Functional Role of C-Terminal Cytoplasmic Tail of Rat Vanilloid Receptor 1

Abstract

The vanilloid receptor [transient receptor potential (TRP)V1, also known as VR1] is a member of the TRP channel family. These receptors share a significant sequence homology, a similar predicted structure with six transmembrane-spanning domains (S1-S6), a pore-forming region between S5 and S6, and the cytoplasmically oriented C- and N-terminal regions. Although structural/functional studies have identified some of the key amino acids influencing the gating of the TRPV1 ion channel, the possible contributions of terminal regions to vanilloid receptor function remain elusive. In the present study, C-terminal truncations of rat TRPV1 have been constructed to characterize the contribution of the cytoplasmic C-terminal region to TRPV1 function and to delineate the minimum amount of C tail necessary to form a functional channel. The truncation of 31 residues was sufficient to induce changes in functional properties of TRPV1 channel. More pronounced effects of C-terminal truncation were seen in mutants lacking the final 72 aa. These changes were characterized by a decline of capsaicin-, pH-, and heat-sensitivity; progressive reduction of the activation thermal threshold (from 41.5 to 28.6 degrees C); and slowing of the activation rate of heat-evoked membrane currents (Q10 from 25.6 to 4.7). The voltage-induced currents of the truncated mutants exhibited a slower onset, markedly reduced outward rectification, and significantly smaller peak tail current amplitudes. Truncation of the entire TRPV1 C-terminal domain (155 residues) resulted in a nonfunctional channel. These results indicate that the cytoplasmic COOH-terminal domain strongly influences the TRPV1 channel activity, and that the distal half of this structural domain confers specific thermal sensitivity.

Carboxyl-terminal Domain of Transient Receptor Potential Vanilloid 1 Contains Distinct Segments Differentially Involved in Capsaicin- and Heat-induced Desensitization

“…Both TRPV1 ctV3 and ctV3(B) showed reduced ⌬H°, suggesting that the altered heat sensitivity of ctV3 and ctV3(B) is commonly derived from the alteration of the distal half of CTD. Altered heat sensitivity conferred by truncation of the distal half of the CTD has been reported in rat TRPV1 and in tissue-specific splicing variants of TRPV1 in vampire bats (28,39,40). In contrast to activation, TRPV1 ctV3(B) displayed heat-induced desensitization similar to WT, whereas ctV3 lacked heat-induced desensitization.…”

“…It is unclear how the distal end of the CTD affects heat-induced desensitization. A previous molecular modeling study based upon the structure of a homologous protein predicted that the distal end of the CTD contains an ␣-helical structure (28). We assume that the presumptive ␣-helix may be involved in heat-induced desensitization, which is affected by truncation of the C⌬7 segment.…”

“…TRPV1 CTD has been implicated in heat activation (6,28,39). The chimeric approach provided a clue as to the roles of TRPV1 CTD in heat-induced desensitization.…”

“…E rev was corrected for junction potentials that were calculated using pClamp software. The temperature coefficient (Q 10 ) and temperature threshold of activation were determined following the methods previously described (28). An Arrhenius plot of logarithm of current amplitudes against the reciprocal of the absolute temperature (K) was generated.…”

Identification of Species-specific Determinants of the Action of the Antagonist Capsazepine and the Agonist PPAHV on TRPV1

“…We did not identify the relevant residues in this region due to the relatively small size of the effect, however, other studies have shown that amino acids in the N-and C-terminal tails can affect capsaicin binding and function (26,27).…”