Ovalbumin Messenger RNA: Evidence That the Initial Product of Transcription Is the Same Size as Polysomal Ovalbumin Messenger

Abstract

The messenger RNA for ovalbumin, the major secretory protein of the chick oviduct, appears not to be made as a high-molecular-weight precursor when artifacts due to aggregation are eliminated. No ovalbumin messenger RNA sequences that will hybridize to complementary DNA made against ovalbumin mRNA are found in concentrated samples of hen oviduct RNA larger than 28 S. The sensitivity of the hybridization assay is sufficient to detect less than one molecule of ovalbumin mRNA precursor per tubular gland cell. Newly synthesized ovalbumin messenger RNA isolated from immature chicks stimulated briefly by estrogen is the same size as that found in hen polyribosomes. We conclude that ovalbumin messenger RNA does not undergo any significant change in molecular weight from its initial transcription to its incorporation into polyribosomes.The synthesis of messenger RNA in animal cells and its transport from the nucleus to the cytoplasm has received much attention in recent years, but our understanding of this process is incomplete and confused. A model for mRNA synthesis has been proposed by Darnell and coworkers (1) which involves the initial synthesis of a large precursor RNA with subsequent post-transcriptional addition of poly(A) sequences to the 3' terminus. This large precursor is degraded except for the 3' portion containing the mRNA, which is then transported to the cytoplasm and incorporated into polysomes.A large amount of evidence supporting this hypothesis has been reported. Both heterogeneous nuclear RNA (HnRNA) and mRNA contain a 3'-terminal poly(A) region of approximately the same length (2). Jelinek et al. (3) have reported that much of the poly(A) synthesized in the nucleus is conserved and transported to the cytoplasm. However, Perry and coworkers (4) have recently suggested that most of the nuclear poly(A) remains in the nucleus and is degraded there. Further evidence for the precursor-product relationship between HnRNA and mRNA has been obtained for specific mRNAs by either translation of the HnRNA in vitro (5,6) or by its hybridization to complementary DNA (7). Such results are not completely convincing because the possibility of aggregation artifacts in sucrose gradients has not been rigorously excluded. Proof that a large precursor exists for a specific mRNA should involve separation of the precursor from polysomal mRNA and a careful demonstration of its covalent integrity by a variety of denaturing techniques. This has not Abbreviations: OV, ovalbumin; HnRNA, heterogeneous nuclear RNA; NaDodSO4, sodium dodecyl sulfate; Me2SO, been reported for any proposed precursor, and in cases where dimethylsulfoxide (Me2SO) gradients have been used, most of the large mRNA sequences dissaggregate to polysomal mRNA sizes (7). In this study we report evidence suggesting that the mRNA coding for chick oviduct ovalbumin (OV) is synthesized, transported to the cytoplasm, and incorporated into polysomes without a measurable change in its molecular weight. MATERIALS AND METHODSChicks, Injections, and RNA Labelin...

Transcription of genetic information in amphibian oocytes

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