Structural basis for pharmacological modulation of the TRPC6 channel

Bai, Yonghong; Yu, Xianjun; Chen, Hao; Horne, Daniel B.; White, Ryan D.; Wu, Xingkun; Lee, Paul; Gu, Yun; Ghimire-Rijal, Sudipa; Lin, Don S.; Huang, Xin

doi:10.7554/elife.53311

Cited by 86 publications

(148 citation statements)

References 50 publications

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“…The TRPC5 lipid binding site that Pico145 interacts with is highly conserved within the TRPC family (Supplementary Table 2). Bound (phospho)lipids have been found in this site in structures of hTRPC3, 35,37 mTRPC4, 33 mTRPC5, 34 and hTRPC6, 35,36 and the TRPC6 activator AM-0883 displaces the ordered lipid bound in this site near the P-loop of hTRPC6 (Supplementary Figure 5F). 36 The hTRPC5 residues F576 and W577 are conserved throughout the TRPC family and are likely to be responsible for lipid binding.…”

Section: Discussionmentioning

confidence: 97%

“…Indeed, a similar density is present in the mTRPC4 structure ( Figure 2C). The hTRPC3 35,37 and hTRPC6 35,36 structures contain lipids that interact with their LFW motifs as well, but with altered geometry (Supplementary Figure 5). Our TRPC5:Pico145 structure also showed density in this region.…”

Section: Pico145 Binds To a Conserved Lipid Binding Site Of Trpc5mentioning

confidence: 99%

“…In the zinc binding domain, the densities were similar (and partially disordered) in the TRPC5:Zn 2+ and TRPC5:Pico145 structures, suggesting that the site may already be fully occupied in the absence of added ZnCl2. A structural alignment showed that the putative zinc binding motif is present in the electron density maps of all TRPC channel structures; it was built into the hTRPC3 35,37 and hTRPC6 35,36 structures, but not into the mTRPC4 33 or mTRPC5 34 structures (Supplementary Figure 10). These findings suggest that TRPC channels have a conserved intracellular zinc binding site.…”

Section: Mutations In the Xanthine Binding Site Alter The Response Ofmentioning

confidence: 99%

“…34 To date, TRPC6 is the only TRPC channel for which small molecule-bound structures have been reported. 35,36 Here, we report the first structure of a TRPC1/4/5 channel in complex with a small-molecule modulator: the homomeric TRPC5:C5 channel in complex with Pico145. We found that Pico145 binds between the trans-membrane domains of two TRPC5 subunits, where it displaces a phospholipid in a conserved lipid binding site.…”

Section: Introductionmentioning

confidence: 99%

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Cryo-EM structures of human TRPC5 reveal interaction of a xanthine-based TRPC1/4/5 inhibitor with a conserved lipid binding site

Wright

Simmons

et al. 2020

Preprint

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TRPC1/4/5 channels are non-specific cation channels implicated in a wide variety of diseases, and TRPC1/4/5 inhibitors have recently entered the first clinical trials. However, fundamental and translational studies require a better understanding of TRPC1/4/5 channel regulation by endogenous and exogenous factors. Although several potent and selective TRPC1/4/5 modulators have been reported, the paucity of mechanistic insights into their modes-of-action remains a barrier to the development of new chemical probes and drug candidates. The xanthine class of modulators includes the most potent and selective TRPC1/4/5 inhibitors described to date, as well as TRPC5 activators. Our previous studies suggest that xanthines interact with a, so far, elusive pocket of TRPC1/4/5 channels that is essential to channel gating. Targeting this pocket may be a promising strategy for TRPC1/4/5 drug discovery. Here we report the first structure of a small molecule-bound TRPC1/4/5 channelhuman TRPC5 in complex with the xanthine Pico145to 3.0 Å. We found that Pico145 binds to a conserved lipid binding site of TRPC5, where it displaces a bound phospholipid. Our findings explain the mode-of-action of xanthine-based TRPC1/4/5 modulators, and suggest a structural basis for TRPC1/4/5 modulation by endogenous factors such as (phospho)lipids and Zn 2+ ions. These studies lay the foundations for the structure-based design of new generations of TRPC1/4/5 modulators.

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Section: Discussionmentioning

confidence: 97%

Section: Pico145 Binds To a Conserved Lipid Binding Site Of Trpc5mentioning

confidence: 99%

Section: Mutations In the Xanthine Binding Site Alter The Response Ofmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Cryo-EM structures of human TRPC5 reveal interaction of a xanthine-based TRPC1/4/5 inhibitor with a conserved lipid binding site

Wright

Simmons

et al. 2020

Preprint

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“…Thanks to the efforts from several groups ( 40, 41 ), three inhibitor binding pockets have been structurally identified in hTRPC5 and hTRPC6, namely inhibitor binding pocket (IBP) A-C (Fig. 6 and Fig.…”

