Structural basis of TRPC4 regulation by calmodulin and pharmacological agents
- Max Planck Institute of Molecular Physiology, Germany
- Julius-Maximilians-Universität Würzburg, Germany
- Max Planck Institute for Molecular Physiology, Germany
- Goldfinch Bio Inc, United States
Abstract
Canonical transient receptor potential channels (TRPC) are involved in receptor-operated and/or store-operated Ca2+ signaling. Inhibition of TRPCs by small molecules was shown to be promising in treating renal diseases. In cells, the channels are regulated by calmodulin. Molecular details of both calmodulin and drug binding have remained elusive so far. Here we report structures of TRPC4 in complex with three pyridazinone-based inhibitors and calmodulin. The structures reveal that all the inhibitors bind to the same cavity of the voltage-sensing-like domain and allow us to describe how structural changes from the ligand binding site can be transmitted to the central ion-conducting pore of TRPC4. Calmodulin binds to the rib helix of TRPC4, which results in the ordering of a previously disordered region, fixing the channel in its closed conformation. This represents a novel calmodulin-induced regulatory mechanism of canonical TRP channels.
Data availability
The atomic coordinates and cryo-EM maps for TRPC4DR in complex with inhibitors, calmodulin and for TRPC4DR in LMNG are available at the Protein Data Bank (PDB) and Electron Microscopy Data Bank (EMDB) databases, under the accession numbers PBD 7B0S and EMD-11970 (TRPC4-GFB8438), PBD 7B16 and EMD-11979 (TRPC4-GFB9289); PBD 7B05 and EMD-11957 (TRPC4-GFB8749); PBD 7B1G and EMD-11985 (TRPC4-Calmodulin) and PBD 7B0J and EMD-11968 (TRPC4-apo in LMNG).
Article and author information
Author details
Funding
Max-Planck-Gesellschaft
- Stefan Raunser
Deutsche Forschungsgemeinschaft (TR240)
- Georg Nagel
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Copyright
© 2020, Vinayagam et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
Metrics
-
- 3,038
- views
-
- 488
- downloads
-
- 53
- citations
Views, downloads and citations are aggregated across all versions of this paper published by eLife.
Download links
A two-part list of links to download the article, or parts of the article, in various formats.
Downloads (link to download the article as PDF)
Open citations (links to open the citations from this article in various online reference manager services)
Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)
Structural basis of TRPC4 regulation by calmodulin and pharmacological agents
eLife 9:e60603.
https://doi.org/10.7554/eLife.60603
Further reading
-
- Structural Biology and Molecular Biophysics
The canonical chemokine receptor CXCR4 and atypical receptor ACKR3 both respond to CXCL12 but induce different effector responses to regulate cell migration. While CXCR4 couples to G proteins and directly promotes cell migration, ACKR3 is G-protein-independent and scavenges CXCL12 to regulate extracellular chemokine levels and maintain CXCR4 responsiveness, thereby indirectly influencing migration. The receptors also have distinct activation requirements. CXCR4 only responds to wild-type CXCL12 and is sensitive to mutation of the chemokine. By contrast, ACKR3 recruits GPCR kinases (GRKs) and β-arrestins and promiscuously responds to CXCL12, CXCL12 variants, other peptides and proteins, and is relatively insensitive to mutation. To investigate the role of conformational dynamics in the distinct pharmacological behaviors of CXCR4 and ACKR3, we employed single-molecule FRET to track discrete conformational states of the receptors in real-time. The data revealed that apo-CXCR4 preferentially populates a high-FRET inactive state, while apo-ACKR3 shows little conformational preference and high transition probabilities among multiple inactive, intermediate and active conformations, consistent with its propensity for activation. Multiple active-like ACKR3 conformations are populated in response to agonists, compared to the single CXCR4 active-state. This and the markedly different conformational landscapes of the receptors suggest that activation of ACKR3 may be achieved by a broader distribution of conformational states than CXCR4. Much of the conformational heterogeneity of ACKR3 is linked to a single residue that differs between ACKR3 and CXCR4. The dynamic properties of ACKR3 may underly its inability to form productive interactions with G proteins that would drive canonical GPCR signaling.
-
- Immunology and Inflammation
- Structural Biology and Molecular Biophysics
A potent class of HIV-1 broadly neutralizing antibodies (bnAbs) targets the envelope glycoprotein’s membrane proximal exposed region (MPER) through a proposed mechanism where hypervariable loops embed into lipid bilayers and engage headgroup moieties alongside the epitope. We address the feasibility and determinant molecular features of this mechanism using multi-scale modeling. All-atom simulations of 4E10, PGZL1, 10E8, and LN01 docked onto HIV-like membranes consistently form phospholipid complexes at key complementarity-determining region loop sites, solidifying that stable and specific lipid interactions anchor bnAbs to membrane surfaces. Ancillary protein-lipid contacts reveal surprising contributions from antibody framework regions. Coarse-grained simulations effectively capture antibodies embedding into membranes. Simulations estimating protein-membrane interaction strength for PGZL1 variants along an inferred maturation pathway show bilayer affinity is evolved and correlates with neutralization potency. The modeling demonstrated here uncovers insights into lipid participation in antibodies’ recognition of membrane proteins and highlights antibody features to prioritize in vaccine design.
