Ribonuclease Dicer Cleaves Triplet Repeat Hairpins into Shorter Repeats that Silence Specific Targets

Król, Jacek; Fiszer, Agnieszka; Mykowska, Agnieszka; Sobczak, Krzysztof; Mezer, Mateusz de; Krzyżosiak, Włodzimierz J.

doi:10.1016/j.molcel.2007.01.031

Cited by 176 publications

(185 citation statements)

References 38 publications

(49 reference statements)

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“…It is also known that MBNL1 interacts mainly with the GC element of the repeated sequence (45). The structural similarity of all CNG repeat hairpins was also observed in the in vitro assay with ribonuclease Dicer (46). On the other hand, these hairpin structures are different targets for the binding of small molecule ligands (47,48).…”

Section: Discussionmentioning

confidence: 79%

Structural Diversity of Triplet Repeat RNAs

Sobczak

Michlewski

Mezer

et al. 2010

Journal of Biological Chemistry

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116

153

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Tandem repeats of various trinucleotide motifs are present in the human transcriptome, but the functions of these regular sequences, which likely depend on the structures they form, are still poorly understood. To gain new insight into the structural and functional properties of triplet repeats in RNA, we have performed a biochemical structural analysis of the complete set of triplet repeat transcripts, each composed of a single sequence repeated 17 times. We show that these transcripts fall into four structural classes. The repeated CAA, UUG, AAG, CUU, CCU, CCA, and UAA motifs did not form any higher order structure under any analyzed conditions. The CAU, CUA, UUA, AUG, and UAG repeats are ordered according to their increasing tendency to form semistable hairpins. The repeated CGA, CGU, and all CNG motifs form fairly stable hairpins, whereas AGG and UGG repeats fold into stable G-quadruplexes. The triplet repeats that formed the most stable structures were characterized further by biophysical methods. UV-monitored structure melting revealed that CGG and CCG repeats form, respectively, the most and least stable hairpins of all CNG repeats. Circular dichroism spectra showed that the AGG and UGG repeat quadruplexes are formed by parallel RNA strands. Furthermore, we demonstrated that the different susceptibility of various triplet repeat transcripts to serum nucleases can be explained by the sequence and structural features of the tested RNAs. The results of this study provide a comprehensive structural foundation for the functional analysis of triplet repeats in transcripts.Trinucleotide repeats (TNRs) 4 belong to a family of microsatellite sequences also known as short tandem repeats or simple sequence repeats. TNRs are present in both prokaryotic and eukaryotic genomes and, similar to other microsatellites, show high mutation rates resulting in frequent length variability. Polymorphic TNRs are better tolerated than dinucleotide and tetranucleotide repeats in translated sequences because their length variation does not change the open reading frame.A recent survey of the human genome reference sequence showed that it harbors more than 32,000 tracts of uninterrupted TNR sequences composed of six or more repeated units. In annotated human exons, which account for less than 3% of the genomic sequence, there are as many as 1,030 TNR tracts (61). Some AT-rich TNR types, such as CTT, AAC, and AAT, are particularly underrepresented in exons, whereas GC-rich repeats (CGG, CAG, and CCG) are highly overrepresented, implying that these sequences have a functional significance. The AT-rich and GC-rich TNRs tend to localize preferentially in the 3Ј-UTR and 5Ј-UTR, respectively (1). About 60% of exonic TNRs are localized in the open reading frame. These coding repeats are primarily translated to the poly-Gln, polyAla, poly-Glu, and poly-Leu tracts. However, the amino acid coding property of TNRs is not the only feature for which these sequences are selected in exons. The other properties of TNR sequences that manifest thems...

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Section: Discussionmentioning

confidence: 79%

Structural Diversity of Triplet Repeat RNAs

Sobczak

Michlewski

Mezer

et al. 2010

Journal of Biological Chemistry

Self Cite

116

153

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“…18,20,24 It has also been shown that long CUG repeats degrade, resulting in the accumulation of short CUG repeats detectable in DM1 cells. 42 Because the accumulation of short CUG repeats causes the DM1 phenotype in DM1 murine models, 24 we decided to determine whether high levels of short CUG repeats would elevate CUGBP1. Analysis of CUGBP1 expression after a single pulse of transcription of CUG 25 showed that CUGBP1 is elevated in cytoplasm at 7 hours after Dox addition ( Figure 4A).…”

