Phosphorylated TRPV1 and ANO1 ⁄ TMEM16A interaction induced by low concentration of capsaicin or innocuous heat stimulation

抄録

Transient receptor potential vanilloid 1 (TRPV1), a capsaicin receptor, and anoctamin 1 (ANO1, also called TMEM16A), a calcium–activated chloride channel, are major ion channels involved in pain sensation in the periph­eral nervous system. Pain–related behaviors dependent on each ion channel are reported­ly reduced in its deficient mice. We previously found that TRPV1 and ANO1 interact with each other upon making a physical complex, and the functional linkage exacerbates capsaicin–induced acute pain sensation. However, the significance of TRPV1 and ANO1 interaction in the inflammatory condition remains unknown. Activation thresholds of TRPV1 become low upon its phosphorylation. Here, we performed whole–cell patch–clamp recordings using phorbol 12–myristate 13–acetate, an activator of protein kinase C to phosphorylate TRPV1, mimicking the inflam­matory conditions in HEK293T cells express­ing mouse TRPV1 and mouse ANO1. We also showed that phosphorylated TRPV1 interacts with ANO1 with a low concentration of capsaicin or innocuous heat stimulation of approxi­mately 37°C. Furthermore, we performed immunoprecipitation to investigate whether TRPV1–ANO1 inter­action is enhanced by phosphorylation. However, the protein–protein interaction was not changed. Thus, ANO1 activa­tion could be enhanced by the acceleration of TRPV1 activity. These facts indicate that interactions between phosphorylated TRPV1 and ANO1 could explain inflammatory pain sensations, for instance, in heat allodynia. Therefore, our findings contribute to clarifying the new molecular mechanisms involved in pathological pain and development of analgesia.