Figure 1. ERBB3 is present in murine EGFR mutant lung adenocarcinomas and interacts with mutant EGFR.
(A) Immunohistochemical staining for ERBB3 is strong in EGFRL858R-induced lung tumors (right) but weak or non-existent in normal lung from control mice (left). (B) EGFRL858R interacts with ERBB3 in transfected cells. Plasmids encoding human EGFRL858R, mouse ERBB3 or both were transfected into 293T cells. Two days after transfection, an antibody to ERBB3 was used to co-precipitate ERBB3 and proteins associated with it. Immunoprecipitates were blotted with antibodies to EGFR and ERBB3. Results from mock-transfected cells are shown as indicated. (C) Mutant EGFRL858R interacts with ERBB3 in mutant EGFR-induced lung tumors. EGFRL858R or IgG immunoprecipitates from two CCSP-rtTA+;TetO-EGFRL858R+ mice (1 and 2) were immunoblotted with antibodies specific to mutant EGFRL858R and ERBB3. (D) Strategy used to conditionally delete Erbb3 in type II pneumocytes in tetracycline-inducible mouse models of EGFR mutant lung cancer. Control mice (top) express mutant EGFR and wild-type Erbb3 in the lung epithelium upon induction of doxycycline. Experimental mice (bottom) express mutant EGFR but not ERBB3 due to the presence of Cre recombinase, which deletes floxed sequences in the Erbb3 locus. (E) ERBB3 protein is not detected in lungs of mice upon deletion of Erbb3. Immunoblots of lung protein extracts with antibodies to EGFRL858R, ERBB3, Surfactant Protein C (SPC) and Actin are shown. Protein levels were normalized for numbers of pneumocytes as judged by levels of surfactant protein C. EGFRL858R+; Erbb3+ (control mice) and EGFRL858R+; Erbb3− (experimental mice). IP, immunoprecipitation.