Shi, J; Miralles, F; Birnbaumer, L; Large, WA; Albert, AP
(2017)
Store-operated interactions between plasmalemmal STIM1 and TRPC1 proteins stimulate PLCβ1 to induce TRPC1 channel activation in vascular smooth muscle cells.
J Physiol, 595 (4).
pp. 1039-1058.
ISSN 1469-7793
https://doi.org/10.1113/JP273302
SGUL Authors: Albert, Anthony Paul Miralles Arenas, Francisco
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Abstract
KEY POINTS: Depletion of Ca(2+) stores activates store-operated channels (SOCs), which mediate Ca(2+) entry pathways that regulate cellular processes such as contraction, proliferation and gene expression. In vascular smooth muscle cells (VSMCs), stimulation of SOCs composed of canonical transient receptor potential channel 1 (TRPC1) proteins requires G protein α q subunit (Gαq)/phospholipase C (PLC)β1/protein kinase C (PKC) activity. We studied the role of stromal interaction molecule 1 (STIM1) in coupling store depletion to this activation pathway using patch clamp recording, GFP-PLCδ1-PH imaging and co-localization techniques. Store-operated TRPC1 channel and PLCβ1 activities were inhibited by STIM1 short hairpin RNA (shRNA) and absent in TRPC1(-/-) cells, and store-operated PKC phosphorylation of TRPC1 was inhibited by STIM1 shRNA. Store depletion induced interactions between STIM1 and TRPC1, Gαq and PLCβ1, which required STIM1 and TRPC1. Similar effects were produced with noradrenaline. These findings identify a new activation mechanism of TRPC1-based SOCs in VSMCs, and a novel role for STIM1, where store-operated STIM1-TRPC1 interactions stimulate Gαq/PLCβ1/PKC activity to induce channel gating. ABSTRACT: In vascular smooth muscle cells (VSMCs), stimulation of canonical transient receptor potential channel 1 (TRPC1) protein-based store-operated channels (SOCs) mediates Ca(2+) entry pathways that regulate contractility, proliferation and migration. It is therefore important to understand how these channels are activated. Studies have shown that stimulation of TRPC1-based SOCs requires G protein α q subunit (Gαq)/phospholipase C (PLC)β1 activities and protein kinase C (PKC) phosphorylation, although it is unclear how store depletion stimulates this gating pathway. The present study examines this issue by focusing on the role of stromal interaction molecule 1 (STIM1), an endo/sarcoplasmic reticulum Ca(2+) sensor. Store-operated TRPC1 channel activity was inhibited by TRPC1 and STIM1 antibodies and STIM1 short hairpin RNA (shRNA) in wild-type VSMCs, and was absent in TRPC1(-/-) VSMCs. Store-operated PKC phosphorylation of TRPC1 was reduced by knockdown of STIM1. Moreover, store-operated PLCβ1 activity measured with the fluorescent phosphatidylinositol 4,5-bisphosphate/inositol 1,4,5-trisphosphate biosensor GFP-PLCδ1-PH was reduced by STIM1 shRNA and absent in TRPC1(-/-) cells. Immunocytochemistry, co-immunoprecipitation and proximity ligation assays revealed that store depletion activated STIM1 translocation from within the cell to the plasma membrane (PM) where it formed STIM1-TRPC1 complexes, which then associated with Gαq and PLCβ1. Noradrenaline also evoked TRPC1 channel activity and associations between TRPC1, STIM1, Gαq and PLCβ1, which were inhibited by STIM1 knockdown. Effects of N-terminal and C-terminal STIM1 antibodies on TRPC1-based SOCs and STIM1 staining suggest that channel activation may involve insertion of STIM1 into the PM. The findings of the present study identify a new activation mechanism of TRPC1-based SOCs in VSMCs, and a novel role for STIM1, in which store-operated STIM1-TRPC1 interactions stimulate PLCβ1 activity to induce PKC phosphorylation of TRPC1 and channel gating.
Item Type: | Article | ||||||||||||
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Additional Information: | © 2016 The Authors. The Journal of Physiology published by John Wiley & Sons Ltd on behalf of The Physiological Society This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited. | ||||||||||||
Keywords: | PLC, STIM1, TRPC, vascular smooth muscle, Physiology, 06 Biological Sciences, 11 Medical And Health Sciences | ||||||||||||
SGUL Research Institute / Research Centre: | Academic Structure > Institute of Medical & Biomedical Education (IMBE) Academic Structure > Institute of Medical & Biomedical Education (IMBE) > Centre for Biomedical Education (INMEBE) Academic Structure > Molecular and Clinical Sciences Research Institute (MCS) Academic Structure > Molecular and Clinical Sciences Research Institute (MCS) > Vascular (INCCVA) |
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Journal or Publication Title: | J Physiol | ||||||||||||
ISSN: | 1469-7793 | ||||||||||||
Language: | eng | ||||||||||||
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Publisher License: | Creative Commons: Attribution 4.0 | ||||||||||||
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PubMed ID: | 27753095 | ||||||||||||
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URI: | https://openaccess.sgul.ac.uk/id/eprint/108322 | ||||||||||||
Publisher's version: | https://doi.org/10.1113/JP273302 |
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