Section: Discussionmentioning

confidence: 99%

Structural basis for human TRPC5 channel inhibition by two distinct inhibitors

Song

Wei

Guo

et al. 2020

Preprint

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17TRPC5 channel is a non-selective cation channel that participates diverse physiological 18 processes. Human TRPC5 inhibitors show promise in the treatment of anxiety disorder, 19 depression and kidney disease. Despite the high relevance of TRPC5 to human health, its 20 inhibitor binding pockets have not been fully characterized due to the lack of structural 21 information, which greatly hinders structure-based drug discovery. Here we show cryo-EM 22 structures of human TRPC5 in complex with two distinct inhibitors, namely clemizole and 23 HC-070, to the resolution of 2.7 Å. Based on the high-quality cryo-EM maps, we uncover the 24 different binding pockets and detailed binding modes for these two inhibitors. Clemizole 25 binds inside the voltage sensor-like domain of each subunit, while HC-070 binds close to the 26 ion channel pore and is wedged between adjacent subunits. Both of them exert the inhibitory 27 function by stabilizing the ion channel in a closed state. These structures provide templates 28 for further design and optimization of inhibitors targeting human TRPC5. 29 70 inhibitory mechanism of these two distinct inhibitors against human TRPC5 (hTRPC5), we 71 embarked structural studies of hTRPC5 in complex with CMZ or HC-070. Our high 72 resolution cryo-EM maps are sufficient to identify their binding pockets and will aid further 73 drug development. 74 75 76 Results 77 78 Structure of human TRPC5 in complex with inhibitors 79To reveal the binding sites of CMZ and HC-070 on hTRPC5, we expressed hTRPC5 in 80 HEK293F cells for structural studies. Similar to the mouse TRPC5 counterpart (mTRPC5) 81 (33), C-terminal truncation of hTRPC5 generated a construct (hTRPC5 1-764 ) that showed 82 higher expression level of the tetramer in comparison with the full length wild type hTRPC5 83 ( Fig. S1A). Moreover, we found hTRPC5 1-764 retained the pharmacological properties of full 84 length hTRPC5 such as activation by EA ( Fig. S1, B to E). Therefore, we used hTRPC5 1-764 85 protein for structural studies. Tetrameric hTRPC5 1-764 channel was purified in detergent 86 micelles (Fig. S1F and G). To obtain the CMZ-bound and the HC-070-bound TRPC5 87 structures, CMZ or HC-070 was added to the concentrated hTRPC5 1-764 protein for cryo-EM 88 sample preparation. We obtained maps of hTRPC5 in CMZ-bound state and HC-070-bound 89 state to the resolution of 2.7 Å (Fig. 1, Figs. S2 to S5; table S1). The map qualities were 90 sufficient for model building and ligand assignment (Fig. S3, D and E, and Fig. S5, D and E).91The overall architecture of inhibitor-bound hTRPC5 is similar to the structure of mTRPC5 in 92 apo state (33), occupying 100 Å × 100 Å × 130 Å in three-dimensional space (Fig. 1). The 93 four-fold symmetric channel has two layer architecture, composed of the intracellular 94 cytosolic domain (ICD) layer and the transmembrane domain (TMD) layer (Fig. 1). In ICD 95 layer, the N-terminal ankyrin repeats domain (ARD) is below the linker-helix domain (LHD) 96 region (Fig .1F). The TRP helix mediates interactions...

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The Calcium Signaling Mechanisms in Arterial Smooth Muscle and Endothelial Cells

Ottolini

Sonkusare

2021

Comprehensive Physiology

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The contractile state of resistance arteries and arterioles is a crucial determinant of blood pressure and blood flow. Physiological regulation of arterial contractility requires constant communication between endothelial and smooth muscle cells. Various Ca 2+ signals and Ca 2+ -sensitive targets ensure dynamic control of intercellular communications in the vascular wall. The functional effect of a Ca 2+ signal on arterial contractility depends on the type of Ca 2+ -sensitive target engaged by that signal. Recent studies using advanced imaging methods have identified the spatiotemporal signatures of individual Ca 2+ signals that control arterial and arteriolar contractility. Broadly speaking, intracellular Ca 2+ is increased by ion channels and transporters on the plasma membrane and endoplasmic reticular membrane. Physiological roles for many vascular Ca 2+ signals have already been confirmed, while further investigation is needed for other Ca 2+ signals. This article focuses on endothelial and smooth muscle Ca 2+ signaling mechanisms in resistance arteries and arterioles. We discuss the Ca 2+ entry pathways at the plasma membrane, Ca 2+ release signals from the intracellular stores, the functional and physiological relevance of Ca 2+ signals, and their regulatory mechanisms. Finally, we describe the contribution of abnormal endothelial and smooth muscle Ca 2+ signals to the pathogenesis of vascular disorders.

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Structural basis for pharmacological modulation of the TRPC6 channel

Cited by 86 publications

References 50 publications

Cryo-EM structures of human TRPC5 reveal interaction of a xanthine-based TRPC1/4/5 inhibitor with a conserved lipid binding site

Cryo-EM structures of human TRPC5 reveal interaction of a xanthine-based TRPC1/4/5 inhibitor with a conserved lipid binding site

Structural basis for human TRPC5 channel inhibition by two distinct inhibitors

The Calcium Signaling Mechanisms in Arterial Smooth Muscle and Endothelial Cells

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