Transforming growth factor‐β regulation of retinoblastoma gene product and E2F transcription factor during cell cycle progression in mouse fibroblasts

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Abstract
The mechanism by which transforming growth factor beta (TGFβ) exerts growth stimulatory effects was examined in C3H/10T1/2 mouse fibroblasts by study of cell cycle regulation of the retinoblastoma gene product (p110Rb) and transcriptional regulation of the p110Rb‐associated transcription factor, E2F. Northern blotting analysis shows that TGFβ and/or epidermal growth factor (EGF) stimulate by three to sixfold the level of Rb mRNA which is also reflected by the increased levels of p110Rb. p110Rb becomes phosphorylated in mid‐G1 and further phosphorylated at the G1/S transition. Hyperphosphorylation of p110Rb by TGFβ can be observed when cells are in S phase. TGFβ stimulates by three to fourfold the activity of cdk2 kinase consistent with the observed phosphorylation of p110Rb and also with the possibilit that the kinase is involved in phosphorylating p110Rb close to the G1/S transition. Thus, TGFβ as a growth stimulator induces, as does EGF, the phosphorylation of p110Rb during cell cycle progression. Transient transfection of E2F promoter constructs was used to analyze the effect of TGFβ on the modulation of E2F‐mediated transcription. The data revealed that TGFβ can stimulate wild‐type adenoviral E2 promoter activity by 12‐fold. Taken together, TGFβ‐induced phosphorylation of p110Rb in mouse fibroblasts appears to exert a positive regulatory function upon genes that have a pivotal role in the G1/S transition of the cell cycle.