Oncogenic RAS signaling leads to neoplastic transformation in immortalized cells and oncogene induced senescence in primary cells. Our laboratory has found that in both primary and immortalized cells oncogenic RAS induces constitutive perinuclear re-localization of p-ERK, CK2, and the MAPK scaffolding protein, KSR1. KSR1 binds to p-ERK and CK2, forming assemblies that we call “perinuclear signaling complexes” (PSCs). We observe PSCs in all cancer cell lines tested and in KRasG12V-driven mouse lung tumors, indicating that PSC formation is an important and integral characteristic of tumor cells. Our results show that KSR1 is essential for PSCs, as RNAi-mediated depletion of KSR1 or knockout of KSR1 disrupted their formation. We also demonstrated a key role for endosomal trafficking in this localized RAS signaling, as pharmacological inhibition of endocytosis prevented PSC formation. Furthermore, Rab11A, a Rab GTPase involved in trafficking of recycling endosomes, is required for establishment of PSCs and co-localizes with a pool of cellular CK2 and KSR1, but not p-ERK. Moreover, Rab11A knockdown caused decreased proliferation and survival of A549 cells (human KRAS mutant non-small cell lung adenocarcinoma). Our results demonstrate that there may be at least two classes of PSC-containing endosomes, p-ERK bound endosomes and CK2 bound endosomes. We are currently attempting to identify Rab GTPases that may differentiate these two kinds of PSC containing endosomes. Jongsma et al. (Cell, 2016) have shown that endosomes associated with the ubiquitin adaptor proteins EPS15, Tollip or TAXBP1 localize to the perinuclear region via the signaling adaptor, SQSTM1 (p62), which is tethered to the ER. We found that Tollip knockdown in A549 cells disrupted the CK2 bound endosomes but not p-ERK bound endosomes. Tollip knockdown also decreased cell proliferation and cell survival, while increasing apoptosis and autophagy. Whether other adaptor proteins are involved in RAS-mediated PSC formation remains to be determined. Growth factors (GF) also transiently induced PSCs in normal cells with delayed kinetics (4-6 hr). Our findings indicate that transformed cells sustain this novel late phase of GF signaling, localizing RAS pathway kinases to a nuclear-proximal compartment to drive tumorigenesis. We propose that PSCs are hallmarks of cancer cells and may represent useful biomarkers for cancer diagnosis and therapeutic responses.
Citation Format: Srikanta Basu, Sandip K. Basu, Jacqueline Salotti, Peter F. Johnson. Role of Rab GTPases and endosomal adaptor proteins in oncogenic RAS induced formation of perinuclear signaling complexes (PSCs) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4361.