We have demonstrated previously that SV40 T antigen and serum regulate the length of G₁ in exponentially growing NIH‐3T3 cells in part by inhibiting density dependent negative cell cycle regulation. In these studies it was suggested that T antigen positively regulated G₁ in a density independent manner as well. In this report we show that, 24 h after treatment, TGF‐β1 perturbs the cell cycle of exponentially growing fibroblasts in a manner similar to T antigen. However, prior to 24 h, TGF‐β1 produced a negative response, elongating the Gi phase of the cell cycle that was followed by a positive response, both of which were density independent. This biphasic response was measured between 0 and 12 h post‐treatment and was relative to responses from serum. This switch from an early inhibitory effect to a late stimulatory effect was associated with changes in Rb phosphorylation, the timing and magnitude of which indicated that Rb may be directly regulating T_G1 rather than reporting changes in the population. This is further substantiated by abrogation of the inhibitory effect by expression of wild‐type SV40 T antigen and retention of the effect in cells that express an Rb‐binding mutant of T antigen (K1). The biphasic regulatory effects of TGF‐β1 were also displayed in WI‐38 and IMR‐90 human fibroblasts. This suggests that this biphasic effect is a property of fibroblasts.