Abstract
A malignant cell line (clone S1) isolated after co-transfection of normal NIH3T3 DNA and Moloney leukemia virus long terminal repeat (Mo-LTR) sequences has previously been described to contain an activated c-raf oncogene. Here, we report the isolation by molecular cloning and the structural analysis of the LTR-activated c-raf gene. As shown by Southern blot and nucleotide sequence analyses, the transfected Mo-LTR sequences integrated into the 5th intron of the endogenous c-raf proto-oncogene. This intragenic LTR insertion led to the expression of high levels of LTR-U5-c-raf hybrid transcripts indicating an initiation of transcription from the Mo-LTR promoter. Transcriptional activation of c-raf is accompanied by the synthesis of large amounts of cytoplasmic c-raf protein. Immunoblot analysis suggests that the proteins encoded by the LTR-activated c-raf gene are truncated compared with the normal c-raf gene product(s). Our results indicate a promoter insertion mechanism of c-raf activation.