Properties of a small transcribed poly A sequence in heterogeneous nuclear RNA of HeLa cells

Venkatesant, S.; Nakazato, Hiroshi; Kopp, David W.; Edmonds, Mary

doi:10.1093/nar/6.3.1097

Cited by 15 publications

(2 citation statements)

References 19 publications

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“…Recent sequencing of the 3' end of the large rRNA in hamster mitochondria has revealed some sequence heterogeniety due in part to the presence of oligo(A) (10). Oligo(A) has also been found in the heterogeneous nuclear RNA of HeLa cells and is thought to be synthesized by transcription (40). Our detection of short adenylate-rich oligonucleotides in yeast mitochondria is not very surprising in view of the high AT content of mitochondrial DNA (1) and the presence of short adenylate runs adjacent to the amino acid sequence information in several genes (7,20,24,25).…”

Section: Resultsmentioning

confidence: 99%

Oligoadenylate is present in the mitochondrial RNA of Saccharomyces cerevisiae.

Yuckenberg¹,

Phillips²

1982

Mol. Cell. Biol.

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We examined Saccharomyces cerevisiae mitochondrial RNA for polyadenylate. Using hybridization to [3H]polyuridylate as the assay for adenylate sequences, we found adenylate-rich oligonucleotides approximately 8 residues long. Longer polyadenylate was not detected. Most of the adenylate-rich sequence is associated with the large mitochondrial rRNA. The remainder is associated with the 10-12S group of transcripts.In mammalian cells polyadenylation has been associated with the production of functional cytoplasmic mRNA from nuclear precursors (3,8,30) as well as with its stabilization (14,22,26,38). Polyadenylated RNA has been shown to be present in animal cell mitochondria (21, 31). Although it is quite likely that nucleolytic processing of mammalian mitochondrial RNA occurs (32), the role polyadenylation may have in this process is unknown.The unicellular eucaryote Saccharomyces cerevisiae also contains 3' polyadenylated cytoplasmic mRNA. However, there is disagreement in the literature regarding the presence of polyadenylate [poly(A)] in S. cerevisiae mitochondrial RNA (6,17,19,29,33). It is clear from the recent dramatic advances in the analysis of mitochondrial DNA and its transcripts that the production of several mitochondrial RNA species in S. cerevisiae involves extensive posttranscriptional RNA processing (4,18,24,39). Therefore, we reexamined S. cerevisiae mitochondrial RNA for the presence of poly(A), with an emphasis on procedures which permit the detection of short adenylate sequences. We found that S. cerevisiae mitochondrial RNA contains an oligonucleotide sequence approximately 8 nucleotides long that is composed of at least 80% adenylate residues. The bulk of this adenylate-rich oligonucleotide is associated with the large 21S rRNA. Smaller amounts are associated with the 15S RNA and the 10-12S RNA species. Longer poly(A) was not detected in mitochondrial RNA under conditions where cytoplasmic poly(A) is readily observed. MATERIALS AND METHODSCefl culture. S. cerevisiae IL8-8c (at his-trp-CAPr Ery'), used throughout these studies, was a generous Preparation of RNA. Mitochondria were purified by an adaptation of the procedure of Kovac et al. (23). A routine preparation started with the cells from 2 liters of culture. Cells were harvested by centrifugation at 9,000 x g for 10 min and washed by two cycles of resuspension in 500 ml of water and centrifugation. Cells were converted to spheroplasts by treatment at 30C for 30 min with 20 mg of Zymolyase 60,000 in 200 ml of t M sorbitol-20 mM sodium phosphate (pH 7.0) or with 2 ml of Glusulase in 200 ml of 1 M sorbitol.Spheroplasts were recovered and washed free of soluble materials by sedimentation at 1,500 x g for 10 min through a 0.5-volume layer of 2 M sorbitol. The spheroplasts were resuspended in 1 liter of growth medium containing 0.5% galactose and 1 M sorbitol for osmotic stabilization and incubated for 1 h at 30°C to ensure recovery from the metabolic perturbation created during cell wall digestion. The culture was quickly chilled by pouring into ...

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Section: Resultsmentioning

confidence: 99%

Oligoadenylate is present in the mitochondrial RNA of Saccharomyces cerevisiae.

Yuckenberg¹,

Phillips²

1982

Mol. Cell. Biol.

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“…This oligo(dA) tract may be an important template for the small, rapidly-labeled, cordycepin-resistant internal poly(A) sequence found in hnRNA (27,28,29). These oligoadenylate sequences need not serve as a starting point for the addition of a longer (ca.…”

Section: Discussionmentioning

confidence: 99%

Transcriptional analysis of interspersed repetitive polymerase III transcription units in human DNA

et al. 1981

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The template for RNA polymerase III in vitro transcription found on the human DNA clone pJP53 was shown in the previous paper to enclose a member of the Alu famiy of interspersed repetitive DNA sequences. We have mapped this transcript onto its template in greater detail by comparison of the template DNA sequence to the base composition of the Tl ribonuclease digestion products of the in vitro transcript. We find that the 5' end of the transcript lies in close proximity to the 5' end of the conserved Alu family sequence as analyzed in the preceding paper. The 3' end of the transcript appears to terminate in a U-rich region beyond the region of Alu family sequence conservation. Analysis of cellular RNA by Northern blotting and hybridization with a DNA probe derived from another Alu family transcription template demonstrates abundant representation of sequences homologous to the reiterated DNA. Cytoplasmic, nonpolyadenylated RNA from human and murine cells contains a monodisperse, 300 nucleotide species, recently determined by Weiner (4) to be the 7S RNA. In contrast, the Alu-homologous transcripts are heterodisperse in mRNA and hnRNA, with the highest specific representation of Alu family sequences being found in oligo(dT)-retained hnRNA.

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