Expression of the human retinoblastoma gene product in mouse fibroblasts: Effects on cell proliferation and susceptibility to transformation

Expression of the human retinoblastoma gene (RB1) in tumor cells defective of the gene in many instances abrogates the growth of the cells. Here we have evaluated the characteristics and tumor-suppressive functions of the human retinoblastoma gene product in mouse fibroblasts. Human full-length wild-type or mutant RB cDNAs were transfected into NIH 3T3 cells and cell clones expressing high levels of RB protein were isolated and characterized. Stable expression of RB protein was obtained, and cell growth experiments indicated that the human RB expressing clones maintained unaltered growth rates under normal culture conditions. The growth rates of wild-type RB-expressing clones but not those expressing mutant RB were reduced in lower serum concentrations. This indicates that serum withdrawal may bring out some growth suppressive properties of RB. Analysis of the human RB protein produced by mouse fibroblasts by cell synchronization and lmmunoblotting indicated that pRB was phosphorylated and dephosphorylated in a cell cycle-dependent manner. This suggests a functional role for the human pRB also in mouse cells. Moreover, cells expressing wild-type pRB were less susceptible to the transforming effects of SV40 large T antigen than cells expressing mutant pRB as shown by cell transfection studies. The results indicate that human pRB produced by mouse fibroblasts is functionally active and show that pRB can suppress the transforming activity of T antigen.