The properties of virus-like cytoplasmic particles in thymic tissue from normal and lymphomatous AKR mice and the expression of leukemia-related sequences in these tissues have been investigated. The retrovirus nature of the particles was indicated by the presence of RNA-dependent DNA polymerase as judged by the ability of the associated enzymatic activity to carry out endogenous RNase-sensitive DNA synthesis or utilize polycytidylate and poly(2′-O-methylcytidylate) and by the chromatographic properties of the purified enzyme. Equilibrium centrifugation studies of the particles extracted from the two types of tissue revealed a difference in the respective buoyant densities. A density of 1.157 g/ml was estimated for particles isolated from lymphomatous thymic tissue and for the Gross passage A virus, a leukemogenic strain of AKR murine leukemia virus. In contrast, a peak value of reverse transcriptase activity was observed at 1.145 g/ml in the gradient analysis of cytoplasmic particles prepared from the thymus of 1- and 2-month-old mice. Molecular hybridization studies aimed at the detection of differences in the expression of viral-specific information in RNA extracted from the normal and lymphomatous thymic tissues were carried out with a Moloney MuLV complementary DNA probe. Forty to 50% of the probe hybridized with the RNA from normal thymic tissue with an equivalent value of the product of the time (sec) of hybridization and the RNA concentration (moles of nucleotides per liter) at which 50% hybridization is reached indicative of approximately 100 genome equivalents of endogenous viral RNA per cell. About 80% of the probe hybridized with the RNA from lymphomatous tissue. Viral-related RNA in the lymphomatous tissue seemingly consisted of two abundance classes; one class (homologous with 50% of the probe) is hybridized at a rate suggestive of about 2400 viral genome equivalents per cell, while the second class is present at about 10 to 20 genome equivalents per cell. Thus in the lymphomatous thymus, virus-specific RNA expression was not only increased quantitatively (relative to normal thymus) but was characterized by the expression of qualitatively “new” sequences as well. The melting temperature profiles of hybrids of MuLV cDNA with RNA from normal and lymphomatous tissue were identical; the melting temperature was 83°. A melting temperature of 91° was observed for the hybrid composed of the probe and homologous Moloney MuLV 70S RNA.
This investigation was supported by Grants R01-CA16266 and CA18592 and Contracts P01 CA13053-05 and N01 CP1062 awarded by the National Cancer Institute, Department of Health, Education, and Welfare and by the Consiglio Nazionale delle Ricerche of Italy.