EVIDENCE FOR CYTOPLASMIC DNA IN ROOT CELLS OF NICOTIANA

Abstract

Sterile root cultures from Nicotiana tabacum were grown with H3-thymidine added to the medium for various intervals. Incorporation of the labeled nucleoside into nuclear DNA occurred in a fraction of the nuclei which increased with time. In addition, the cytoplasm of all cclls incorporated cnough tritium to be rcadily dctected by autoradiography. The tritium was not rcmovcd by hydrolysis in 1 N HC1 at 60°C for l0 minutes, but was removed by digestion in a DNase solution which also removed nuclear DNA. The amount of tritium in the cytoplasm increased during the first 2 hours, but did not appear to increase significantl Y during the following 5 hours. If thc roots were transfcrrcd to unlabclcd medium aftcr 2 hours, the label was diluted faster than expected by growth without turnover of the labeled componcnt. If FUdR was added to the unlabeled medium, the depletion occurred faster during the first 6 hours, but later appeared to level off so that at l0 hours thesc cultures did not differ from those incubated without FUdR. However, the addition of an excess of unlabelcd carrier had no effect on the rate of depiction of the cytoplasmic label. Actinomycin D, which inhibited the incorporation of H3-cytidine into RNA in the root tips, had no effect on thc incorporation of Ha-thymidine into the cytoplasmic component. Howcver, Mitomycin C or a high concentration of deoxyadenosine inhibited the incorporation of H 3-thymidine into the cytoplasmic component as well as into the nuclear DNA. It is concluded that H3-thymidine is incorporated into a cytoplasmic fraction which has thc characteristics of DNA, with a mcasurable rate of turnover. This fraction is synthesized regardless of whether or not the nucleus is synthesizing DNA. Although the function of cytoplasmic fraction is not yet known, it does not appear to be that of supplying precursors for thc synthesis of the nuclear DNA.