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Regulation of membrane ruffling by polarized STIM1 and ORAI1 in cortactin-rich domains

Cell motility and migration requires the reorganization of the cortical cytoskeleton at the leading edge of cells and extracellular Ca(2+) entry is essential for this reorganization. However the molecular nature of the regulators of this pathway is unknown. This work contributes to understanding the...

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Autores principales: Lopez-Guerrero, Aida M., Tomas-Martin, Patricia, Pascual-Caro, Carlos, Macartney, Thomas, Rojas-Fernandez, Alejandro, Ball, Graeme, Alessi, Dario R., Pozo-Guisado, Eulalia, Martin-Romero, Francisco Javier
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Nature Publishing Group UK 2017
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5428229/
https://www.ncbi.nlm.nih.gov/pubmed/28341841
http://dx.doi.org/10.1038/s41598-017-00331-4
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author Lopez-Guerrero, Aida M.
Tomas-Martin, Patricia
Pascual-Caro, Carlos
Macartney, Thomas
Rojas-Fernandez, Alejandro
Ball, Graeme
Alessi, Dario R.
Pozo-Guisado, Eulalia
Martin-Romero, Francisco Javier
author_facet Lopez-Guerrero, Aida M.
Tomas-Martin, Patricia
Pascual-Caro, Carlos
Macartney, Thomas
Rojas-Fernandez, Alejandro
Ball, Graeme
Alessi, Dario R.
Pozo-Guisado, Eulalia
Martin-Romero, Francisco Javier
author_sort Lopez-Guerrero, Aida M.
collection PubMed
description Cell motility and migration requires the reorganization of the cortical cytoskeleton at the leading edge of cells and extracellular Ca(2+) entry is essential for this reorganization. However the molecular nature of the regulators of this pathway is unknown. This work contributes to understanding the role of STIM1 and ORAI1 in the promotion of membrane ruffling by showing that phospho-STIM1 localizes at the leading edge of cells, and that both phospho-STIM1 and ORAI1 co-localize with cortactin (CTTN), a regulator of the cytoskeleton at membrane ruffling areas. STIM1-KO and ORAI1-KO cell lines were generated by CRISPR/Cas9 genome editing in U2OS cells. In both cases, KO cells presented a notable reduction of store-operated Ca(2+) entry (SOCE) that was rescued by expression of STIM1-mCherry and ORAI1-mCherry. These results demonstrated that SOCE regulates membrane ruffling at the leading edge of cells. Moreover, endogenous ORAI1 and overexpressed ORAI1-GFP co-immunoprecipitated with endogenous CTTN. This latter result, in addition to the KO cells’ phenotype, the preservation of ORAI1-CTTN co-localization during ruffling, and the inhibition of membrane ruffling by the Ca(2+)-channel inhibitor SKF96365, further supports a functional link between SOCE and membrane ruffling.
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spelling pubmed-54282292017-05-15 Regulation of membrane ruffling by polarized STIM1 and ORAI1 in cortactin-rich domains Lopez-Guerrero, Aida M. Tomas-Martin, Patricia Pascual-Caro, Carlos Macartney, Thomas Rojas-Fernandez, Alejandro Ball, Graeme Alessi, Dario R. Pozo-Guisado, Eulalia Martin-Romero, Francisco Javier Sci Rep Article Cell motility and migration requires the reorganization of the cortical cytoskeleton at the leading edge of cells and extracellular Ca(2+) entry is essential for this reorganization. However the molecular nature of the regulators of this pathway is unknown. This work contributes to understanding the role of STIM1 and ORAI1 in the promotion of membrane ruffling by showing that phospho-STIM1 localizes at the leading edge of cells, and that both phospho-STIM1 and ORAI1 co-localize with cortactin (CTTN), a regulator of the cytoskeleton at membrane ruffling areas. STIM1-KO and ORAI1-KO cell lines were generated by CRISPR/Cas9 genome editing in U2OS cells. In both cases, KO cells presented a notable reduction of store-operated Ca(2+) entry (SOCE) that was rescued by expression of STIM1-mCherry and ORAI1-mCherry. These results demonstrated that SOCE regulates membrane ruffling at the leading edge of cells. Moreover, endogenous ORAI1 and overexpressed ORAI1-GFP co-immunoprecipitated with endogenous CTTN. This latter result, in addition to the KO cells’ phenotype, the preservation of ORAI1-CTTN co-localization during ruffling, and the inhibition of membrane ruffling by the Ca(2+)-channel inhibitor SKF96365, further supports a functional link between SOCE and membrane ruffling. Nature Publishing Group UK 2017-03-24 /pmc/articles/PMC5428229/ /pubmed/28341841 http://dx.doi.org/10.1038/s41598-017-00331-4 Text en © The Author(s) 2017 This work is licensed under a Creative Commons Attribution 4.0 International License. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in the credit line; if the material is not included under the Creative Commons license, users will need to obtain permission from the license holder to reproduce the material. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/
spellingShingle Article
Lopez-Guerrero, Aida M.
Tomas-Martin, Patricia
Pascual-Caro, Carlos
Macartney, Thomas
Rojas-Fernandez, Alejandro
Ball, Graeme
Alessi, Dario R.
Pozo-Guisado, Eulalia
Martin-Romero, Francisco Javier
Regulation of membrane ruffling by polarized STIM1 and ORAI1 in cortactin-rich domains
title Regulation of membrane ruffling by polarized STIM1 and ORAI1 in cortactin-rich domains
title_full Regulation of membrane ruffling by polarized STIM1 and ORAI1 in cortactin-rich domains
title_fullStr Regulation of membrane ruffling by polarized STIM1 and ORAI1 in cortactin-rich domains
title_full_unstemmed Regulation of membrane ruffling by polarized STIM1 and ORAI1 in cortactin-rich domains
title_short Regulation of membrane ruffling by polarized STIM1 and ORAI1 in cortactin-rich domains
title_sort regulation of membrane ruffling by polarized stim1 and orai1 in cortactin-rich domains
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5428229/
https://www.ncbi.nlm.nih.gov/pubmed/28341841
http://dx.doi.org/10.1038/s41598-017-00331-4
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