Section: Cugbp1 Elevation Is a Primary Event Of The Accumulation Of Tmentioning

confidence: 99%

RNA Foci, CUGBP1, and ZNF9 Are the Primary Targets of the Mutant CUG and CCUG Repeats Expanded in Myotonic Dystrophies Type 1 and Type 2

Jones

Jin

Iakova

et al. 2011

The American Journal of Pathology

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Expansions of noncoding CUG and CCUG repeats in myotonic dystrophies type 1 (DM1) and DM2 cause complex molecular pathology, the features of which include accumulation of RNA aggregates and misregulation of the RNA-binding proteins muscleblind-like 1 (MBNL1) and CUG-binding protein 1 (CUGBP1). CCUG repeats also decrease amounts of the nucleic acid binding protein ZNF9. Using tetracycline (Tet)-regulated monoclonal cell models that express CUG and CCUG repeats, we found that low levels of long CUG and CCUG repeats result in nuclear and cytoplasmic RNA aggregation with a simultaneous increase of CUGBP1 and a reduction of ZNF9. Elevation of CUGBP1 and reduction of ZNF9 were also observed before strong aggregation of the mutant CUG/CCUG repeats. Degradation of CUG and CCUG repeats normalizes ZNF9 and CUGBP1 levels. Comparison of short and long CUG and CCUG RNAs showed that great expression of short repeats form foci and alter CUGBP1 and ZNF9; however, long CUG/CCUG repeats misregulate CUGBP1 and ZNF9 much faster than high levels of the short repeats. These data suggest that correction of DM1 and DM2 might be achieved by complete and efficient degradation of CUG and CCUG repeats or by a simultaneous disruption of CUG/CCUG foci and correction of CUGBP1 and ZNF9. (Am J Pathol

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“…Nonviral approaches to reduce CUG exp RNA include short interfering RNA (siRNA), which has demonstrated efficacy in DM1 fibroblasts. 64 Together, these data support a role for degradation of the mutant RNA as a potential therapeutic strategy in DM1. Clinical application of these strategies necessitate development of efficient means of systemic delivery of viral-mediated antisense, siRNA, or both.…”

Section: Reduction Of Rna-mediated Toxicitymentioning

confidence: 77%

Myotonic Dystrophy: Therapeutic Strategies for the Future

Wheeler

2008

Neurotherapeutics

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Summary: Myotonic dystrophy (DM) is a dominantly inherited neurodegenerative disorder for which there is no cure or effective treatment. Investigation of DM pathogenesis has identified a novel disease mechanism that requires development of innovative therapeutic strategies. It is now clear that DM is not caused by expression of a mutant protein. Instead, DM is the first recognized example of an RNA-mediated disease. Expression of the mutated gene gives rise to an expanded repeat RNA that is directly toxic to cells. The mutant RNA is retained in the nucleus, forming ribonuclear inclusions in affected tissue. A primary consequence of RNA toxicity in DM is dysfunction of two classes of RNA binding proteins, which leads to abnormal regulation of alternative splicing, or spliceopathy, of select genes. Spliceopathy now is known to cause myotonia and insulin resistance in DM. As our understanding of pathogenesis continues to improve, therapy targeted directly at the RNA disease mechanism will begin to replace the supportive care currently available. New pharmacologic approaches to treat myotonia and muscle wasting in DM type 1 are already in early clinical trials, and therapies designed to reverse the RNA toxicity have shown promise in preclinical models by correcting spliceopathy and eliminating myotonia. The well-defined ribonuclear inclusions may serve as convenient therapeutic targets to identify new agents that modify RNA toxicity. Continued development of appropriate model systems will allow testing of additional therapeutic strategies as they become available. Although DM is a decidedly complex disorder, its RNA-mediated disease mechanism may prove to be highly susceptible to therapy.

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Ribonuclease Dicer Cleaves Triplet Repeat Hairpins into Shorter Repeats that Silence Specific Targets

Cited by 176 publications

References 38 publications

Structural Diversity of Triplet Repeat RNAs

Structural Diversity of Triplet Repeat RNAs

RNA Foci, CUGBP1, and ZNF9 Are the Primary Targets of the Mutant CUG and CCUG Repeats Expanded in Myotonic Dystrophies Type 1 and Type 2

Myotonic Dystrophy: Therapeutic Strategies for the Future

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