Phosphoinositide-interacting regulator of TRP (PIRT) has opposing effects on human and mouse TRPM8 ion channels

Jacob K. Hilton, Taraneh Salehpour, Nicholas J. Sisco, Parthasarathi Rath, Wade D. Van Horn

Research output: Contribution to journalArticlepeer-review

8 Scopus citations


Transient receptor potential melastatin 8 (TRPM8) is a coldsensitive ion channel with diverse physiological roles. TRPM8 activity is modulated by many mechanisms, including an interaction with the small membrane protein phosphoinositide-interacting regulator of TRP (PIRT). Here, using comparative electrophysiology experiments, we identified species-dependent differences between the human and mouse TRPM8?PIRT complexes. We found that human PIRT attenuated human TPRM8 conductance, unlike mouse PIRT, which enhanced mouse TRPM8 conductance. Quantitative Western blot analysis demonstrates that this effect does not arise from decreased trafficking of TRPM8 to the plasma membrane. Chimeric human/mouse TRPM8 channels were generated to probe the molecular basis of the PIRT modulation, and the effect was recapitulated in a pore domain chimera, demonstrating the importance of this region for PIRT-mediated regulation of TRPM8. Moreover, recombinantly expressed and purified human TRPM8 S1?S4 domain (comprising transmembrane helices S1?S4, also known asthe sensing domain, ligand-sensing domain, or voltage sensing-like domain) and full-length human PIRT were used to investigate binding between the proteins. NMR experiments, supported by a pulldown assay, indicated that PIRT binds directly and specifically to the TRPM8 S1?S4 domain. Binding became saturated as the S1?S4:PIRT mole ratio approached 1. Our results have uncovered species-specific TRPM8 modulation by PIRT. They provide evidence for a direct interaction between PIRT and the TRPM8 S1?S4 domain with a 1:1 binding stoichiometry, suggesting that a functional tetrameric TRPM8 channel has four PIRT-binding sites.

Original languageEnglish (US)
Pages (from-to)9423-9434
Number of pages12
JournalJournal of Biological Chemistry
Issue number24
StatePublished - Jun 15 2018

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology


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