Cellular context determines primary characteristics of human TRPC5 as a cold‐activated channel

• Published in: Journal of cellular physiology 2022-09, Vol.237 (9), p.3614-3626
• Main Authors: Ptakova, Alexandra, Mitro, Michal, Zimova, Lucie, Vlachova, Viktorie
• Format: Article
• Language: English
• Subjects: Calcium; Calcium (intracellular); Calcium (reticular); Calcium homeostasis; Calcium permeability; Channel gating; Cold; cold temperature; Dwell time; Electrostatic properties; Endoplasmic reticulum; Enthalpy; Entropy; HEK293T; Homeostasis; Inflammation; Ion channels; Mutants; Nervous system; Odontoblasts; Pain perception; Peripheral nervous system; Phosphorylation; Proteins; Receptors; Room temperature; Sensitivity; single channels; STIM1 protein; stromal interaction molecule 1; Synovium; Temperature dependence; Temperature gradients; thermosensitive TRP; Transient receptor potential proteins; TRPC cation channels
• ISSN: 0021-9541; 1097-4652
• DOI: 10.1002/jcp.30821

The human transient receptor potential canonical 5 (TRPC5) is a calcium‐permeable, nonselective cation channel expressed in the central and peripheral nervous system and also in other tissues such as the kidney, synovium, and odontoblasts. TRPC5 has been recently confirmed to play a key role in spontaneous, inflammatory mechanical, and cold pain. Although TRPC5 activation is known to be cold sensitive, it is unclear whether this property is intrinsic to the channel protein and whether or to what extent it may be determined by the cellular environment. In this study, we explored the cold sensitivity of human TRPC5 at the single‐channel level using transiently transfected HEK293T cells. Upon decreasing the temperature, the channel demonstrated prolonged mean open dwell times and a robust increase in the open probability (Po), whereas the amplitude of unitary currents decreased ~1.5‐fold per 10°C of temperature difference. In the absence of any agonists, the temperature dependence of Po was sigmoidal, with a steep slope within the temperature range of 16°C–11°C, and exhibited saturation below 8–5°C. Thermodynamic analysis revealed significant changes in enthalpy and entropy, suggesting that substantial conformational changes accompany cold‐induced gating. The mutant channel T970A, in which the regulation downstream of G‐protein coupled receptor signaling was abrogated, exhibited higher basal activity at room temperature and a less steep temperature response profile, with an apparent threshold below 22°C. An even more pronounced decrease in the activation threshold was observed in a mutant that disrupted the electrostatic interaction of TRPC5 with the endoplasmic reticulum calcium sensor stromal interaction molecule 1. Thus, TRPC5 exhibits features of an intrinsically cold‐gated channel; its sensitivity to cold tightly depends on the phosphorylation status of the protein and intracellular calcium homeostasis. The human transient receptor potential canonical 5 (TRPC5) can be activated by cold in intact cells. The cold‐induced activity depends on the electrostatic interaction between TRPC5 (residues D652/E653) with the cytoplasmic C terminus of stromal interaction molecule 1 (STIM1), which stabilizes the channel activity through the K‐domain over the whole tested temperature range (25–5°C). Upon cooling, STIM1 stabilizes the closed conformation of the channel.