Exploring functional roles of TRPV1 intracellular domains with unstructured peptide-insertion screening.
TRPV1 is a polymodal nociceptor for diverse physical and chemical stimuli that interact with different parts of the channel protein. Recent cryo-EM studies revealed detailed channel structures, opening the door for mapping structural elements mediating activation by each stimulus. Towards this goal, here we have combined unstructured peptide-insertion screening (UPS) with electrophysiological and fluorescence recordings to explore structural and functional roles of the intracellular regions of TRPV1 in mediating various activation stimuli. We found that most of the tightly packed protein regions did not tolerate structural perturbation by UPS when tested, indicating that structural integrity of the intracellular region is critical. In agreement with previous reports, Ca2+-dependent desensitization is strongly dependent on both intracellular N- and C-terminal domains; insertions of an unstructured peptide between these domains and the transmembrane core domain nearly eliminated Ca2+-dependent desensitization. In contrast, channel activations by capsaicin, low pH, divalent cations, and even heat are mostly intact in mutant channels containing the same insertions. These observations suggest that the transmembrane core domain of TRPV1, but not the intracellular domains, is responsible for sensing these stimuli.
Department of Physiology and Membrane Biology, University of California School of Medicine, Davis CA 95616, USA.|Institute for Molecular Bioscience, University of Queensland, St Lucia, QLD 4072, Australia.
Linlin
L
Ma
Department of Physiology and Membrane Biology, University of California School of Medicine, Davis CA 95616, USA.
Fan
F
Yang
Department of Physiology and Membrane Biology, University of California School of Medicine, Davis CA 95616, USA.
Simon
S
Vu
Department of Physiology and Membrane Biology, University of California School of Medicine, Davis CA 95616, USA.
Jie
J
Zheng
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