Gene for M1 subunit of ribonucleotide reductase is amplified in hydroxyurea-resistant hamster cells.

The hydroxyurea-resistant Chinese hamster cell line 600H has been shown to have greatly elevated quantities of ribonucleotide reductase. This increase in enzyme activity is due to an increased level of both the M1 and M2 subunit activities. The M1 subunit has been purified from the 600H cell line and shown to consist of a series of six protein spots with apparent molecular weights of 88,000 daltons, but with varying isoelectric points in the range of pH 6.5-7.0. Western blot analyses with antisera against the M1 and M2 proteins indicated that both subunit proteins are present in elevated quantities in the 600H cell line when compared to the wild-type V79 cell line. Southern blot analyses with genomic DNA from the series of stepwise-selected hydroxyurea-resistant cell lines leading to 600H showed that, in latter steps of selection, genomic sequences homologous to a mouse M1 cDNA have undergone a fivefold amplification. This was accompanied by a four- to eightfold increase in the single M1 homologous mRNA.

Temperature-sensitive herpes simplex virus type 1 mutants defective in the shutoff of cellular DNA synthesis and host polypeptide synthesis.

Two temperature-sensitive herpes simplex virus type 1 mutants, ts 1-8 and ts 199, belonging to different complementation groups, were isolated. Both mutants were defective in the shutoff of host DNA synthesis at 39.5 degrees C (nonpermissive temperature). ts 1-8 exhibited intermediate levels of viral DNA synthesis at 39.5 degrees C, while ts 199 was completely deficient in viral DNA synthesis at 39.5 degrees C. Comparative polyacrylamide gel electrophoresis of the ts 1-8, ts 199 and wild-type viral-coded polypeptides and cellular proteins produced in vivo at 34 degrees C and 39.5 degrees C during various periods post infection was performed. The results indicated that ts 1-8 and ts 199 were temperature-sensitive for the secondary suppression of host polypeptide synthesis. Production of the beta (early) and gamma (late) viral polypeptides was slightly delayed in the mutant-infected cells at early times post infection at both 34 degrees C and 39.5 degrees C. This delayed protein production was not evident at later times post-infection. The ts 1-8 and ts 199 mutants were distinct from the HSV-1 viron-associated host shutoff (vhs) mutants of Read and Frenkel (J. Virol. 46 (1983) 498).