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Somatosensory Channel for Irritants


Nature 520, 511517 (23 April 2015)

Structure of the TRPA1 ion channel suggests regulatory mechanisms

Candice E. Paulsen,

Department of Physiology, University of California, San Francisco, California 94158-2517, USA

Keck Advanced Microscopy Laboratory, Department of Biochemistry and Biophysics, University of California, San Francisco, California 94158-2517, USA


The TRPA1 ion channel (also known as the wasabi receptor) is a detector of noxious chemical agents encountered in our environment or produced endogenously during tissue injury or drug metabolism. These include a broad class of electrophiles that activate the channel through covalent protein modification. TRPA1 antagonists hold potential for treating neurogenic inflammatory conditions provoked or exacerbated by irritant exposure. Despite compelling reasons to understand TRPA1 function, structural mechanisms underlying channel regulation remain obscure. Here we use single-particle electron cryo- microscopy to determine the structure of full-length human TRPA1 to ~4  resolution in the presence of pharmacophores, including a potent antagonist. Several unexpected features are revealed, including an extensive coiled-coil assembly domain stabilized by polyphosphate co-factors and a highly integrated nexus that converges on an unpredicted transient receptor potential (TRP)-like allosteric domain. These findings provide new insights into the mechanisms of TRPA1 regulation, and establish a blueprint for structure-based design of analgesic and anti-inflammatory agents.

TRP ion channels have crucial roles in somatosensation by serving as sensors for thermal and chemical stimuli. In mammals, the TRPA1 subtype (so named for its extensive amino-terminal ankyrin repeat domain) is expressed by primary afferent nociceptors, where it detects structurally diverse noxious compounds that elicit pain and neurogenic inflammation. Such activators include pungent irritants from mustard, onion and garlic, as well as volatile environmental toxins and endogenous pro-algesic agents. TRPA1 is also activated downstream of phospholipase-C-coupled receptors and has been proposed to function as a sensor of noxious cold. TRPA1 is associated with persistent pain, respiratory and chronic itch syndromes and is therefore a promising target for treating these and other neurogenic inflammatory conditions. While selective TRPA1 antagonists have been developed, their sites and mechanisms of action remain unclear.

Many TRPA1 agonists are potent electrophiles that activate the channel through covalent modification of conserved cysteine or lysine residues within the cytoplasmic N terminus14, 15. While these and other functional properties have been deduced from electrophysiological studies of TRPA1 in whole cells, channel activity is not readily retained in excised membrane patches. This run down can be mitigated if membranes are excised into solutions containing polyphosphates, suggesting that obligate cytoplasmic co-factors support TRPA1 function in intact cells. Therefore, determining the biophysical and structural basis of polyphosphate regulation is key to understanding how TRPA1 is regulated in vivo, and how it can be efficiently manipulated in artificial systems for more detailed functional characterization.

A transformative step in addressing these questions would be to determine the three-dimensional (3D) atomic structure of the channel. TRP channels have posed particular challenges in this regard, probably reflecting their conformationally dynamic nature and diverse intracellular elements. Electron microscopy provides a potential means to achieve this goal, although for TRPA1 this approach has until now yielded densities of relatively low resolution (~16 ). However, advances in single-particle electron cryo-microscopy (cryo-EM) have recently enabled de novo structural analysis of TRPV1 to near-atomic (≤4.0 ) resolution. Here, we exploit this approach to determine the structure of the full-length human TRPA1 channel to ~4  resolution, revealing the structural basis of subunit assembly, polyphosphate action and antagonist binding.

TRPA1 is a sensor for chemical irritants and a major contributor to chemonociception. We now show that key residues involved in irritant detection are solvent accessible and lie within a putative allosteric nexus converging on an unpredicted TRP-like domain, suggesting a structural basis in which TRPA1 functions as a sensitive, low-threshold electrophile receptor. An important next step is to visualize electrophile-evoked conformational changes that are associated with gating, a goal that will require robust stabilization of TRPA1 under conditions amenable to structural studies.

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