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Polyadenylate sequences have been found covalently linked in heterogeneous DNA-like nuclear RNA of HeLa cells. This poly(A) material seems homogeneous in size and accounts for about 0.5% of such RNA. Similar poly(A) sequences were found in rapidlylabeled polyribosomal RNA, thought to be messenger RNA. A possible model for mRNA synthesis from large heterogeneous nuclear RNA precursor molecules is discussed.The synthesis of a relatively homogeneous population of polyriboadenylic acid [poly (A) I sequences of 150-200 nucleotides in the nucleus of Ehrlich ascites cells was recently described (1). The suppression of this synthesis with actinomycin D suggested that AMP-rich sequences were transcribed on DNA. Knowledge of the precise location of these sequences with respect to those RNA species which characterize eukaryotic cells was required to interpret these observations. Suspension cultures of HeLa cells were selected for such studies, not only for the techniques available for the isolation of wellcharacterized subcellular components (2) and RNA species (3, 4), but particularly for the detailed knowledge, available from kinetic analyses, of the flow of radioactivity through and into various RNA species of these cells (5).The poly(A) content of RNA fractions from the nucleus and cytoplasm of HeLa cells has been measured by a technique dependent on the hydridization of poly(A) sequences to polydeoxythymidylate oligomers covalently bound to cellulose (6). Highly purified poly(A) can be quantitatively recovered from suitably prepared RNA samples on temperaturecontrolled filters of poly(dT)-cellulose (7).We show here a broad distribution of poly(A) sequences within two classes of RNA that are themselves characteristically heterogeneous, i.e. the rapidly labeled polydisperse RNA (HnRNA) of the nucleus, and the polyribosome-associated RNA with the sedimentation characteristics of messenger RNA (mRNA). The poly(A) sequences recovered from all size classes of both types of RNA are rather homogeneous and show similar electrophoretic mobilities in acrylamide gel that correspond to an approximate length of 150-200 nucleotides.Evidence is presented that the poly(A) sequences detected after ribonuclease digestion of the HnRNA are in fact covalently bonded within that RNA. Equally compelling evidence for the covalent linkage of poly(A) detected in mRNA to that RNA has not yet been obtained, although such a linkage is considered likely.The implications of the detection of poly(A) sequences of similar length in both cytoplasmic mRNA and nuclear HnRNA will be considered with respect to a precursorproduct relationship proposed (8) to link these RNA species. MATERIALS AND METHODS Cell cultureSuspension cultures of HeLa cells were maintained as previously described (9), at cell concentrations of 2-5 X 105 cells/ml. Cell labeling and fractionationCells were concentrated to 3 X 106 cells/ml for labeling with 32PO4 (0.2 mCi/ml) in phosphate-free Eagle's minimal essential medium, supplemented with 5% normal calf serum. After the la...

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