The role of receptor tyrosine kinase RON in the response of cancer cells to cellular stress

Abstract

Epithelial carcinogenesis is a complex multistep process that entails the progressive acquisition of alterations to survive within rigorous microenvironment such as hypoxia nutrient starvation and changes in pH Receptuer d’Origine Nantatise (RON) is a member of the c-Met RTK family Based on its response to starvation stress we hypothesized that RON receptor protein can translocate into nucleus of cancer cells and functions as a stress-responsive regulator for cell survival Experiments in vitro showed that full-length RON translocates into nucleus of TSGH8301 bladder cancer cells as early as 3 hr after hypoxia and is independent of EGFR Transient knock-down of HIF-1alpha did not disturb the nuclear translocation under hypoxia The interaction of nuclear RON with HIF-1alpha could be demonstrated in vivo Domain mapping revealed that tyrosine kinase truncation (RONdeleteTyrK) abolishes the interaction of RON with HIF-1alpha; while transmembrane domain deletion (RONdeleteTM) enhances the interaction of RON with HIF-1alpha and is localized in the nucleus The luciferase promoter assay showed that RON and HIF-1alpha synergistically transactivate the c-JUN promoter Both knockdown and putative binding-site mutation experiments demonstrated that nuclear RON seems more important than HIF-1alpha for activating c-JUN promoter which leads to increased cell proliferation (p <0 001) survival adaptation (p <0 01) and migration (p <0 001) in vitro and tumorigenicity in vivo during hypoxia Co-immunoprecipitation and subcellular fractionation experiments verified the interaction of nuclear RON with Ku70 and DNA-PK after hypoxia for 3 6 12 and 24 h respectively Stable cells with over-expression of RON had a higher survival rate than transmembrane truncated RON after treatment with Epirubicin within two dose ranges (0 56 and 1 125 uM) (p <0 05) Cancer cells with over-expression of RONFL or RONdeleteTM had a better survival in the presence of Epirubicin under hypoxia than that seen under normoxia (p <0 05 respectively) These results suggest that nuclear translocation of RON occurs in response to hypoxia in human bladder cancer cells and is associated with HIF-1alpha Ku70 and DNA-PK leading to activation of c-JUN promoter activation of non- homologous end joining DNA repair Elucidation of the mechanisms underlying nuclear RON may help to develop a novel co-targeting strategy for cancer patients with over-expression of RON

Date of Award	2014 Aug 2
Original language	English
Supervisor	Nan-Haw Chow (Supervisor)