The role of the retinoblastoma gene product (pRb) as a critical controlling element for inhibiting cell proliferation was first suggested by the demonstration that mutations in the retinoblastoma gene are frequently associated with several specific tumors, and that in gene transfer studies its overexpression in transformed cells suppresses cell growth. Additional evidence supporting such a role are the observations that its state of phosphorylation is altered with the cell cycle. Because of our interest in the cellular and molecular factors that control SMC proliferation, we are studying the influence of suppressor genes, like Rb, on this process. Recent studies on simian virus 40 T antigen (Tag) have shown that one domain of this multifunctional protein binds with pRb, and thereby inhibits its suppressor activity. To demonstrate the effect of pRb on the cell cycle of SMC we used, in collaboration with Dr. Bruce Howard, a novel system that allows sensitive assessment of the effects of transfected suppressor gene activity on cell proliferation. Rat SMCs were transiently transfected to express wild type Tag or a mutant. The mutated form of Tag is defective in binding pRb. Thus, if pRb does play a role in SMC proliferation, its binding by Tag should stimulate quiescent SMCs to proliferate. Our results suggest that the product of the retinoblastoma gene is indeed expressed in SMC in vitro, and that this product seems to exert a suppressor function in the maintenance of SMC quiescence.
