Genetic variants affecting human TRPA1 or TRPM8 structure can be classified in vitro as ‘well expressed’, ‘poorly expressed’ or ‘salvageable’

Abstract

SynopsisMultiple mis-sense variants of TRPA1 (transient receptor potential A1) and TRPM8 (transient receptor potential M8) are recorded in the human genome single nt polymorphism (SNP) database, but their potential impact on channel signalling in patho-physiology is not fully explored. Variants, mostly quite rare in the general human population, alter sites in different structural domains of these homo-tetrameric ion channel proteins. The effects of individual SNPs affecting the large cytoplasmic N-terminal domain have not been completely documented for TRPM8 or TRPA1. We examined the Ca 2 + signalling properties of a short-list of eight variants affecting the N-terminal domain by individual expression in human embryonic kidney HEK293 or neuroblastoma (SH-SY5Y) cell lines (four SNP variants for TRPM8: G150R, K423N, R475C, R485W and four for TRPA1: Y69C, A366D, E477K, D573A). These were compared with TRPA1 SNP variants affecting intracellular loops located beyond the N-terminal domain and associated with gain of function (such as increased sensitivity to agonists: TRPA1 R797T and N855S). A substitution in TRPA1 (Y69C) exhibited high expression/sensitivity to agonists (high iCa 2 + max (maximum level of intracellular calcium ion), similar to R797T, but less sensitive than N855S), whereas each of the other non-conservative substitutions exhibited poor signalling response (low iCa 2 + max). Responses from these poorly expressed variants could be salvaged, to different extents, by pre-treating cells with the Src (Src protein) family inhibitor protein kinase inhibitor PP2 (PP2: 4-Amino-3-(4-chlorophenyl)-1-(t-butyl)-1H-pyrazolo [3,4-d]pyrimidine, 4-Amino-5-(4-chlorophenyl)-7-(tbutyl)pyrazolo [3,4-d]pyrimidine), or with micromolar Zn 2 + . The TRPA1 variants and several experimental mutants (TRPA1 Y97F, Y226F and YY654-655FF) expressed poorly in SH-SY5Y compared with HEK293 cells. More in-depth studies are needed to identify SNP variants eliciting gain of function in these TRP (transient receptor potential) channels and to assess their roles in medical conditions.

Activation of mutated TRPA1 ion channel by resveratrol in human prostate cancer associated fibroblasts (CAF)

“…Indeed, it has been described that the channels with Y69C and K186N substitutions exhibited high expression and sensitivity to TRPA1 agonists. In this report, a substitution in TRPA1 (Y69C) exhibited high expression/sensitivity to agonists . These observations could explain the activation of TRPA1 expressed in human prostate CAFs while the wt‐hTRPA1 is inhibited by the polyphenol.…”

“…In this report, a substitution in TRPA1 (Y69C) exhibited high expression/ sensitivity to agonists. 23…”

TRPM8 and Migraine

“…It remains to be tested whether and how the three variants alter the TRPM8 expression level, as suggested by their locations in regions involved in transcriptional regulation. However, a recent study suggests that some TRPM8 SNP variants causing amino acid substitutions affect the expression level as well as the function of TRPM8 channels in cell lines, providing support for the idea that the variants identified in migraine patients may indeed impact channel expression/function.…”

The Development of a Menthol Solution for Use during Sport and Exercise

“…Perception of menthol mouth swill concentration may be highly individual in nature, with 200 gene variants of the receptor responsible for menthol detection (TRPM8 [32]). This may in part explain the broad standard deviations and confidence intervals overlapping zero in our results.…”