Giant-size rapidly labeled nuclear ribonucleic acid and cytoplasmic messenger ribonucleic acid in immature duck erythrocytes

Abstract

The pattern of RNA synthesis has been investigated in immature erythrocytes present in the blood from normal and anemic ducks and maintained in vitro in a suitable medium. The majority of the RNA synthesized in vitro by these cells remains associated with the nuclear fraction and consists of RNA species of high sedimentation coefficient. This heavy RNA appears to include two distinct types: (a) ribosomal-type RNA, representing presumably precursors of ribosomal RNA, sedimenting between 30 and 50 s; (b) RNA different from ribosomal RNA in base ratios and with sedimentation coefficients ranging from 30 to 80 s or more. Under the conditions of cell maintenance in vitro used in the present work, the synthesis of presumptive ribosomal RNA precursors appears to be limited in time, ceasing after a few hours: in addition, only a relatively small conversion of such precursors to mature ribosomal RNA takes place in immature erythrocytes. The heavy non-ribosomal type nuclear RNA represents a heterogeneous mixture of molecules characterized by high U and relatively low GC content. All the evidence accumulated in the present work using enzymic tests, changes in the extraction procedure and in the conditions of sedimentation analysis, thermal denaturation tests, reconstruction experiments and base composition studies, supports the conclusion that the high sedimentation coefficients of these molecules are a reflection of their large size, corresponding to molecular weights from 2 × 10 6 to 10 7 or possibly more. These giant-size molecules are metabolically unstable, with a halflife of about 30 min; under the experimental conditions used in the present work, they continue to be synthesized and destroyed for as long as two days. A minor portion of the RNA synthesized in vitro by immature duck erythrocytes during a two-to three-hours incubation is found to be associated with the polysome fraction, and contains, in addition to a variable but in general relatively small amount of ribosomal RNA, a heterogeneous RNA fraction with sedimentation coefficients between 6 and 35 s, and base ratios different from those of ribosomal RNA and reflecting rather the base composition of DNA: this fraction can be identified as messenger RNA. In blood cell populations rich in immature erythrocytes at early stages of development, the labeling of the polysomal messenger RNA increases dramatically, and a large fraction of it sediments as a relatively sharp peak with a sedimentation coefficient of about 9 s: this corresponds to a molecular weight of about 150,000, which would be expected for the messengers of hemoglobin chains. Both base composition and kinetics data strongly suggest that no relationship of precursor-to-product type or, at any rate, not a simple one, exists between the heavy non-ribosomal type nuclear RNA and the cytoplasmic messenger RNA.