Transcription of the viral genome in papillomas and carcinomas induced by the Shope virus.
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Biomedical subjects
Publications and source records attributed to F O Wettstein.
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In an initial efforts to characterize the virological basis of neoplasia in the Shope papilloma-carcinoma system, the extent to which the viral genome is present in non-virus-producing benign and malignant tumors in domestic rabbits was established. Employing nick-translated radioactive viral DNA purified from productively infected papillomas on cotton tail rabbits as a probe, it was found that (i) papillomas, primary carcinomas, and metastatic carcinomas contain 10 to about 100 copies of the viral genome per diploid cell equivalent of DNA and (ii) viral DNA is present in detectable amounts in essentially all neoplastic cells. These results are consistent with the suggestion that continued presence of the viral genome is necessary for induction and maintenance of malignant as well as benign neoplasms.
Immunoglobulin M synthesized by a human lymphoblastoid cell line, LA173, was found to bind specifically to the protein A-bearing Cowan I strain of Staphylococcus aureus. The (3H)-leucine-labeled, secreted IgM from these LA173 cells also formed precipitin complexes with purified protein A. Soluble complexes formed at high protein A/IgM ratios retained the ability to bind to the bacterial surface. Precipitin complexes also were observed in double diffusion Ouchterlony gels with a line of identity formed between the IgM, protein A, and anti-IgM in adjacent wells. Intracellular IgM species from detergent-lysed LA173 cells were bound to S. aureus. Labeled 19S pentamers, 8S monomers, and HL subunits were eluted from the bacteria and identified by velocity sedimentation and SDS agarose-acrylamide gel electrophoresis. In addition, several intermediates migrating between 8S and 19S were detected and shown to contain authentic H and L chains. Binding of the labeled IgM 19S pentamers to staphylococci was not inhibited by prior treatment of the bacteria with an excess of unlabeled human IgG. However, S. aureus saturated with unlabeled IgG did not bind either labeled IgM monomers or labeled IgG. The interaction of this human IgM with S. aureus exhibited a high degree of specificity with quantitative recovery of secreted 19S IgM. Intracellular IgM species were bound selectively by the bacteria with little if any contamination by other cytoplasmic proteins.
Virus-specific polysome-associated RNA (psRNA) and RNA after dissociation of polysomes were analyzed by direct hybridization with unlabeled viral RNA (vRNA) and complementary RNA (cRNA). psRNA after a 30-min pulse with [3H]uridine contained 28% labeled cRNA, 70% host RNA, and no vRNA. After dissociation, psRNA sedimented heterogeneously. Heavy RNA (greater than 60S), ribosomal subunit RNA (rsuRNA, 30-60S), free mRNA (fmRNA, 10-30S), and light RNA (less than 10S) contained 16%, 54%, 70% and 28% cRNA, respectively, but no vRNA. When actinomycin D (AcD) was added at 2 h postinfection, the nature of the psRNA depended on the concentration of AcD and the condition of the labeling. At AcD concentrations of 1 mug or more per ml, no detectable vRNA or cRNA was associated with polysomes. At 0.2 mug of AcD per ml (a concentration that partially inhibited cRNA synthesis) and 2 h of labeling at 2.5 h postinfection, psRNA contained 40% viral-specific RNA, which included both vRNA and cRNA in almost equal amounts. When polysomes were dissociated, however, viral-specific fm RNA from AcD-treated cells contained exclusively cRNA and no detectable vRNA. Increasing amounts of labeled vRNA were present in the heavy region of the gradient (and in the pellet), which also contained varying amounts of cRNA. The labeled vRNA appears to be associated with polysomes in a cesium chloride density gradient (rho = 1.525 g/ml). Although we have ruled out the trivial explanation of viral ribonucleoprotein contamination,the nature of the complex containing both polysomes and vRNA is unknown.
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