Orai1α, but not Orai1β, co-localizes with TRPC1 and is required for its plasma membrane location and activation in HeLa cells

• Published in: Cellular and molecular life sciences : CMLS Vol. 79; no. 1; p. 33
• Main Authors: Sanchez-Collado, Jose, Lopez, Jose J., Jardin, Isaac, Berna-Erro, Alejandro, Camello, Pedro J., Cantonero, Carlos, Smani, Tarik, Salido, Gines M., Rosado, Juan A.
• Format: Journal Article
• Language: English
• Published: Cham Springer International Publishing 01.01.2022; Springer Nature B.V
• Subjects: Agonists; Amino acids; Antibodies; Biochemistry; Biomarkers; Biomedical and Life Sciences; Biomedicine; Biophysics; Calcium - metabolism; Calcium channels; Calcium influx; Calcium ions; Calcium signalling; Cations; Cell activation; Cell Biology; Cell Membrane - metabolism; Cloning; Depletion; Fluorescence; HeLa Cells; Humans; Immunoprecipitation; Inactivation; Life Sciences; Mammalian cells; Membranes; Mutant Proteins - metabolism; Orai1 protein; ORAI1 Protein - metabolism; Original; Original Article; Oscillations; Physiology; Plasma; Plasmids; Pore formation; Protein Binding; Proteins; Reagents; Smooth muscle; Stromal Interaction Molecule 1 - metabolism; TRPC Cation Channels - metabolism; Orai1α; STIM1; TRPC1; Store-operated calcium entry; Orai1β
• ISSN: 1420-682X; 1420-9071
• DOI: 10.1007/s00018-021-04098-w

The identification of two variants of the canonical pore-forming subunit of the Ca 2+ release-activated Ca 2+ (CRAC) channel Orai1, Orai1α and Orai1β, in mammalian cells arises the question whether they exhibit different functional characteristics. Orai1α and Orai1β differ in the N-terminal 63 amino acids, exclusive of Orai1α, and show different sensitivities to Ca 2+ -dependent inactivation, as well as distinct ability to form arachidonate-regulated channels. We have evaluated the role of both Orai1 variants in the activation of TRPC1 in HeLa cells. We found that Orai1α and Orai1β are required for the maintenance of regenerative Ca 2+ oscillations, while TRPC1 plays a role in agonist-induced Ca 2+ influx but is not essential for Ca 2+ oscillations. Using APEX2 proximity labeling, co-immunoprecipitation and the fluorescence of G-GECO1.2 fused to Orai1α our results indicate that agonist stimulation and Ca 2+ store depletion enhance Orai1α–TRPC1 interaction. Orai1α is essential for TRPC1 plasma membrane location and activation. Thus, TRPC1 function in HeLa cells depends on Ca 2+ influx through Orai1α exclusively.