[en] [ENUsing a 4-hydroxytamoxifen (4OHT)-inducible, conditional Sos1-null mutation, we analyzed wild-type (WT), single Sos1-KO, Sos2-KO and double Sos1/2 KO primary mouse embryonic fibroblasts (MEF) with an aim at evaluating the functional specificity or redundancy of the Sos1 and Sos2 alleles at the cellular level. The 4OHT-induced Sos1-KO and Sos1/2-DKO MEFs exhibited distinct flat morphology, enlarged cell perimeter and altered cytoskeletal organization that were not observed in the WT and Sos2-KO counterparts. The Sos1-KO and Sos1/2-DKO MEFs also displayed significant accumulation, in comparison with WT and Sos2-KO MEFs, of cytoplasmic vesicular bodies identified as autophagosomes containing degraded mitochondria by means of electron microscopy and specific markers. Cellular proliferation and migration were impaired in Sos1-KO and Sos1/2-DKO MEFs in comparison with WT and Sos2-KO MEFs, whereas cell adhesion was only impaired upon depletion of both Sos isoforms. RasGTP formation was practically absent in Sos1/2-DKO MEFs as compared with the other genotypes and extracellular signal-regulated kinase phosphorylation showed only significant reduction after combined Sos1/2 depletion. Consistent with a mitophagic phenotype, in vivo labeling with specific fluorophores uncovered increased levels of oxidative stress (elevated intracellular reactive oxygen species and mitochondrial superoxide and loss of mitochondrial membrane potential) in the Sos1-KO and the Sos1/2-DKO cells as compared with Sos2-KO and WT MEFs. Interestingly, treatment of the MEF cultures with antioxidants corrected the altered phenotypes of Sos1-KO and Sos1/2-DKO MEFs by restoring their altered perimeter size and proliferative rate to levels similar to those of WT and Sos2-KO MEFs. Our data uncover a direct mechanistic link between Sos1 and control of intracellular oxidative stress, and demonstrate functional prevalence of Sos1 over Sos2 with regards to cellular proliferation and viability.

[es] Utilizando un mutante condicional de Sos1 inducible por 4-hidroxitamoxifeno (4OHT), analizamos fibroblastos embrionarios de rat?n primarios (MEF) de tipo salvaje (WT), Sos1-KO simple, Sos2-KO y doble Sos1/2 KO con el objetivo de para evaluar la especificidad funcional o redundancia de los alelos Sos1 y Sos2 a nivel celular. Los MEF Sos1-KO y Sos1/2-DKO inducidos por 4OHT una morfolog?a plana distinta, un per?metro celular ampliado y una organizaci?n del citoesqueleto alterada que no se observaron en las c?lulas WT y Sos2-KO. Los MEF Sos1-KO y Sos1/2-DKO tambi?n mostraron una acumulaci?n significativa, en comparaci?n con los MEF WT y Sos2-KO, de cuerpos vesiculares citoplasm?ticos identificados mediante microscop?a electr?nica como autofagosomas que contienen mitocondrias degradadas. Tambi?n la proliferaci?n y migraci?n celular se vieron afectadas en los MEF Sos1-KO y Sos1/2-DKO en comparaci?n con los MEF WT y Sos2-KO, mientras que la adhesi?n celular solo se vio afectada tras la eliminaci?n de ambas isoformas de Sos. La formaci?n de RasGTP estuvo pr?cticamente ausente en los MEF Sos1/2-DKO en comparaci?n con los otros genotipos y la fosforilaci?n de ERK mostr? solo una reducci?n significativa despu?s de la eliminaci?n combinada de Sos1 y Sos2. Un marcaje in vivo con fluor?foros espec?ficos descubri? niveles elevados de estr?s oxidativo (especies reactivas de ox?geno intracelular elevadas y super?xido mitocondrial y p?rdida del potencial de membrana mitocondrial) en las c?lulas Sos1-KO y Sos1/2-DKO en comparaci?n con Sos2-KO y WT MEF. Curiosamente, el tratamiento de los cultivos de MEFs con antioxidantes corrigi? los fenotipos alterados de los MEF Sos1-KO y Sos1/2-DKO al restaurar su tama?o y su tasa proliferativa a niveles similares a los de los MEFs WT y Sos2-KO. Nuestros datos descubren un v?nculo mec?nico directo entre Sos1 y el control del estr?s oxidativo intracelular, y demuestran la prevalencia funcional de Sos1 sobre Sos2 con respecto a la proliferaci?n y viabilidad celular.