DNA synthesis in neuronal, glial and liver nuclei isolated from the adult guinea pig.

Abstract

Abstract— [3H]Deoxythymidine‐5′‐triphosphate incorporation into P51 (51% neuronal nuclei: 49% glial nuclei), P3 (3% neuronal nuclei: 97% glial nuclei) and liver nuclear preparations, isolated from the adult guinea pig, was determined in the presence of the other three complementary deoxyribonucleo‐tides. The enzymic characteristics of the DNA synthesis reaction were studied and DNA polymerase contents were estimated in neuronal, glial and liver nuclei. (1) Cerebral and liver nuclei exhibited similar enzymic properties for DNA synthesis activities with a few discrepancies. (2) P51 nuclei synthesized DNA 2.4‐fold more actively than P3 nuclei. Liver nuclei carried out the most active DNA synthesis. The proportion of chromatin DNA available as template and primer was estimated by comparison with native calf thymus DNA. The available proportions found, in terms of the total chromatin DNA. were 2.39% for P51 nuclei, 1.38% for P3 nuclei and 37.6% for liver nuclei. (3) Exogenous native and heat‐denatured calf thymus DNA were utilized as template and primer by DNA polymerase in nuclei in different ways depending on the nuclear species. The enzyme was saturated with native DNA by elevating the concentration and the activity reached a plateau. Denatured DNA inhibited the activity at the higher concentrations. (4) From the enzyme activities at a saturation concentration of exogenous DNA, DNA polymerase contents were estimated: P51 nuclei, 39.2 ± 2.6 (s.e.m.) units (fmol of TMP incorporated/30 min at 31°C)/μg of nuclear DNA; P3 nuclei. 24.5 ± 1.6; and liver nuclei, 72.5 ± 8.1; the specific activity obtained on a protein basis was 1.55 times higher with P3 nuclei than with P51 nuclei. (5) Denatured DNA inhibited the nuclear DNA polymerase activity dependent on native DNA. The efficiency of inhibition was in the order: P3 > P51 > liver nuclei.