The primary transcription unit of the human (eqn)a2 globin gene defined by quantitative RT/PCR

Owczarek, Catherine M.; Enriquez-Harris, Pita; Proudfoot, Nick J.

doi:10.1093/nar/20.4.851

Cited by 10 publications

(7 citation statements)

References 28 publications

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“…Radioactive PCR products were excised and quantitated. The radioactivity of each PCR product is given as a percentage of that recorded for the PCR product produced with oligonucleotide 1. between RNA input and PCR product was determined, as described in Owczarek et al (1992) and was found to exist after 20 cycles of amplification, for 50 ng of S.pombe RNA (data not shown). Therefore a measure of the relative number of transcripts was made by comparing the counts incorporated into the PCR products separated electrophoretically ( Figure 2B) after 20 cycles and is presented in Figure 2A.…”

Section: Resultsmentioning

confidence: 99%

RNA 3′ end signals of the S.pombe ura4 gene comprise a site determining and efficiency element.

1994

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We have defined sequences in the 3′ non‐coding region of the Schizosaccharomyces pombe ura4 gene that are required for efficient mRNA 3′ end formation. Three separate sequence elements have been identified. Two of these are site determining elements which specify alternative sites of polyadenylation [the major poly(A) site and a minor downstream poly(A) site]. The third sequence, located downstream of both poly(A) sites, functions as an efficiency element that enhances utilization of either polyadenylation site. By employing sensitive RT‐PCR analysis, we demonstrate that although low levels of transcripts are detected up to the efficiency element, none is detected beyond this point. The downstream site determining element and efficiency element have both been delineated to specific 16 nt sequences which we show are together sufficient for ura4 mRNA 3′ end formation. We have further characterized the interaction between these two elements and show that the efficiency element behaves in a position‐independent, orientation‐dependent manner, but cannot form 3′ ends independently of the site determining element. Surprisingly, we find that the efficiency element can be functionally replaced by a second copy of either site determining element. We present a model for the mechanism of RNA 3′ end formation of the ura4 gene and note that this bipartite structure for a poly(A) signal in S.pombe may be related to the AAUAAA and downstream GU‐rich sequences of poly(A) signals in mammalian genes.

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Section: Resultsmentioning

confidence: 99%

RNA 3′ end signals of the S.pombe ura4 gene comprise a site determining and efficiency element.

1994

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“…The first to be characterized was the ␣2 element (30), which very likely acts as an auxiliary element in poly(A)-dependent termination (61). In one report, run-on transcription of the cloned ␣2 gene showed pausing in the neighborhood of this element but no termination that was poly(A)-dependent (30).…”

Section: Poly(a)-dependent Pausing As a Checkpoint For Coupling Procementioning

confidence: 99%

“…More likely, as previously described for the CCAAT sequence (51) these elements operate as weak poly(A)-independent terminators when acting alone. Thus, whereas the ␣2 and C2 elements undoubtedly activate poly(A) signals (30,60,63) and probably also termination (30,60,61) and possibly even splicing (64), there is no consistent evidence that they actually pause RNA polymerase II in vivo or that their function is related to pausing.…”

Section: Poly(a)-dependent Pausing As a Checkpoint For Coupling Procementioning

confidence: 99%

The Poly(A) Signal, without the Assistance of Any Downstream Element, Directs RNA Polymerase II to Pause in Vivo and Then to Release Stochastically from the Template

Orozco

Kim

Martinson

2002

Journal of Biological Chemistry

114

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Genes encoding polyadenylated mRNAs depend on their poly(A) signals for termination of transcription. Typically, transcription downstream of the poly(A) signal gradually declines to zero, but often there is a transient increase in polymerase density immediately preceding the decline. Special elements called pause sites are traditionally invoked to account for this increase. Using run-on transcription from the nuclei of transfected cells, we show that both the pause and the gradual decline that follow a poly(A) site are generated entirely by the poly(A) signal itself in a series of model constructs. We found no other elements to be involved and argue that the elements called pause sites do not function through pausing. Both the poly(A)-dependent pause and the subsequent decline occurred earlier for a stronger poly(A) signal than for a weaker one. Because the gradual decline resembles the abortive elongation that occurs downstream of many promoters, one model has proposed that the poly(A) signal flips the polymerase from the elongation mode to the abortive mode like a binary switch. We compared abortive elongators with poly(A) terminators and found a 4-fold difference in processivity. We conclude that poly(A) terminating polymerases do not merely revert to their prior state of low processivity but rather convert to a new terminationprone condition.

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“…A few studies, however, have identified defined termination sequences at which efficient transcription termination occurs (Enriquez-Harris et al 1991;Owczarek et al 1992;Tantravahi et al 1993). These sequences are different in different genes, but a common feature is an A-rich region.…”

Section: Termination Of Transcriptionmentioning

confidence: 99%

Transcriptional termination in the Balbiani ring 1 gene is closely coupled to 3′-end formation and excision of the 3′-terminal intron

Baurén

Беликов²,

Wieslander³

1998

Genes Dev.

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We have analyzed transcription termination, 3 -end formation, and excision of the 3 -terminal intron in vivo in the Balbiani ring 1 (BR1) gene and its pre-mRNA. We show that full-length RNA transcripts are evenly spaced on the gene from a position 300 bp upstream to a region 500-700 bp downstream of the polyadenylation sequence. Very few full-length transcripts and no short, cleaved, nascent transcripts could be observed downstream of this region. Pre-mRNA with 10-20 adenylate residues accumulates at the active gene and then rapidly leaves from the gene locus. Only polyadenylated pre-mRNAs could be detected in the nucleoplasm. Our results are consistent with the hypothesis that transcription termination occurs in a narrow region for the majority of transcripts, simultaneous with 3 -end formation. Excision of the 3 -terminal intron occurs before 3 -end formation in about 5% of the nascent transcripts. When transcription terminates, 3 cleavage takes place and 10-20 adenylate residues are added, the 3 -terminal intron is excised from additionally about 75% of the pre-mRNA at the gene locus. Our data support a close temporal and spatial coupling of transcription termination and the cleavage and initial polyadenylation of 3 -end formation. Excision of the 3 -terminal intron is highly stimulated as the cleavage/polyadenylation complex assembles and 3 -end formation is initiated.

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The primary transcription unit of the human (eqn)a2 globin gene defined by quantitative RT/PCR

Cited by 10 publications

References 28 publications

RNA 3′ end signals of the S.pombe ura4 gene comprise a site determining and efficiency element.

RNA 3′ end signals of the S.pombe ura4 gene comprise a site determining and efficiency element.

The Poly(A) Signal, without the Assistance of Any Downstream Element, Directs RNA Polymerase II to Pause in Vivo and Then to Release Stochastically from the Template

Transcriptional termination in the Balbiani ring 1 gene is closely coupled to 3′-end formation and excision of the 3′-terminal intron

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