THE G(1) PERIOD AND TRANSLATABLE MESSENGER-RNA CHANGES IN CULTURED CHICK EMBRYO FIBROBLASTS (DNA, CELL CYCLE, GROWTH FACTORS)

摘要

The temporal growth response of primary cultures of confluent chick embryo fibroblasts stimulated by serum and other putative growth factors, especially insulin, was determined. The G(,1) period was determined to be approximately 4.5 hours. A small percentage of the cells entered S in response to insulin, however, the majority of cells required additional factors. An insulin deficient serum preparation was made by treatment of serum with 50mM dithiothreitol (DTT-serum). This serum is unable to induce DNA formation by the chick cells. The growth promoting ability could be largely restored by addition of insulin indicating a complementing activity in the serum. The activity in the DTT-serum could be partially replaced by fibroblast growth factor (FGF) or a crude platelet-derived growth factor (PDGF) preparation but epidermal growth factor (EGF) was ineffective.;Serum stimulation of resting chick embryo fibroblasts results in increased RNA synthesis. Cell-free translation of mRNA populations indicates a selective accumulation of several mRNA species following serum stimulation. Three of these messages coded for polypeptides of approximately 34kd and, a fourth, for a polypeptide of about 30kd. These mRNAs appeared between 45 to 120 minutes after resting cells were treated with serum and their appearance was blocked by actinomycin D. Inhibitors of RNA synthesis, camptothecin and 5,6-dichloro-1-(beta)-D-ribofuranosylbenzimidazole (DRB), however, did not block their appearance. In fact, DRB enhanced the accumulation of the 30kd coding mRNA. FGF and PDGF also enhanced the appearance of the 30kd coding mRNA whereas insulin, EFG, and DTT-serum only induced the appearance of this mRNA weakly, it at all. The 34kd coding mRNAs were not detectably affected by the growth factors. In addition, an mRNA coding for a second more acidic 30 kd polypeptide was detected after FGF and PDGF treatment and to a lesser extent after EGF treatment. A possible role for these mRNAs in the growth response at the competency step is discussed.