Point Mutations Upstream of the Yeast <i>ADH2</i> Poly(A) Site Significantly Reduce the Efficiency of 3′-End Formation

Hyman, Linda E.; Seiler, Stephanie H.; Whoriskey, John; Moore, Claire

doi:10.1128/mcb.11.4.2004-2012.1991

Cited by 13 publications

(13 citation statements)

References 43 publications

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“…Plasmid pL101 and ADH2 3′-end sequence was described previously (23) and is shown in Figure 1A. The plasmid pBEVY-U (Fig.…”

Section: Plasmid Constructionmentioning

confidence: 99%

“…Yeast total RNA was prepared according to the described protocol (32). RNAs were separated by electrophoresis on 1.5% agarose-formaldehyde gels and northern analyses were carried out as described (23). Mapping the 3′ ends of mRNAs was described previously (33).…”

Section: Yeast Total Rna Preparation Northern Blot Analysis and Rt-pc...mentioning

confidence: 99%

“…Previous studies have demonstrated that the sequences located at the end of the ADH2 gene constitute a strong termination signal (23,24). By examining a 327 nt fragment inserted within an intron in a chimeric gene linked to LacZ (Fig.…”

Section: The Effect Of the (Ua) 3 Deletion On 3′-end Formation In Vivomentioning

confidence: 99%

“…Tel: +1 504 584 2941; Fax: +1 504 584 2739; Email: lhyman@mailhost.tcs.tulane.edu (17)(18)(19)(20). Mutation of the cis-acting signal, critical for cleavage and polyadenylation in mammalian cells and yeast results in aberrant termination of transcription by RNA polymerase II (21)(22)(23)(24). However the most direct evidence for coupling of the processing machinery with transcription termination comes from two recent observations.…”

Section: Introductionmentioning

confidence: 99%

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A specific RNA-protein interaction at yeast polyadenylation efficiency elements

Chen

Hyman

1998

Nucleic Acids Research

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The specific RNA-protein interactions responsible for the production of mature 3' ends of eukaryotic mRNAs are not well understood. Sequence elements at the 3' ends of yeast genes have been identified that specify the position of the poly(A) site and the efficiency of polyadenylation. To provide additional insights into the interaction between important sequences that direct 3'-end formation in vivo and nuclear proteins, we utilized gel mobility shift assays and UV-crosslinking studies. The data indicate that a protein, with an apparent molecular weight of 80 kDa, interacts specifically with pre-mRNA at the (UA)3efficiency element. Although the interaction is specific, it can be competed by RNA sequences that do not contain the same type of efficiency element; that is, a sequence lacking a (UA)3repeat. This result implies that the protein binding site is flexible. Using immunoprecipitation techniques, the protein has been identified as Hrp1, a heteronuclear RNA binding protein. The role of Hrp1p in 3'-end formation including RNA processing and transcription termination is addressed.

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“…Plasmid pL101 and ADH2 3′-end sequence was described previously (23) and is shown in Figure 1A. The plasmid pBEVY-U (Fig.…”

Section: Plasmid Constructionmentioning

confidence: 99%

Section: Yeast Total Rna Preparation Northern Blot Analysis and Rt-pc...mentioning

confidence: 99%

Section: The Effect Of the (Ua) 3 Deletion On 3′-end Formation In Vivomentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

A specific RNA-protein interaction at yeast polyadenylation efficiency elements

Chen

Hyman

1998

Nucleic Acids Research

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“…An involvement of mRNA secondary structure has been proposed to explain the ability of some 3′ flanks to function in both orientations (8,28). In fact, inverted repeats have been described in these regions in yeast genes such as URA3 (32), ADH2 (33), GAL1, GAL7, GAL10 (8) and CYC1 (34), but a requirement for such secondary structures has not been demonstrated. The ability to direct transcription termination in both orientations has only been studied systematically for CYC1 (25,26).…”

Section: Introductionmentioning

confidence: 99%

The yeast FBP1 poly(A) signal functions in both orientations and overlaps with a gene promoter

Aranda

Pérez‐Ortín

Moore

et al. 1998

Nucleic Acids Research

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This report provides an analysis of a region of chromosome XII in which the FBP1 and YLR376c genes transcribe in the same direction. Our investigation indicates that the Saccharomyces cerevisiae FBP1 gene contains strong signals for polyadenylation and transcription termination in both orientations in vivo . A (TA)14 element plays a major role in directing polyadenylation in both orientations. While this region has four nonoverlapping copies of a TATATA hexanucleotide, which is a very potent polyadenylation efficiency element in yeast, it alone is not sufficient for full activation in the reverse orientation of a cluster of downstream poly(A) sites, and an additional upstream sequence is required. The putative RNA hairpin formed from the (TA)14 element is not involved in 3'-end formation. Surprisingly, deletion of the entire (TA)14 stretch affects transcription termination in the reverse orientation, in contrast to our previous results with the forward orientation, indicating that the transcription termination element operating in the reverse orientation has very different sequence requirements. Promoter elements for the YLR376c gene overlap with the signal for FBP1 3'-end formation. To our knowledge, this is the first time that overlapping of both types of regulatory signals has been found in two adjacent yeast genes.

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How is polyadenylation restricted to 3′‐untranslated regions?

Struhl

2023

Yeast

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Polyadenylation occurs at numerous sites within 3′‐untranslated regions (3′‐UTRs) but rarely within coding regions. How does Pol II travel through long coding regions without generating poly(A) sites, yet then permits promiscuous polyadenylation once it reaches the 3′‐UTR? The cleavage/polyadenylation (CpA) machinery preferentially associates with 3′‐UTRs, but it is unknown how its recruitment is restricted to 3′‐UTRs during Pol II elongation. Unlike coding regions, 3′‐UTRs have long AT‐rich stretches of DNA that may be important for restricting polyadenylation to 3′‐UTRs. Recognition of the 3′‐UTR could occur at the DNA (AT‐rich), RNA (AU‐rich), or RNA:DNA hybrid (rU:dA‐ and/or rA:dT‐rich) level. Based on the nucleic acid critical for 3′‐UTR recognition, there are three classes of models, not mutually exclusive, for how the CpA machinery is selectively recruited to 3′‐UTRs, thereby restricting where polyadenylation occurs: (1) RNA‐based models suggest that the CpA complex directly (or indirectly through one or more intermediary proteins) binds long AU‐rich stretches that are exposed after Pol II passes through these regions. (2) DNA‐based models suggest that the AT‐rich sequence affects nucleosome depletion or the elongating Pol II machinery, resulting in dissociation of some elongation factors and subsequent recruitment of the CpA machinery. (3) RNA:DNA hybrid models suggest that preferential destabilization of the Pol II elongation complex at rU:dA‐ and/or rA:dT‐rich duplexes bridging the nucleotide addition and RNA exit sites permits preferential association of the CpA machinery with 3′‐UTRs. Experiments to provide evidence for one or more of these models are suggested.

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Point Mutations Upstream of the Yeast ADH2 Poly(A) Site Significantly Reduce the Efficiency of 3′-End Formation

Cited by 13 publications

References 43 publications

A specific RNA-protein interaction at yeast polyadenylation efficiency elements

A specific RNA-protein interaction at yeast polyadenylation efficiency elements

The yeast FBP1 poly(A) signal functions in both orientations and overlaps with a gene promoter

How is polyadenylation restricted to 3′‐untranslated regions?

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