Cytoplasmic RNA-dependent RNA-synthesis in maturing chicken erythrocytes.

Doctoral Thesis


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University of Cape Town

An RNA-dependent RNA polymerase from ribosomes of maturing chicken erythrocytes was investigated. Concurrent with an increase in globin and RNA synthesis during phenylhydrazine induced anaemia, the total and specific activities of ribosome bound RNA-dependent RNA polymerase increased 40 and 9.4 times respectively, indicating the possible involvement of this enzyme activity in the synthesis or control of synthesis of globin. A partially purified RNA-dependent RNA polymerase was prepared from ribosomes of immature chicken erythrocytes. This preparation was shown to be a predominantly primer dependent enzyme activity, incorporating UTP, CTP and to a lesser extent ATP and GTP into homopoly-ribonucleotide material. The ribosome-bound enzyme preparation demonstrated in addition, the synthesis of heteropolyribonucleotide material. A complementary DNA copy of chicken globin mRNA containing-9S RNP, was prepared. Unlabelled complementary DNA was used to demonstrate that no globin mRNA replicase activity was detectable in the ribosome fraction of immature chicken erythrocytes. The radioactively labelled product obtained after incubation of ribosomes with [³H]-UTP was shown by sucrose density gradient centrifugation to have a sedimentation coefficient of 4S; the ratio of non-terminal to terminal incorporation of UTP was 6.08. The product synthesized after incubation of intact cells in the presence of [³H]-uridine and actinomycin D was shown to be very similar with respect to S-value and the ratio of UMP/uridine incorporated. This similarity suggests the presence of a ribosome bound RNA-dependent RNA polymerase activity in intact cells.

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