Transcriptional control of the fibronectin gene in chick embryo fibroblasts transformed by Rous sarcoma virus

Abstract

Rous sarcoma virus transformed-chick embryo fibroblasts contain decreased amounts of the adhesive protein fibronectin that results from a lowering of fibronectin mRNA levels (Fagan, J.B., Sobel, M.E., Yamada, K.M., de Crombrugghe, B., and Pastan, I. (1981) J. Biol. Chem. 256, 520-525). In the present study, nuclei isolated from chick embryo fibroblasts (CEF) and Rous sarcoma virus transformed-chick embryo fibroblasts (RSV-CEF) were used to measure relative rates of fibronectin RNA synthesis. [³²P]RNA was synthesized by preinitiated endogenous RNA polymerases in nuclei, and the radioactive RNA was subsequently hybridized to DNA fragments spanning approximately 40 kilobase pairs (approximately 85%) of the fibronectin gene. Transformation of CEF by RSV resulted in a 7- to 8-fold reduction in the amount of ³²P-labeled nuclear RNA hybridizing with fibronectin DNA sequences when the nuclei were incubated for either 3.5 or 45 min. These results indicate that decreased transcription and not reduced RNA stability plays a major role in lowering fibronectin mRNA levels in RSV-CEF. When nuclear RNA was transcribed in the presence of Sarkosyl, diminished RNA synthesis was still observed suggesting that reduced fibronectin transcription in RSV-CEF is not due to a block in RNA chain elongation. Our data indicate that transcriptional regulation of the fibronectin gene is an important mechanism of lowering steady state levels of fibronectin mRNA in RSV-CEF.