EXPRESSION CLONING OF LFC, A NOVEL ONCOGENE WITH STRUCTURAL SIMILARITIES TO GUANINE-NUCLEOTIDE EXCHANGE FACTORS AND TO THE REGULATORY REGION OF PROTEIN-KINASE-C

In order to identify cDNAs that can induce oncogenic transformation, a retroviral vector was used to transfer a library of cDNAs from the mu rine 32D hemopoietic cell line into NIH 3T3 fibroblasts. We have ident ified and recovered a provirus containing a 1.8-kilobase pair cDNA who se expression causes morphological transformation in NIH 3T3 cells. Th e transforming cDNA contains a complete open reading frame that encode s a protein (designated Lfc) with a region of sequence similarity to t he product of the lbc oncogene. This region includes a domain that is characteristic of the CDC24 family of guanine nucleotide exchange fact ors in tan dem with a pleckstrin homology (PH) domain. The Lfc protein is distinguished from Lbc by a 150-amino acid NH2-terminal extension that contains a cysteine- and histidine-rich domain similar to the dia cylglycerol-binding site (zinc butterfly) found in protein kinase C. N H2- and COOH-terminal deletion analysis revealed that both the PH and putative guanine nucleotide exchange factor domains are required, but the zinc butterfly is dispensable, for transformation. Although the re moval of the PH domain of the Lfc protein completely eliminated its ab ility to transform MH 3T3 cells, replacement of this domain with an is oprenylation site restored all of its transforming activity. This sugg ests that a PH domain-dependent recruitment of the Lfc protein to the cellular membrane is a necessary step for cellular transformation. The lfc gene is expressed in a broad range of tissues as well as in a var iety of hemopoietic and non-hemopoietic cell lines. Lfc appears to be a new member of a growing family of proteins that are likely to act as activators of Ras-like proteins in a developmental or cell-lineage sp ecific manner.