Critical amino acid residues regulating TRPA1 Zn 2+ response: A comparative study across species.

Cellular zinc ions (Zn ²⁺ ) are crucial for signal transduction in various cell types. The transient receptor potential (TRP) ankyrin 1 (TRPA1) channel, known for its sensitivity to intracellular Zn ²⁺ ([Zn ²⁺ ] _i ), has been a subject of limited understanding regarding its molecular mechanism. Here, we used metal ion-affinity prediction, three-dimensional structural modeling, and mutagenesis, utilizing data from the Protein Data Bank and AlphaFold database, to elucidate the [Zn ²⁺ ] _i binding domain (IZD) structure composed by specific AAs residues in human (hTRPA1) and chicken TRPA1 (gTRPA1). External Zn ²⁺ induced activation in hTRPA1, while not in gTRPA1. Moreover, external Zn ²⁺ elevated [Zn ²⁺ ] _i specifically in hTRPA1. Notably, both hTRPA1 and gTRPA1 exhibited inherent sensitivity to [Zn ²⁺ ] _i , as evidenced by their activation upon internal Zn ²⁺ application. The critical AAs within IZDs, specifically histidine at 983/984, lysine at 711/717, tyrosine at 714/720, and glutamate at 987/988 in IZD1, and H983/H984, tryptophan at 710/716, E854/E855, and glutamine at 979/980 in IZD2, were identified in hTRPA1/gTRPA1. Furthermore, mutations, such as the substitution of arginine at 919 (R919) to H919, abrogated the response to external Zn ²⁺ in hTRPA1. Among single-nucleotide polymorphisms (SNPs) at Y714 and a triple SNP at R919 in hTRPA1, we revealed that the Zn ²⁺ responses were attenuated in mutants carrying the Y714 and R919 substitution to asparagine and proline, respectively. Overall, this study unveils the intrinsic sensitivity of hTRPA1 and gTRPA1 to [Zn ²⁺ ] _i mediated through IZDs. Furthermore, our findings suggest that specific SNP mutations can alter the responsiveness of hTRPA1 to extracellular and intracellular Zn ²⁺ .
Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article.
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