Direct evidence for functional TRPV1/TRPA1 heteromers

Area of Science:

• Neuroscience
• Molecular Biology
• Biophysics

Background:

• Transient Receptor Potential Cation Channel, subfamily V, member 1 (TRPV1) is crucial for sensing environmental hazards and pain.
• TRPV1 and Transient Receptor Potential Cation Channel, subfamily A, member 1 (TRPA1) often coexist in cells, with evidence suggesting potential interactions.
• A TRPV1-TRPA1 heteromeric channel has been hypothesized but lacked direct structural and functional evidence.

Purpose of the Study:

• To investigate direct evidence for TRPV1 and TRPA1 interaction and heteromeric channel formation.
• To characterize the functional properties and subunit stoichiometry of TRPV1-TRPA1 channels.
• To determine the functional consequences of TRPA1 co-assembly on TRPV1 channel activity.

Main Methods:

• Construction and functional characterization of TRPV1::TRPV1 and TRPV1::TRPA1 subunit concatemers.
• Atomic Force Microscopy (AFM) for molecular volume and structural analysis.
• Antibody labeling to determine subunit arrangement and stoichiometry.

Main Results:

• TRPV1::TRPV1 concatemers exhibited properties similar to native TRPV1 channels.
• TRPV1::TRPA1 concatemers formed functional channels activated by TRPV1 agonists and heat, but not TRPA1 agonists.
• AFM and antibody labeling indicated TRPV1::TRPA1 forms a heterotetramer with a distinct subunit arrangement compared to TRPV1 homotetramers.
• TRPV1::TRPA1 channels displayed reduced capsaicin binding sites and current amplitude compared to TRPV1.
• TRPA1 co-assembly led to functional inhibition of TRPV1 activity, evidenced by altered gating and sensitization by PKC.

Conclusions:

• Direct evidence supports the formation of TRPV1-TRPA1 heteromeric channels.
• The stoichiometry and arrangement of TRPV1::TRPA1 channels differ from TRPV1 homotetramers.
• TRPA1 incorporation into TRPV1 channels results in functional inhibition of TRPV1 activity, impacting channel gating and ligand sensitivity.
• These findings reveal a novel mechanism of TRPV1 channel regulation by TRPA1 in pain and sensory pathways.