Purification and Immunological Characterization of the Major Internal Protein of the RD-114 Virus

115

Mammalian type-C viruses contain a major internal polypeptide of about 30,000 daltons that is characterized by both intraspecies and interspecies antigenic reactivities. Radioimmunoprecipitation assays were used for measurernent of this protein; the assay was based upon interspecies reactivities of the protein. As little as 5 ng of the group-specific antigen of murine leukemia virus can be measured by radioimmunoprecipitation assays, thus providing an approximate 10,000-fold increase in sensitivity over the standard immunodiffusion procedure. The type-C viruses that were recently isolated from a woolly monkey and gibbon ape each have an interspecies type-C antigenic reactivity. The primate viruses, however, could be distinguished from the type-C viruses of murine, rat, hamster, and feline origin that were more highly related to each other. The interspecies reactivity of the 30,000-dalton polypeptide is an immunological marker of the mammalian type-C viruses, since even with this sensitive assay other mammnalian viruses with RNAdependent DNA polymerase activity did not contain the type-C interspecies antigen.

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Radioimmunoassay of Mammalian Type-C Viral Proteins: Interspecies Antigenic Reactivities of the Major Internal Polypeptide

Parks

Scolnick

1972

115

“…When these tumor cells are inoculated into fetal cats in utero, tumors develop. From one of these tumors, a type-C virus (2) called RD-i 14 was isolated that was shown to have a group-specific (gs) antigen and an RNA-dependent DNA polymerase that differed immunologically from that of the type-C viruses previously isolated from cats (3)(4)(5). The suggestion was therefore made that this virus may have been derived from the human RD cells or from some recombinational event between genetic information in the RD cells and the cat host in which the cells grew.…”

mentioning

confidence: 99%

A Type-C Virus in Human Rhabdomyosarcoma Cells After Inoculation into NIH Swiss Mice Treated with Antithymocyte Serum

Todaro¹,

Arnstein²,

Parks³

et al. 1973

113

A type-C RNA virus has been isolated that replicates readily in human and other primate cells. It was obtained from a human rhabdomyosarcoma cell (RD) that had been serially transplanted in immunosuppressed NIH Swiss mice, a strain of mouse from which infectious type-C virus has not been isolated. Various other human tumor cells, similarly transplanted, remained free of overt type-C virus expression. The virus growing in the RD cells, AT-124, has a group-specific antigen and an RNA-dependent DNA polymerase immunologically related to murine type-C viruses, but a host range similar to that of the RD-114 virus. The new isolate is either a previously undescribed, endogenous type-C virus from NIH Swiss mice or a recombinant with both mouse and human type-C genetic information.

“…One comparison of critical importance was between FeLV and RD-114, both of which now appear to be of cat origin (10,(29)(30)(31)(32). Previous studies (23,(33)(34)(35) with the immunological results in showing a lack of crosshybridization at equivalent RNA concentrations. Further, no evident relationship between RD-114 and the primate viruses is seen.…”

mentioning

confidence: 99%

Specificity of the DNA Product of RNA-Dependent DNA Polymerase in Type C Viruses: II. Quantitative Analysis

Okabe

Gilden

Hatanaka

1973

A number of mammalian Type C viruses were analyzed for relatedness by the technique of DNA. RNA hybridization. Viral DNAs were prepared in singlestranded form from complexes with 70S viral RNA formed during endogenous polymerase reactions. Extent of hybridization was assayed with the single-strand nuclease (S-i) from Aspergillus oryzae. Results obtained indicated a high degree of viral specificity, with significant crossreactions being observed only with viruses obtained from within a species, as in the case of mouse and cat viruses, or in the special case of woolly monkey-gibbon comparisons.Comparisons of RD-114 virus, recently determined to be of feline origin, and conventional feline Type C viruses (FeLV), revealed minimal relatedness, especially when feline virus was grown on human cells, thus indicating the possibility of coexistence of greatly disparate Type C viruses within one species. A rat-specific virus, recovered from tumors induced by murine sarcoma virus, was found to contain genetic material common to both the original mouse virus and viruses indigenous to the rat, even though only rat-specific proteins have been detected during infection by this virus.The discovery of the RNA-dependent DNA polymerase (reverse transcriptase) in RNA tumor viruses (1, 2), and subsequent findings of base sequence complementarity between template viral RNA and product DNA (3-5), have made possible studies of viral interrelationships (6) and have shown presence of viral-specific RNA in infected (7,8) and apparently normal cells of several species (9-i 1). In previous studies utilizing cesium sulfate gradient centrifugation to detect RNA -DNA hybrids, mouse, hamster, cat, and viper Type C viruses were found to be completely distinguishable, with no evidence of interviral hybridization (6). Several mouse viruses did, however, show clear evidence of crosshybridization (6), as did chicken viruses (12). Because of recent findings of crosshybridization between mouse Type C virus DNA products and RNA from human malignant cells (13,14) and availability of more viruses for comparison, the question of viral interrelationship was re-examined by the use of sensitive and quantitative techniques employing the singlestrand specific nuclease (S-i) from Aspergillus oryzae. The viral 70S RNA and the [3H ]DNA products of the RNA-dependent DNA polymerase reaction used for hybridization experiments were purified by sucrose gradient centrifugation (10). To insure specificity, the DNA used was obtained from hybrid complexes with 70S RNA. In kinetic analyses such products gave results similar to those obtained using single-stranded probes prepared in the presence of actinomycin D (H. Okabe, submitted for publication). The latter preparations represented (in our experiments) at least 60% of the viral genome, based on protection of homologous 70S RNA from RNase digestion after annealing. Specific activity of the DNA products averaged 85 X 108 dpm/ng. RNA and DNA (after digestion of RNA) were dissolved in one tenth standard saline-citra...