Directionality of Temperature Activation in Mouse TRPA1 Ion Channel Can Be Inverted by Single-Point Mutations in Ankyrin Repeat Six

Several transient receptor potential (TRP) ion channels are activated with high sensitivity by either cold or hot temperatures. However, structures and mechanism that determine temperature directionality (cold versus heat) are not established. Here we screened 12,000 random mutant clones of the cold-activated mouse TRPA1 ion channel with a heat stimulus. We identified three single-point mutations that are individually sufficient to make mouse TRPA1 warm activated, while leaving sensitivity to chemicals unaffected. Mutant channels have high temperature sensitivity of voltage activation, specifically of channel opening, but not channel closing, which is reminiscent of other heat-activated TRP channels. All mutations are located in ankyrin repeat six, which identifies this domain as a sensitive modulator of thermal activation. We propose that a change in the coupling of temperature sensing to channel gating generates this sensitivity to warm temperatures. Our results demonstrate that minimal changes in protein sequence are sufficient to generate a wide diversity of thermal sensitivities in TRPA1.