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Biomedical subjects

E F Baril

Publications and source records attributed to E F Baril.

31 records · Page 2Linked to original sources
PubMed1978

HeLa DNA polymerase alpha activity in vitro: specific stimulation by a non-enzymic protein factor.

A non-enzymic protein factor that increases the in vitro rate of synthesis by HeLa DNA polymerase alpha 15- to 30-fold with denatured DNA as template has been partially purified from the cytoplasmic fraction of HeLa cells. The stimulatory effect is highly specific for HeLa DNA polymerase alpha and for DNA templates that contain extensive regions of single-strandedness. Synthesis with denatured DNA as template presumably proceeds from 3'-hydroxyl termini formed at loop-back regions since the synthesized DNA product and template are covalently linked. The stimulatory protein factor chromatographs as a basic protein, has an approximate molecular weight of 30,000 daltons and binds with moderate affinity to denatured DNA cellulose, being eluted by o.4M NaCl. The purified factor lacks detectable DNA polymerase, exo- and endodeoxyribonuclease and RNA polymerase activities. It also does not promote helix-coil transitions with poly[d(A-T)] and Clostridium perfringens DNA.

DNA Polymerase II↗
PubMed1976

Induction of DNA polymerase alpha during liver regeneration in rats on controlled feeding schedules.

The activity of 2 nonmitochondrial forms of DNA polymerase, designated DNA polymerases alpha and beta, was investigated during liver regeneration in regimented rats. In accord with Barbiroli and Potter, we observed that regimentation of rats with respect to temperature, light and darkness, and availability of food resolves the DNA synthesis response to partial hepatectomy into 2 peaks, one occurring at a fixed time after operation and the other entrained by the environmental conditions. The peaks can be fused or separated depending on the timing of the operation. For this study, operation times were selected to give both patterns of DNA synthesis as measured by the uptake of radioactive thymidine into DNA. For both operation times, DNA polymerase activity in the nuclear extract correlated temporally and qualitatively with radioactive thymidine uptake into DNA. At the times of maximal DNA synthesis and polymerase activity, the DNA polymerase was purified from extracts of isolated nuclei. DNA polymerase alpha represented 70% and DNA polymerase beta represented 30% of the recovered activity from the nuclear extract. This is in agreement with the previous observation in nonregimented rats that DNA polymerase alpha is the major activity in nuclei during liver regeneration. For both operation times, DNA polymerase activity in the postmicrosomal fraction was sedimentable and increased 3 to 4 times above the level observed with this same fraction from normal rat liver. This activity was shown to be due to DNA polymerase alpha only with this subcellular fraction. DNA polymerase alpha activity with this fraction peaked 4 to 6 hr after the time of maximal radioactive thymidine incorporation into DNA. DNA polymerase activity in the microsome fraction did not change significantly after partial hepatectomy. This activity has been shown to represent DNA polymerase beta. Prior administration of cycloheximide and actinomycin abolished the rise in DNA polymerase alpha activity in the nucleus and postmicrosomal fraction. Hydroxyurea did not prevent the rise in DNA polymerase alpha activity with those subcellular fractions but did inhibit over 90% of the uptake of radioactive thymidine into DNA. These data suggest, but do not prove, that DNA polymerase alpha activity is induced in response to the stimulus(i) for liver regeneration.

Animals↗
PubMed1975

Nuclear DNA polymerases and the HeLa cell cycle.

Purified nuclei of HeLa S3 cells contain two DNA-dependent DNA polymerases that have distinct physical and enzymatic properties. We have investigated the variations in their activity during the cell cycle of a synchronized culture. Cells were synchronized by a double thymidine block, harvested at various phases of the cycle, and the two DNA polymerases were purified partially by DEAE-cellulose and phosphocellulose chromatography. The activity of DNA polymerase I (low molecular weight, N-ethylmaleimide-insensitive) remains essentially constant throughout the cycle. The activity of DNA polymerase II (high molecular weight, N-ethylmaleimide-sensitive), however, increases during G1 to mid-S and declines, 7- to 10-fold between late-S and G2. Addition of cycloheximide (60 mug/ml) to cultures 12 hours after the release from thymidine block abolishes the rise in the activity of DNA polymerase II. Cycloheximide also reduced the activity of DNA polymerase I by 60%. Addition of hydroxyurea (1mM) at 1 hour after release has no effect on the activity of either enzyme. We conclude that in HeLa cells, DNA polymerase I and II are distinct enzymes, that DNA polymerase II probably functions in DNA replication and is probably induced in response to stimuli for DNA biosynthesis.

Cell Division↗
PubMed1975

Inhibition of rat liver DNA polymerase by nitrosoureas and isocyanates.

The effects of 1,3-bis(2-chloroethyl)-1-nitrosourea, 1-(2-chloroethyl)-3-cyclohexl-1-nitrosourea, and 1-(2-chloroethyl)-3-(trans-4-methylcyclohexyl)-1-nitrosourea on two nonmitochondrial DNA polymerases (I and II) purified from rat liver and hepatoma were examined. The activity of DNA polymerase I was not altered by treatment with any of the nitrosoureas or the corresponding isocyanates, 2-chloroethyl isocyanate and cyclohexyl isocyanate. Incubation of DNA polymerase II with the nitrosoureas (1 mM) inhibited its enzymatic activity 30 to 45%. DNA polymerase II was inhibited 75 and 90% by 1.mM 2-chloroethyl isocyanate and cyclohexyl isocyanate, respectively. The nitrosoureas appear to exert their inhabitory action on the enzyme (DNA polymerase II) rather than on the DNA template. Pretreatment of the enzyme increased the degree of inhibition by 1 mM nitrosourea (50 to 60% inhibition) or 2-chloroethul isocyanate (greater than 90% inhibition), whereas pretreatment of the DNA template did not enhance the inhibitory effect. The three nitrosoureas are equally effective as inhibitors of DNA polymerase II. 2-Chloroethyl isocyanate and cyclohexyl isocyanate are better inhibitors than are the nitrosoureas. Since further decomposition products of the isocyanates, 2-chloroethylamine and cyclohexylamine, do not inhibit DNA polymerase II, we conclude that the isocyanates, which are decomposition products of the nitrosoureas, are the active inhibitors of the enzyme.

Animals↗
PubMed1970

Amino acid transport in hepatoma cell cultures during tyrosine aminotransferase induction.

Alpha-aminoisobutyric acid transport in hepatoma cells in culture was increased by insulin but not by hydrocortisone. Both of these agents induce tyrosine aminotransferase activity in this system. The apparent increase in alpha-aminoisobutyric acid transport and tyrosine aminotransferase activity produced by glucagon is probably caused by insulin contamination. Insulin did not increase transport in this system until after tyrosine aminotransferase activity had reached maximum levels. The mechanisms underlying increased alpha-aminoisobutyric acid transport appear to differ from those for tyrosine aminotransferase induction with hydrocortisone despite their close association in previous whole animal experiments.

Aminoisobutyric Acids↗
PubMed1970

DNA polymerase activities associated with smooth membranes and ribosomes from rat liver and hepatoma cytoplasm.

Deoxyribonucleic acid polymerase activity that prefers denatured DNA primer concentrates with the smooth membranes during sucrose gradient centrifugation of rat liver and hepatoma cytoplasmic extracts. The activity in this fraction is eightfold higher in hepatoma tissue. Deoxyribonucleic acid polymerase activity that prefers native DNA primer concentrates with the free ribosome fraction from both tissues.

Ammonium Chloride↗