C. Birchmeier

Isolation and characterization of a new cellular oncogene encoding a protein with multiple potential transmembrane domains.

We have cloned and sequenced a new human oncogene and have named it mas. This oncogene was detected by its tumorigenicity in nude mice using the cotransfection and tumorigenicity assay previously described. The mas oncogene has a weak focus-inducing activity in transfected NIH 3T3 cells. A DNA rearrangement in the 5 noncoding sequence, which occurred during transfection, is probably responsible for activation of the mas gene. The cDNA sequence of the mas oncogene reveals a long open reading frame that codes for a 325 amino acid protein. This protein is very hydrophobic and has seven potential transmembrane domains. In this respect, the structure of the mas protein is novel among cellular oncogene products and may reflect a new functional class of oncogenes.

The S. cerevisiae CDC25 gene product regulates the RAS/adenylate cyclase pathway

The gene corresponding to the S. cerevisiae cell division cycle mutant cdc25 has been cloned and sequenced, revealing an open reading frame encoding a protein of 1589 amino acids that contains no significant homologies with other known proteins. Cells lacking CDC25 have low levels of cyclic AMP and decreased levels of Mg2+-dependent adenylate cyclase activity. The lethality resulting from disruption of the CDC25 gene can be suppressed by the presence of the activated RAS2val19 gene, but not by high copy plasmids expressing a normal RAS2 or RAS1 gene. These results suggest that normal RAS is dependent on CDC25 function. Furthermore, mutationally activated alleles of CDC25 are capable of inducing a set of phenotypes similar to those observed in strains containing a genetically activated RAS/adenylate cyclase pathway, suggesting that CDC25 encodes a regulatory protein. We propose that CDC25 regulates adenylate cyclase by regulating the guanine nucleotide bound to RAS proteins.

RAS proteins can induce meiosis in xenopus oocytes

Injection of human H-ras protein induces maturation of Xenopus oocytes; that is, progression from prophase to metaphase of meiosis. The oncogenic protein encoded by H-rasval12 is nearly a 100-fold more potent than the protein encoded by the wild-type gene. We do not observe any measurable increase or decrease in cyclic AMP concentration in injected oocytes, and the effects of H-ras protein are only partially blocked by cholera toxin. Our results suggest that not all, if any, of the effects of H-rasval12 protein in this system are mediated by adenylate cyclase.

Expression of three mammalian cDNAs that interfere with RAS function in Saccharomyces cerevisiae.

Saccharomyces cerevisiae strains expressing the activated RAS2Val19 gene or lacking both cAMP phosphodiesterase genes, PDE1 and PDE2, have impaired growth control and display an acute sensitivity to heat shock. We have isolated two classes of mammalian cDNAs from yeast expression libraries that suppress the heat shock-sensitive phenotype of RAS2Val19 strain. Members of the first class of cDNAs also suppress the heat shock-sensitive phenotype of pde1- pde2- strains and encode cAMP phosphodiesterases. Members of the second class fail to suppress the phenotype of pde1- pde2- strains and therefore are candidate cDNAs encoding proteins that interact with RAS proteins. We report the nucleotide sequence of three members of this class. Two of these cDNAs share considerable sequence similarity, but none are clearly similar to previously isolated genes.

Characterization of an activated human ros gene.

A human oncogene, mcf3, previously detected by a combination of DNA-mediated gene transfer and a tumorigenicity assay, derives from a human homology of the avian v-ros oncogene. Both v-ros and mcf3 can encode a protein with homology to tyrosine-specific protein kinases, and both mcf3 and v-ros encode a potential transmembrane domain N terminal to the kinase domain. mcf3 probably arose during gene transfer from a normal human ros gene by the loss of a putative extracellular domain. There do not appear to be any other gross rearrangements in the structure of mcf3.

Characterization of Two New Human Oncogenes

The first oncogenes discovered were the transforming genes of the oncogenic viruses (reviewed by Bishop 1985). The subsequent discovery that the oncogenes of retroviruses were derived from normal host cellular genes provided the first direct evidence that cellular genomes contain genes with transforming potential. More recently, the development of techniques for DNA transfer in eukaryotic cells led to the discovery of cellular transforming genes in tumor cells by their ability to induce foci of transformed NIH-3T3 cells (reviewed by Land et al. 1983). Several new oncogenes have been discovered this way, including N-ras (Shimizu et al. 1983), met (Cooper et al. 1984), neu (Bargmann et al. 1986), and possible others (Goubin et al. 1983; Lane et al. 1984; Takahashi et al. 1985).

Conservation and Divergence of RAS Protein Function during Evolution

The ras genes were first isolated as the transforming genes of Harvey and Kirsten sarcoma virus (Ellis et al. 1981). At least three different ras genes, Ha-ras, Ki-ras, and N-ras, exist in mammals and code for three very similar 21-kD proteins (Shimizu et al. 1983b). The ras proteins are localized in the plasma membrane (Willingham et al. 1980), bind guanine nucleotides (Shih et al. 1980, 1982), and have weak GTPase activity (Gibbs et al. 1984; McGrath et al. 1984; Sweet et al. 1984). A large number of tumor cells contain structurally mutated ras genes that are capable of tumorigenic transformation of NIH-3T3 cells upon DNA-mediated gene transfer (Reddy et al. 1982; Tabin et al. 1982; Taparowsky et al. 1982; Capon et al. 1983; Shimizu et al. 1983a; Yuasa et al. 1983)