TPA-inducible proteins may account for sensitivity to promotion of transformation

The preneoplastic JB6 mouse epidermal cell system includes cell lines sensitive (P+) or resistant (P-) to tumor promoter induced neoplastic transformation. The authors investigated whether a difference in TPA-inducible proteins may explain this differential sensitivity. The synthesis of a 39 Kd cytoplasmic protein (Major Excreted Protein) was TPA-inducible, but to a similar extent in both P+ and P- cells. TPA stimulated phosphorylation but not synthesis of the previously described stress protein pp80 in both P+ and P- cells from 1 to 5 hr after treatment. Pulse labelling of P+ and P- cells with 35S-methionine revealed a TPA dependent P+ specific transient increase in the synthesis of 58Kd protein. Induction was observed at 1 hr, and returned to basal levels by 4 hr and 20 hr, in nuclear and cytoplasmic fractions, respectively. This protein is not phosphorylated in response to TPA treatment. P+ cells differ from P- cells in one or more genes that specify sensitivity to promotion of transformation, designated pro genes. Antibodies to three peptides representing the pro-1 open reading frame were used in immunoprecipitation and Western blotting to isolate the pro-1 gene product. A 43 Kd protein was immunologically responsive to the pro-1 peptide antibodies, and showed an increased signal 40 min after TPA treatment. Since the predicted molecular weight of a pro-1 gene product is only 7 Kd, the possibility of a modification of the protein by poly(ADP-ribosylation) or glycosylation is being investigated