Pulmonary foam cells in rats of different age.
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Biomedical subjects
Publications and source records attributed to G Magnusson.
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During in vitro incubation, nuclei from polyoma-infected cells elongate the daughter strands of the replicative intermediate of polyoma DNA. This process is now shown to involve the transient formation of short fragments (4-5 S), a process that is stimulated by the addition of ribonucleoside triphosphates. The presence of stretches of RNA at the 5'-end of short DNA chains was determined from Cs(2)SO(4) equilibrium centrifugation and from the finding that isotope from alpha-(32)P-labeled deoxynucleoside triphosphates was recovered in 2'(3')-ribonucleotides after alkaline hydrolysis. Transfer occurred preferentially with [alpha-(32)P]dCTP as substrate. Starvation for deoxynucleotides by in vivo treatment with hydroxyurea resulted in the accumulation of short fragments that are deficient in RNA. Our results suggest that a late step during the discontinuous synthesis of polyoma DNA is selectively inhibited when deoxynucleotides are in short supply.
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Hydroxyurea treatment of 3T6 mouse fibroblast cells infected with polyoma virus resulted within 15 min in more than a 20-fold reduction of the rate of both viral and cellular DNA synthesis. After the initial rapid inhibition, the rate of DNA synthesis remained essentially constant for at least 2 h. In the inhibited cells viral DNA accumulated as short chains with a sedimentation coefficient of about 4S (hydroxyurea fragments). A variable proportion of these fragments was released from the template strands when the viral DNA was extracted by the Hirt procedure. Reannealing experiments demonstrated that hydroxyurea fragments were polyoma-specific and probably synthesized on both parental strands at the replication forks.
The synthesis of polyoma DNA was studied in isolated nuclei from hydroxyurea-inhibited 3T6 cells infected with polyoma virus. During incubation of nuclei under conditions suitable for polyoma DNA synthesis in vitro, the short DNA fragments with a sedimentation coefficient of 4S formed in vivo (hydroxyurea fragments) became associated with preformed, replicating DNA strands. Centrifugation in dye-buoyant density gradients showed that the fragments formed part of the structure of the replicative intermediate of polyoma DNA. The proportion of "young" replicative intermediates was larger after hydroxyurea inhibition than in uninhibited controls. Hydroxyurea fragments appear to be closely related to the 4S fragments formed as normal intermediates during discontinuous synthesis of polyoma DNA.
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