A 1.6-Kb cDNA (A-raf) has been isolated from a murine spleen cDNA library, which encodes part of a protein related to the v-raf oncogene. Its amino acid sequence has 85% homology to raf in central protein of 100 amino acids. When incorporated into a retrovirus, the resulting gag-A-raf fusion gene causes transformation in vitro and induces tumors in newborn mice. Later on we isolated the complete 2453- nucleotide sequence of the human A-raf gene from a human T-cell cDNA library. When the 5' deleted fragment of the cDNA is incorporated into a murine retrovirus, the resulting gag-A-raf fusion gene causes transformation in vitro and in vivo. Whereas, the full-lengths of c-raf-1 and human A-raf were not transforming when they were constructed under the control of a murine leukemia virus promoter (long terminal repeat). Moreover, when we deleted 20 amino acids from the N-terminal of c-raf-1 and incorporated them into a murine retrovirus, the resulting gag-c-raf-1 fusion gene caused transformation of fibroblasts. In trying to define the minimal sequences of raf oncogenes required for their transforming ability, we found that deletions of 14 N-terminal and 13 C-terminal amino acids were dispensable, but deletion of 28 or more amino acids from v-raf at the carboxy terminal abolished all transforming activity. Furthermore, we made four different XHOI linker insertion mutants of c-raf-1. These mutants will be incorporated into retroviral vectors for expression.
