TRPV1 Channels Are Intrinsically Heat Sensitive and Negatively Regulated by Phosphoinositide Lipids

Abstract

Summary The capsaicin receptor, TRPV1, is regulated by phosphatidylinositol-4,5-bisphosphate (PIP2), although the precise nature of this effect (i.e., positive or negative) remains controversial. Here, we reconstitute purified TRPV1 into artificial liposomes, where it is gated robustly by capsaicin, protons, spider toxins and, notably, heat, demonstrating intrinsic sensitivity of the channel to both chemical and thermal stimuli. TRPV1 is fully functional in the absence of phosphoinositides, arguing against their proposed obligatory role in channel activation. Rather, introduction of various phosphoinositides, including PIP2, PI4P and PI, inhibits TRPV1, supporting a model whereby phosphoinositide turnover contributes to thermal hyperalgesia by disinhibiting the channel. Using an orthogonal chemical strategy, we show that association of the TRPV1 C-terminus with the bilayer modulates channel gating, consistent with phylogenetic data implicating this domain as a key regulatory site for tuning stimulus sensitivity. Beyond TRPV1, these findings are relevant to understanding how membrane lipids modulate other “receptor-operated” TRP channels.

Molecular sensors and modulators of thermoreception

“…However, there is a continuing controversy regarding the exact role of PIP 2 in TRPV1 activation, with some supporting an inhibiting role, 60,61 and some advocating an stimulating effect, [62][63][64][65][66][67][68][69] whereas others favoring both activating and inhibiting roles depending on certain conditions. 70,71 Different approaches used for manipulating the cellular PIP 2 level may underlie the difference, with some including PIP 2 into the whole-cell recording pipette, 65,67 and some applying PIP 2 directly to an inside-out excised patch, 63,68,69 whereas others reconstituting PIP 2 and purified TRPV1 in an artificial liposome or into planar lipid bilayers.…”

Mechanism for phosphoinositide selectivity and activation of TRPV1 ion channels

“…PI(4,5)P 2 has been proposed to inhibit TRPV1, mediating the sensitizing actions of bradykinin and nerve growth factor (Chuang et al, 2001;Cao et al, 2013a). In contrast, others (including us) have proposed that PI(4,5)P 2 activates TRPV1 (Stein et al, 2006;Lukacs et al, 2007Lukacs et al, , 2013aKlein et al, 2008), and the depletion of PI(4,5)P 2 by phospholipase C may play a role in Ca 2+ -dependent desensitization (Mercado et al, 2010;Lukacs et al, 2013b).…”

Regulation of thermoTRPs by lipids

“…Prescott and Julius as well as Cao et al, have shown that the negatively-charged phosphate group of PIP 2 interacts with a stretch of positively-charged amino acids (777-820) in the C-terminus. 10,18 Other groups have proposed that PIP 2 interacts with a more proximal region of the C-terminus. 19 Specifically, it has been shown that the important amino acid residues for PIP 2 interaction with TRPV1 include R701 and K710 19 and/or R721.…”

“…24 In contrast, Cao and collaborators performed experiments were they incorporated purified TRPV1 into artificial lipid vesicles and showed that, under conditions where phosphoinositides were added, the sensitivity of the channel to heat was decreased. 25 Thus, the role of PIP 2 on thermal sensitivity related to TRPV1 activation needs further clarification.…”

“…18 LPC is an endogenous activator of the TRPV2 channel that induces Ca 2C influx through this protein. By activating Gq/Go-protein and phosphatidylinositol-3,4 kinase signaling, LPC causes translocation of TRPV2 to the plasma membrane and leads to migration in cells of the prostate cancer cell line PC3.…”