Binding of Extracellular Ca2+ to the Specific Amino Acids is Required for Heat-Evoked Activation of TRPA1

Abstract

Transient receptor potential, ankyrin 1 (TRPA1) is a homotetrameric nonselective cation- permeable channel with six transmembrane domains handed by cytoplasmic N and C termini. Although many TRP channel family members contain the range of 3-6 ankyrin repeats (AR) within the N-terminal region, TRPA1 is distinguished by having an unusually large number of such repeats (16-17 ARs). A number of studies have shown that both intracellular and extracellular Ca2+ is a key regulator of many TRP channels, including TRPA1. In the previous study, we found that extracellular Ca2+, but not intracellular Ca2+ plays an important role in heat-evoked activation of green anole TRPA1 (gaTRPA1). In this study, we focus on extracellular Ca2+-dependent heat sensitivity of gaTRPA1 by comparing gaTRPA1 with other heat-activated TRPA1s from rat-snake (rsTRPA1) and chicken (chTRPA1). We found that rsTRPA1 and chTRPA1s are activated by heat with small inward currents in the absence of extracellular Ca2+. We identified several negatively charged amino acid residues (glutamate and aspartate) near outer pore vestibule in gaTRPA1, chTRPA1 and rsTRPA1 for activation by heat in the presence of extracellular Ca2+. These results suggest that the neutralization of the acidic amino acids by extracellular Ca2+ is important for the heat-evoked activation of gaTRPA1, chTRPA1 and rsTRPA1.