Proteomic analysis of a neoplastic mouse lung epithelial cell line whose tumorigenicity has been abrogated by transfection with the gap junction structural gene for connexin 43, Gja1.
In order to examine how tumorigenicity is abrogated by gap junctional intercellular communication (GJIC), protein expression was analyzed in four related mouse lung epithelial cell lines that vary in their GJIC status and neoplastic potential. Since alterations in protein expression underlie neoplastic behavior, this proteomic analysis provides insights into the molecular pathogenesis of lung cancer. E10, an immortalized but non-tumorigenic cell line derived from alveolar type II pneumocytes, has functional GJIC. E9, a spontaneous transformant of E10, is GJIC-deficient and is tumorigenic upon injection into a syngeneic mouse. Stable transfection of E9 with Gja1, the gene for the gap junctional protein, connexin 43, re-established GJIC and rendered this line (designated E9-2) non-tumorigenic; the vector transfection control line, E9-41, remains tumorigenic. Proteins extracted from these cell lines were separated by two-dimensional electrophoresis (2DE) and visualized by Coomassie blue staining. We consistently observed differential expression of 27 proteins between E10 and E9 and identified 11 of these by peptide mass mapping. The functions of these proteins include stress response, cytoskeletal structure, signal transduction, apoptosis, immune response, pre-mRNA processing, and carbohydrate metabolism. Gja1 transfection affected the concentrations of four of these proteins, viz. PDI, alpha-enolase, aldolase A, and gelsolin-like protein. PDI concentration was most profoundly affected; E10 cells contain twice as much PDI as E9, and PDI was restored to E10-like levels in the E9-2 transfectant line while remaining at E9-like levels in the vector control E9-41 cells. An association between connexin 43 and PDI expression was also observed in a second set of independently derived type II cell lines. The PDI superfamily has multiple cellular roles including chaperoning assembled glycoproteins, regulating the activities of transcription factors, and regulating disulfide bond formation.
Peebles
Katherine A
KA
Department of Pharmaceutical Sciences, University of Colorado Health Sciences Center, 4200 East Ninth Avenue, Denver 80262, USA.
Duncan
Mark W
MW
Ruch
Randall J
RJ
Malkinson
Alvin M
AM
D000818:Animals; D018031:Connexin 43 / Q000235:genetics*; D004847:Epithelial Cells / Q000473:pathology; D005634:Fructose-Bisphosphate Aldolase / Q000378:metabolism; D017629:Gap Junctions / Q000235:genetics*; D018260:Gelsolin / Q000378:metabolism; D005796:Genes; D008175:Lung Neoplasms / Q000201:enzymology / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D051379:Mice; D010751:Phosphopyruvate Hydratase / Q000378:metabolism; D040901:Proteomics; D014162:Transfection; D014407:Tumor Cells, Cultured
D018031:Connexin 43; D018260:Gelsolin; D005634:Fructose-Bisphosphate Aldolase; D010751:Phosphopyruvate Hydratase