Regulation of Membrane Ruffling by Polarized STIM1 and ORAI1 in Cortactin-rich Domains

Overview

Journal Sci Rep

Specialty Science

Date 2017 Mar 26

PMID 28341841

Citations 10

Authors

Aida M Lopez-Guerrero

Patricia Tomas-Martin

Carlos Pascual-Caro

Thomas Macartney

Alejandro Rojas-Fernandez

Graeme Ball

Dario R Alessi

Eulalia Pozo-Guisado

Francisco Javier Martin-Romero

Affiliations

Soon will be listed here.

Abstract

Cell motility and migration requires the reorganization of the cortical cytoskeleton at the leading edge of cells and extracellular Ca 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 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-channel inhibitor SKF96365, further supports a functional link between SOCE and membrane ruffling.

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