Cloning of TPA-inducible early (TIE) genes by differential hybridization using TPA-nonresponsive variant of mouse 3T3-L1 cells.

The tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) induces DNA synthesis in quiescent 3T3-L1 cells but not in its variant VT-1 cells. A lambda gt10 cDNA library was constructed using poly(A)+ RNA from 3T3-L1 cells that were stimulated by TPA for 20 min. Radioactive cDNA probes were prepared from mRNAs of TPA-treated 3T3-L1 and VT-1 cells and used for screening of the 3T3-L1 cDNA library by differential hybridization. Nine of 6000 phase plaques hybridized only to the 3T3-L1 cDNA probe. Analysis of the nucleotide sequence of five of these clones indicated a high degree of homology with human or mouse type I and type III collagen genes. Three other independent clones showed no homology with any known DNA sequences. These isolated clones of TPA-inducible early (TIE) genes may be useful to study the signal transduction pathway of phorbol esters.