Deficiency of joining of Okazaki-type fragments in absence of cellular protein synthesis.

Abstract

The dependence of integration of newly formed DNA chain ( less than 10 S) into larger DNA on concomitant protein synthesis was studied in a special cellular system. Exponentially growing Ehrlich ascites tumor cells in vivo show decreasing rated and finally complete cessation of protein and DNA synthesis upon transfer into an isotonic but non-nutritive environment (Hanks' balanced salt solution). Both protein and DNA synthesis is stimulated in these cells for a period of 30 min when they are placed into fresh Hanks' balanced salt solution; however, stimulation of protein synthesis is completely prevented in Hanks' balanced salt solution containing cycloheximide. This system allowed us to investigate the formation and fate of newly formed DNA chains ( less than 10 S) in dependence of protein synthesis. Analysis of DNA produced in [3H]thymidine pulses showed that DNA chains smaller than 18 S were still formed during the phase of totally delayed protein synthesis and in the presence of cycloheximide, but they were not converted into DNA molecules sedimenting faster than 18 S under these conditions. Stimulation of protein synthesis for a period of 30 min allowed the short DNA pieces to be chased into larger DNA 30 min post stimulation of protein synthesis. The results clearly indicate that DNA chain growth, by sealing of DNA chains smaller than 18 S, is strongly dependent of concomitant protein synthesis. Direct chain elongation by addition of new deoxyribonucleotides is less dependent on concomitant cellular protein synthesis.