CONDITIONAL TRANSFORMATION OF A PANCREATIC BETA-CELL LINE DERIVED FROM TRANSGENIC MICE EXPRESSING A TETRACYCLINE-REGULATED ONCOGENE

Conditional oncogene expression in transgenic mice is of interest for studying the oncoprotein requirements during tumorigenesis and for der iving cell lines that can be induced to undergo growth arrest and enha nce their differentiated functions. We utilized the bacterial tetracyc line (Tet)-resistance operon regulatory system (tet) from Tn10 of Esch erichia coil to control simian virus 40 (SV40) large tumor (T) antigen (TAg) gene expression and to generate conditionally transformed pancr eatic beta cells in transgenic mice. A fusion protein containing the t et repressor (tetR) and the activating domain of the herpes simplex vi rus protein VP16, which converts the repressor into a transcription ac tivator, was produced in beta cells of transgenic mice under control o f the insulin promoter. In a separate lineage of transgenic mice, the TAg gene was introduced under control of a tandem array of tet operato r sequences and a minimal promoter, which by itself is not sufficient for gene expression. Mice from the two lineages were then crossed to g enerate double-transgenic mice. Expression of the tetR fusion protein in beta cells activated TAg transcription, resulting in the developmen t of beta-cell tumors, Tumors arising in the absence of Tet were cultu red to derive a stable beta-cell line. Cell incubation in the presence of Tet led to inhibition of proliferation, as shown by decreased BrdU rd and [H-3]thymidine incorporation. The Tet derivative anhydrotetracy cline showed a 100-fold stronger inhibition compared with Tet, When ad ministered in vivo, Tet efficiently inhibited beta-cell proliferation. These findings indicate that transformed beta cells selected for grow th during a tumorigenesis process in vivo maintain a dependence on the continuous presence of the TAg oncoprotein for their proliferation, T his system provides an approach for generation of beta-cell lines for cell therapy of diabetes as well als conditionally transformed cell li nes from other cell types of interest.