The mechanism of reversion of adenovirus-transformed cells in response to the selection of drug-resistant variants resistant to methylglyoxal-bis-(guanylhydrazone) was studied. The results presented here suggest that there is no direct relationship between development of MGBG resistance and cellular detransformation. All MGBG resistant variants displayed the transformation revertant phenotype in that these cell lines (i) had a flat morphology, (ii) exhibited anchorage-dependence, (iii) did not produce tumors in nude mice and (iv) possessed the viral transforming genes integrated into the cellular genome. The present study deals principally with investigating the basis of drug-resistance and the cause of detransformation in these MGBGR-variants. Drug resistance in these cells may have resulted from two possible events: (a) the target enzyme, SAM-decarboxylase, may be altered leading to development of resistance to MGBG, or (b) the cell has an altered drug transport system. None of the MGBG-resistant variants were altered in the SAM-decarboxylase. By contrast, analysis of the transport mechanism showed a decreased influx of MGBG, and no change in the efflux rate. These results demonstrate that these cells are transport mutants that are defective in MGBG uptake. Detransformation could have been brought about through three different ways: (a) loss or inactivation of the adenovirus transforming genes, (b) mutation in a cellular gene required for transformation and (c) activation of a cellular tumor suppressor gene. Three of the cell lines studied came under category (a) : The G4F cell line had lost the integrated viral transforming genes. The G2a and G5 cell lines had undergone a mutation which prevented the transcription of the viral transforming genes. None of the cell lines exhibited the properties of a cell line expected of those in category (b). Finally, a single cell line, G2 exhibited the properties of a cell line with an activated tumor suppressor gene. This conclusion was based on the following experimental observations: (1) G2 cells expressed a biologically active Ela transforming protein as shown by (i) resistance to retransformation by Ela plasmids, (ii) positive complementation of an Ela deletion mutant, and (iii) Ela-mediated tumor necrosis factor cytolysis, and (2) somatic cell hybridization between the parental transformed F4 cell line and the detransformed G2 cell line resulted in hybrids which were non-tumorigenic. These hybrids had about double the number of chromosomes, carried both parental drug-resistance markers and expressed Ela proteins. Thus, the G2 cell line belongs to the class of recently reported revertants that result from activation of a dominant transformation-suppressor gene(s)
