Evidence that Orai1 does not contribute to store-operated TRPC1 channels in vascular smooth muscle cells

• Autores: Shi, Jian; Miralles, Francesc; Kinet, Jean Pierre; Birnbaumer, Lutz; Large, William A.; Albert, Anthony P.
• Año de publicación: 2017
• Idioma: inglés
• Tipo de recurso: artículo
• Materia: ORAI1; PLC; STIM1; STORE-OPERATED; TRPC1; VASCULAR SMOOTH MUSCLE
• Institución: Consejo Nacional de Investigaciones Científicas y Técnicas

Ca2+-permeable store-operated channels (SOCs) mediate Ca2+ entry pathways which are involved in many cellular functions such as contraction, growth, and proliferation. Prototypical SOCs are formed of Orai1 proteins and are activated by the endo/sarcoplasmic reticulum Ca2+ sensor stromal interaction molecule 1 (STIM1). There is considerable debate about whether canonical transient receptor potential 1 (TRPC1) proteins also form store-operated channels (SOCs), and if they do, is Orai1 involved. We recently showed that stimulation of TRPC1-based SOCs involves store depletion inducing STIM1-evoked Gαq/PLCβ1 activity in contractile vascular smooth muscle cells (VSMCs). Therefore the present work investigates the role of Orai1 in activation of TRPC1-based SOCs in freshly isolated mesenteric artery VSMCs from wild-type (WT) and Orai1−/− mice. Store-operated whole-cell and single channel currents recorded from WT and Orai1−/− VSMCs had similar properties, with relatively linear current-voltage relationships, reversal potentials of about +20mV, unitary conductances of about 2pS, and inhibition by anti-TRPC1 and anti-STIM1 antibodies. In Orai1−/− VSMCs, store depletion induced PLCβ1 activity measured with the fluorescent phosphatidylinositol 4,5-bisphosphate/inositol 1,4,5-trisphosphate biosensor GFP-PLCδ1-PH, which was prevented by knockdown of STIM1. In addition, in Orai1−/− VSMCs, store depletion induced translocation of STIM1 from within the cell to the plasma membrane where it formed STIM1-TRPC1 interactions at discrete puncta-like sites. These findings indicate that activation of TRPC1-based SOCs through a STIM1-activated PLCβ1 pathway are likely to occur independently of Orai1 proteins, providing evidence that TRPC1 channels form genuine SOCs in VSMCs with a contractile phenotype.
Fil: Shi, Jian. University of Leeds; Reino Unido
Fil: Miralles, Francesc. University of London; Reino Unido
Fil: Kinet, Jean Pierre. Harvard Medical School; Estados Unidos
Fil: Birnbaumer, Lutz. Pontificia Universidad Católica Argentina ; Argentina
Fil: Large, William A.. University of London; Reino Unido
Fil: Albert, Anthony P.. University of London; Reino Unido