Thermosensory and Nonthermosensory Isoforms of Drosophila melanogaster TRPA1 Reveal Heat-Sensor Domains of a ThermoTRP Channel

Abstract

SUMMARY Specialized somatosensory neurons detect temperatures ranging from pleasantly cool or warm to burning hot and painful (nociceptive). The precise temperature ranges sensed by thermally sensitive neurons is determined by tissue specific expression of ion channels of the Transient Receptor Potential (TRP) family. We show here, that in Drosophila, TRPA1 is required for sensing of nociceptive heat. We identify two new protein isoforms of dTRPA1named dTRPA1-C and dTRPA1-D that explain this requirement. A dTRPA1-C/D reporter was exclusively expressed in nociceptors and dTRPA1-C rescued thermal nociception phenotypes when restored to mutant nociceptors. However, surprisingly, we find that dTRPA1-C is not a direct heat sensor. Alternative splicing generates at least four isoforms of dTRPA1. Our analysis of these isoforms reveals a 37 amino acid intracellular region (encoded by a single exon) that is critical for dTRPA1 temperature responses. The identification of these amino acids opens the door to a biophysical understanding of a molecular thermosensor.

Drosophila TRPA1 isoforms detect UV light via photochemical production of H 2 O 2

“…We found that ectopic expression of these H 2 O 2 -sensitive dTRPA1 isoforms conferred UV sensitivity to light-insensitive cultured HEK293 cells and a few types of light-insensitive Drosophila neurons. The light and H 2 O 2 sensitivities are specific to certain dTRPA1 isoforms, consistent with previous findings that different dTRPA1 isoforms exhibit distinct thermal sensitivities (20,21). Strikingly, the H 2 O 2 -sensitive dTRPA1 was as effective as channelrhodopsin-2 (ChR2) in triggering light-produced motor responses when expressed in a specific group of motor neurons that control proboscis extension…”

“…Our results suggest specific H 2 O 2 -sensitive TRPA1 isoforms can be activated by UV via a photochemical transduction cascade and that these isoforms may be exploited as a UV-dependent optogenetic tool for controlling neuronal activities. (20,21) (Fig. 1A).…”

“…The dTRPA1(A)10a and dTRPA1(A)10b isoforms are derived from the A promoter, and both contain a stretch of 97 aa at their N terminus that is absent in dTRPA1(B)10a. The exact physiological function of these A promoter-derived isoforms has not been fully characterized in vivo, but ectopic expression experiments in vitro have shown that both isoforms are sensitive to allyl isothiocyanate (AITC, a major chemical component of wasabi) and that dTRPA1(A)10a, but not dTRPA1(A)10b, can also sense high temperature (>34°C) (21).…”

“…First, a previous report has shown that CC cells showed a clear signal when stained with an antibody against dTRPA1 (18). Second, a GAL4 strain that is expected to report expression of H 2 O 2 -sensitive dTRPA1 isoforms has been shown to be active in CC cells (21). To determine whether the H 2 O 2 -sensitive dTRPA1 isoforms are expressed in CC cells, we performed RT-PCR experiments.…”

“…Indeed, a recent finding has suggested that two specific GPCRs-GUR-3 and LITE-1-both of which play important roles in light sensing in C. elegans, may also be H 2 O 2 sensitive (16). Moreover, TRPA1, an evolutionarily conserved TRP channel known for its role in sensing many chemical irritants, has been shown to sense H 2 O 2 (17)(18)(19)(20)(21)(22)(23)(24), thus TRPA1 may be activated by light via sensing light-induced H 2 O 2 production. Some recent evidence has demonstrated a role for TRPA1 in light sensing.…”