Reorganización del citoesqueleto regulada por STIM1 y ORAI1

Laburpena

In this Doctoral Thesis we have described the role of STIM1 and ORAI1, the two major components of SOCE, in the promotion of membrane ruffling required for the reorganization of the cortical cytoskeleton at the leading edge of migrating cells. Previously to the cytoskeletal study we described the interdependence of ERK1/2 and store operated calcium entry (SOCE). In this regard, our results revealed that the activation of ERK1/2 is STIM1- and SOCE-independent. Regarding to the study of the cytoskeleton, in the present work we described that phospho-STIM1 localizes at the leading-edge of cells, and that both phospho-STIM1 and ORAI1 co-localizes with cortactin (CTTN), a regulator of the cytoskeleton at membrane ruffling areas. STIM1-KO and ORAI1-KO cell lines were generated by CRISPR/Cas9 system in U2OS cells. In both cases, KO cells presented a notable reduction of SOCE, together with a striking decrease of the membrane ruffling, compared with U2OS wild-type cells. These results demonstrated that SOCE regulates membrane ruffling at the leading edge of cells. Moreover, we showed that ORA1 co-immunoprecipitated with CTTN. This latter result, in addition to the KO cells phenotype and the preservation of ORAI1-CTTN co-localization during ruffling, further support a functional link between SOCE and membrane ruffling.