)-diene as the starting material. Guaia-6,10(14)-diene was systematically engineered in Escherichia coli and Saccharomyces cerevisiae using the CRISPR/Cas9 system and produced with high titers. This provided us the opportunity to execute a concise chemical synthesis of EA and the two related guaianes (−)-oxyphyllol and (+)-orientalol E. The potentially scalable approach combines the advantages of synthetic biology and chemical synthesis and provides an efficient and economical method for producing EA as well as its analogs.

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