Oncogenic RAS isoforms show a hierarchical requirement for the guanine nucleotide exchange factor SOS2 to mediate cell transformation

Abstract

About a third of tumors have activating mutations in <i>HRAS</i>, <i>NRAS</i>, or <i>KRAS</i>, genes encoding guanosine triphosphatases (GTPases) of the RAS family. In these tumors, wild-type RAS cooperates with mutant RAS to promote downstream effector activation and cell proliferation and transformation, suggesting that upstream activators of wild-type RAS are important modulators of mutant RAS-driven oncogenesis. The guanine nucleotide exchange factor (GEF) SOS1 mediates KRAS-driven proliferation, but little is understood about the role of SOS2. We found that RAS family members have a hierarchical requirement for the expression and activity of SOS2 to drive cellular transformation. In mouse embryonic fibroblasts (MEFs), SOS2 critically mediated mutant KRAS-driven, but not HRAS-driven, transformation. <i>Sos2</i> deletion reduced epidermal growth factor (EGF)-dependent activation of wild-type HRAS and phosphorylation of the kinase AKT in cells expressing mutant RAS isoforms. Assays using pharmacological inhibitors revealed a hierarchical requirement for signaling by phosphoinositide 3-kinase (PI3K) in promoting RAS-driven cellular transformation that mirrored the requirement for SOS2. KRAS-driven transformation required the GEF activity of SOS2 and was restored in <i>Sos2</i>^<i>-/-</i> MEFs by expression of constitutively activated PI3K. Finally, CRISPR/Cas9-mediated deletion of <i>SOS2</i> reduced EGF-stimulated AKT phosphorylation and synergized with MEK inhibition to revert the transformed phenotype of human <i>KRAS</i> mutant pancreatic and lung tumor cells. These results indicate that SOS2-dependent PI3K signaling mediates mutant KRAS-driven transformation, revealing therapeutic targets in KRAS-driven cancers. Our data also reveal the importance of three-dimensional culture systems in investigating the mediators of mutant KRAS.