Sequence Dependence of Substrate Recognition and Cleavage by Yeast RNase III

Lamontagne, Bruno; Ghazal, Ghada; Lebars, Isabelle; Yoshizawa, Satoko; Fourmy, Dominique; Elela, Sherif Abou

doi:10.1016/s0022-2836(03)00231-6

Cited by 39 publications

(110 citation statements)

References 49 publications

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“…Although, Rnt1p has been shown to form a stable RNP complex with short RNA hairpins in vitro (Lamontagne et al, 2003), this phenomenon has yet to be demonstrated in vivo. Indeed, demonstrating the effects of Rnt1p-RNA interaction on the cell functions would be difficult because all mutations that affect RNA binding also impair RNA cleavage (Lamontagne et al, 2000;Tremblay et al, 2002) or perturb Rnt1p localization (our unpublished data).…”

Section: Novel Functions Of Rnt1p Unrelated To Rna Processingmentioning

confidence: 99%

Cell Cycle-dependent Nuclear Localization of Yeast RNase III Is Required for Efficient Cell Division

Catala

Lamontagne

Larose

et al. 2004

MBoC

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Members of the double-stranded RNA-specific ribonuclease III (RNase III) family were shown to affect cell division and chromosome segregation, presumably through an RNA interference-dependent mechanism. Here, we show that in Saccharomyces cerevisiae, where the RNA interference machinery is not conserved, an orthologue of RNase III (Rnt1p) is required for progression of the cell cycle and nuclear division. The deletion of Rnt1p delayed cells in both G1 and G2/M phases of the cell cycle. Nuclear division and positioning at the bud neck were also impaired in ⌬rnt1 cells. The cell cycle defects were restored by the expression of catalytically inactive Rnt1p, indicating that RNA cleavage is not essential for cell cycle progression. Rnt1p was found to exit from the nucleolus to the nucleoplasm in the G2/M phase, and perturbation of its localization pattern delayed the progression of cell division. A single mutation in the Rnt1p N-terminal domain prevented its accumulation in the nucleoplasm and slowed exit from mitosis without any detectable effects on RNA processing. Together, the data reveal a new role for a class II RNase III in the cell cycle and suggest that at least some members of the RNase III family possess catalysis-independent functions.

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Section: Novel Functions Of Rnt1p Unrelated To Rna Processingmentioning

confidence: 99%

Cell Cycle-dependent Nuclear Localization of Yeast RNase III Is Required for Efficient Cell Division

Catala

Lamontagne

Larose

et al. 2004

MBoC

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“…RNA transcripts used for cleavage were generated and 5Ј-end labeled with ␥-32 P-ATP as described before Lamontagne et al 2003). In vitro cleavage using purified Rnt1p was performed as described by Lamontagne and Abou Elela (2001).…”

Section: In Vitro Analysis Of Rnt1p Activitymentioning

confidence: 99%

The RNA catabolic enzymes Rex4p, Rnt1p, and Dbr1p show genetic interaction with trans-acting factors involved in processing of ITS1 in Saccharomyces cerevisiae pre-rRNA

Faber¹,

Vos

Vos³

et al. 2004

RNA

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Eukaryotes have two types of ribosomes containing either 5.8S L or 5.8S S rRNA that are produced by alternative pre-rRNA processing. The exact processing pathway for the minor 5.8S L rRNA species is poorly documented. We have previously shown that the trans-acting factor Rrp5p and the RNA exonuclease Rex4p genetically interact to influence the ratio between the two forms of 5.8S rRNA in the yeast Saccharomyces cerevisiae. Here we report a further analysis of ITS1 processing in various yeast mutants that reveals genetic interactions between, on the one hand, Rrp5p and RNase MRP, the endonuclease required for 5.8S S rRNA synthesis, and, on the other, Rex4p, the RNase III homolog Rnt1p, and the debranching enzyme Dbr1p. Yeast cells carrying a temperature-sensitive mutation in RNase MRP (rrp2-1) exhibit a pre-rRNA processing phenotype very similar to that of the previously studied rrp5-⌬3 mutant: ITS2 processing precedes ITS1 processing, 5.8S L rRNA becomes the major species, and ITS1 is processed at the recently reported novel site A4 located midway between sites A2 and A3. As in the rrp5-⌬3 mutant, all of these phenotypical processing features disappear upon inactivation of the REX4 gene. Moreover, inactivation of the DBR1 gene in rrp2-1, or the RNT1 gene in rrp5-⌬3 mutant cells also negates the effects of the original mutation on pre-rRNA processing. These data link a total of three RNA catabolic enzymes, Rex4p, Rnt1p, and Dbr1p, to ITS1 processing and the relative production of 5.8S S and 5.8S L rRNA. A possible model for the indirect involvement of the three enzymes in yeast pre-rRNA processing is discussed.

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“…One explanation of this phenomenon is that Dicer binding itself stabilizes the complex formed between the guide and the target RNA, while increased stability of the bipartite complex reduces product release. Previous studies using yeast RNase III indicated that strengthening the RNA duplex formation by increasing the GC content often leads to an increase in the binding affinity and a decrease in the catalytic rate (Lamontagne et al 2003).…”

Section: Discussionmentioning

confidence: 99%

Short RNA duplexes guide sequence-dependent cleavage by human Dicer

Bergeron

Perreault²,

Elela³

2010

RNA

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Dicer is a member of the double-stranded (ds) RNA-specific ribonuclease III (RNase III) family that is required for RNA processing and degradation. Like most members of the RNase III family, Dicer possesses a dsRNA binding domain and cleaves long RNA duplexes in vitro. In this study, Dicer substrate selectivity was examined using bipartite substrates. These experiments revealed that an RNA helix possessing a 2-nucleotide (nt) 39-overhang may bind and direct sequence-specific Dicer-mediated cleavage in trans at a fixed distance from the 39-end overhang. Chemical modifications of the substrate indicate that the presence of the ribose 29-hydroxyl group is not required for Dicer binding, but some located near the scissile bonds are needed for RNA cleavage. This suggests a flexible mechanism for substrate selectivity that recognizes the overall shape of an RNA helix. Examination of the structure of natural pre-microRNAs (pre-miRNAs) suggests that they may form bipartite substrates with complementary mRNA sequences, and thus induce seed-independent Dicer cleavage. Indeed, in vitro, natural pre-miRNA directed sequence-specific Dicer-mediated cleavage in trans by supporting the formation of a substrate mimic.

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Sequence Dependence of Substrate Recognition and Cleavage by Yeast RNase III

Cited by 39 publications

References 49 publications

Cell Cycle-dependent Nuclear Localization of Yeast RNase III Is Required for Efficient Cell Division

Cell Cycle-dependent Nuclear Localization of Yeast RNase III Is Required for Efficient Cell Division

The RNA catabolic enzymes Rex4p, Rnt1p, and Dbr1p show genetic interaction with trans-acting factors involved in processing of ITS1 in Saccharomyces cerevisiae pre-rRNA

Short RNA duplexes guide sequence-dependent cleavage by human Dicer

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