Steven R. Tronick

Expression of the normal human sis/PDGF-2 coding sequence induces cellular transformation

The human sis proto-oncogene contains the coding sequence for one of two polypeptide chains present in preparations of biologically active human platelet-derived growth factor (PDGF). A human clone, c-sis clone 8, which contains all of the v-sis-related sequences present in human DNA, was transcriptionally inactive when transfected into NIH/3T3 cells. When placed under the control of a retrovirus LTR, the clone was transcribed at levels comparable to that observed in cells transformed by SSV DNA. However, c-sis clone 8 DNA did not express detectable sis/PDGF-2 proteins and lacked biologic activity. A putative upstream exon was identified by its ability to detect the 4.2 kb sis-related transcript in certain human cells. When this sequence was inserted in the proper orientation between the LTR and c-sis clone 8, the chimeric molecule acquired high titered transforming activity, comparable to that of SSV DNA. Transformants containing this construct expressed human sis/PDGF-2 translational products. Thus the normal coding sequence for a human growth factor has transforming activity when expressed in an appropriate assay cell.

Generation of BALB-MuSV and Ha-MuSV by type C virus transduction of homologous transforming genes from different species

The nature of the cell-derived (bas) sequences of BALB-MuSV, a spontaneous mouse sarcoma virus isolate, was determined. Molecularly cloned bas sequences demonstrated no detectable homology with the onc genes of other mouse transforming viruses, but exhibited a high degree of sequence homology with the ras gene of the rat-derived Harvey murine sarcoma virus (Ha-MuSV) genome. The Ha-MuSV cell-derived sequence (ras) shared a colinear 750 bp region of homology with bas. Moreover, BALB-MuSV transformation was associated with the expression of high levels of a 21,000 dalton protein, immunologically related to the ras gene products, p21. Thus bas and ras represent retroviral transforming gene homologs that were independently transduced by mouse type C viruses from the genomes of different species.

Distribution of three classes of endogenous type-C RNA viruses among inbred strains of mice.

Abstract Mouse cells contain genetic information for type-C RNA viruses. Three prototype endogenous viruses with biologically distinguishable properties have been isolated. The present studies show that viruses immunologically indistinguishable from these prototype viruses are either chemically inducible or partially expressed in cells of many inbred mouse strains of widely diverse geographic origin. The evidence further indicates that the expression of these three virus classes is differentially regulated within cells of the strains examined.

Chromosomal localization of three human ras genes by in situ molecular hybridization

Three human rasfamily protooncogenes, c-Ki-ras-1, and c-Ki-ras-2, and N-ras,have been mapped to chromosome bands 6p11–12, 12p11.1–12.1, and 1p11–13, respectively by in situ molecular hybridization. Certain human cancers display consistent and specific alterations involving chromosomes 1, 6, and 12. The precise chromosomal localization of ras genes will permit evaluation of thepossible effect of these chromosome changes on the structure and activities of ras protooncogenes in human neoplasia.

Murine leukemia virus mutants with temperature-sensitive defects in precursor polypeptide cleavage

Abstract Among temperature-sensitive conditional lethal mutants of the Rauscher strain of murine leukemia virus (R-MuLV), several were previously shown to express viral antigens in the absence of complete virus release at the nonpermissive temperature. In the present study, analysis of the size distribution of viral polypeptides of molecular weight 30,000 and 12,000 daltons in extracts of cells infected with two temperature-sensitive mutants at the nonpermissive temperature revealed a marked accumulation of both antigens in a size region corresponding to 60,000–70,000 daltons. The precursor was partially purified and shown to contain the polypeptides of molecular weight 15,000 daltons, as well as 30,000 and 12,000 daltons, in relative concentrations similar to those found in purified virions.

Analysis of type specific antigenic determinants of two structural polypeptides of mouse RNA C-type viruses

Abstract Radioimmunoassay techniques have previously been shown to be highly sensitive and specific for the detection of intra- and interspecies determinants of antigens of RNA C-type viruses. In the present report, two murine leukemia virus (MuLV) polypeptides, with molecular weights of 30,000 and 12,000, are shown to contain type-specific determinants by use of appropriate radioimmunologic procedures. These methods have made it possible to distinguish three groups of MuLV. The type-specific antigenic properties of the viral polypeptides examined are shown to be independent of previously described host range and neutralization properties.

Immunological characterization of a low molecular weight polypeptide of murine leukemia virus.

Abstract A low molecular weight polypeptide (MW 16,000) of the Rauscher strain of mouse leukemia virus (MuLV) has been isolated. A very sensitive and highly specific radioimmunoassay has been developed for its quantitation. The present studies indicate that this polypeptide is virus-coded and antigenically distinct from another virion protein, the group-specific (gs) antigen Different strains of MuLV contain antigens immunologically cross-reactive with the low molecular weight polypeptide. Studies are presented comparing the level of expression of this virion antigen in normal and transformed cells.

Demonstration of two immunologically distinct xenotropic type C RNA viruses of mouse cells

Abstract Several independent type C virus isolates were obtained by transplantation of human tumor cell lines into immunosuppressed NIH Swiss mice. Each isolate appeared to be of mouse origin as determined by the antigenicity of its major virion polypeptide, p30. The virus isolates were indistinguishable from each other in serologic and host range properties and also closely resembled BALB:virus-2, one endogenous virus of BALB c mouse cells. However, in immunoassays for a highly type-specific virion polypeptide, designated p12, every isolate was found to differ from BALB: virus-2. These results provide evidence for the existence of immunologically distinct xenotropic viruses of mouse cells.

Evidence for genetic recombination between endogenous and exogenous mouse RNA type C viruses.

Abstract Two viruses isolated following prolonged growth of serologically distinct mouse type C RNA viruses in human cells have previously been shown to have acquired common envelope properties distinct from those of either parental virus. Virus neutralization tests show that the viruses selected in human cells possess envelope antigens identical to those of endogenous mouse type C viruses of cells in which the parental viruses had been propagated. In contrast, the p12 polypeptide of each virus selected in human cells is antigenically indistinguishable from that of its respective parental virus and different from those of known endogenous mouse type C viruses. Molecular hybridization indicates significant differences in the genetic sequences of one virus and its parent, excluding the possibility that it arose from a point mutation. These findings indicate that the viruses selected in human cells represent genetic recombinants between exogenous and endogenous mouse type C viruses.

Comparative immunological studies of RNA C-type viruses: radioimmunoassay for a low molecular weight polypeptide of woolly monkey leukemia virus.

Abstract A 12,000-dalton polypeptide was partially purified from the woolly monkey RNA C-type virus. A sensitive and specific radioimmunoassay for this polypeptide detected major differences in the reactivities of woolly monkey and gibbon ape viruses. In contrast, these viruses were indistinguishable in immunoassays for two other woolly monkey virion proteins, the major structural polypeptide and the reverse transcriptase. Each assay readily differentiated the woolly and gibbon from other mammalian RNA C-type viruses.