Turnover of nucleic acids in a multiplying animal cell. 2. Retention studies

Abstract

The retention of [8-C14]adenine, [2-C14]-uridylic acid and [p32]orthophosphate in the ribonucleic acid (RNA) of multiplying HeLa cells was studied. The results were in general agreement with previous observations that was made on non-multiplying cells. Turnover of RNA was demonstrated, which appeared to involve mainly nuclear RNA. The cells accumulated large and persistent pools of radioactive precursors of nucleic acids even after brief exposures to radioactive medium. When radioactive cells were transferred to non-radioactive medium, radioactivity continued for several hours to enter deoxyribo-nucleic acid (DNA) and both nuclear RNA and cytoplasmic RNA. These "pool effects" were greatest when [P32]orthophosphate was used to label the cells. The magnitude of the "pool effect" could be diminished by introducing unlabelled RNA precursors into the non-radioactive medium. When the cells were labelled with [8-C14]adenine, the precursors that most effectively diminished the continued rise in radioactivity in RNA were adenosine and the 2[image]-, 3[image]- and 5[image]-phosphates of adenosine. Adenine was less effective, and hypoxanthine, inosine and inosine 5[image]-phosphate had little effect. When retention of radioactive adenine was studied in medium containing adenosine, the specific radioactivity of adenine in the nuclear RNA fell rapidly, but it was not possible to prevent a rise in the specific radioactivity of adenine in the cytoplasmic RNA. The rate at which the specific radioactivity of adenine in nuclear RNA fell varied with the length of the initial period of growth in radioactive medium. The nuclear RNA contained at least 3 fractions that had half-lives of turnover between 45 minutes and 5 hours. A fraction of nuclear RNA with a fast rate of turnover was degraded to acid-soluble material during the phenol extraction of RNA. The results were not consistent with a simple precursor-product relationship between nuclear and cytoplasmic RNA.