Autoregulation of MBNL1 function by exon 1 exclusion from MBNL1 transcript

Konieczny, Patryk; Stepniak-Konieczna, Ewa; Taylor, Katarzyna; Sznajder, Łukasz J.; Sobczak, Krzysztof

doi:10.1093/nar/gkw1158

Cited by 42 publications

(54 citation statements)

References 66 publications

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“…To gain further insight, we first asked whether alterations of RNA secondary structure encompassing MBNL-binding motifs modify the MBNL1 affinity and its ability to regulate AS. For this purpose, we selected three MBNL-sensitive pre-mRNA regulatory elements from Mbnl1 exon1 (E1) ( 45 ), Atp2a1 intron 22 (I22) and Pphln1 intron 6 (I6) ( 44 ). Each of these AS substrates contains a few conserved MBNL-recognized 5′-YGCY-3′ motifs ( 33 , 55 ).…”

Section: Resultsmentioning

confidence: 99%

“…The analysis was conducted as previously described ( 45 ) with concentrations of S1, T1 and T2 endonucleases indicated in Supplementary Figures S1a, S3a, S8a and S9a . The analyzed RNA sequences were also run through the following secondary structure prediction software programs: RNAfold ( 46 ), Mfold ( 47 ), RNAstructure ( 48 ), using biochemically defined constrains or default settings.…”

Section: Methodsmentioning

confidence: 99%

“…Co-transfection was conducted with 200 ng of minigene and 500 ng of eGFP-MBNL, mCherry or eGFP expressing vectors (or as indicated in the figures). eGFP-MBNL constructs were previously described ( 30 , 45 ). Competition assay was conducted with 200 ng of minigene, 250 ng of MBNL1 (different isoforms) and increasing concentrations of MBNL3 or MBNL1 (different isoforms) as indicated in the figures, and substituted with eGFP.…”

Section: Methodsmentioning

confidence: 99%

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MBNL splicing activity depends on RNA binding site structural context

Taylor

Sznajder

Cywoniuk

et al. 2018

Nucleic Acids Research

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Muscleblind-like (MBNL) proteins are conserved RNA-binding factors involved in alternative splicing (AS) regulation during development. While AS is controlled by distribution of MBNL paralogs and isoforms, the affinity of these proteins for specific RNA-binding regions and their location within transcripts, it is currently unclear how RNA structure impacts MBNL-mediated AS regulation. Here, we defined the RNA structural determinants affecting MBNL-dependent AS activity using both cellular and biochemical assays. While enhanced inclusion of MBNL-regulated alternative exons is controlled by the arrangement and number of MBNL binding sites within unstructured RNA, when these sites are embedded in a RNA hairpin MBNL binds preferentially to one side of stem region. Surprisingly, binding of MBNL proteins to RNA targets did not entirely correlate with AS efficiency. Moreover, comparison of MBNL proteins revealed structure-dependent competitive behavior between the paralogs. Our results showed that the structure of targeted RNAs is a prevalent component of the mechanism of alternative splicing regulation by MBNLs.

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

See 1 more Smart Citation

MBNL splicing activity depends on RNA binding site structural context

Taylor

Sznajder

Cywoniuk

et al. 2018

Nucleic Acids Research

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“…To test whether circRNA levels are decreased in DM1 and to verify the role of MBNLs in the biogenesis of circRNA, we analyzed the expression level of up to twenty circRNAs in myoblast cell lines and skeletal muscle samples derived from patients with DM1 and healthy controls. Among the selected circRNAs were those with a relatively high number (n≥10) of potential MBNL-binding motifs in flanking introns, as well as circMBNL1, which is regulated by MBNL1 (37). Additionally, circCDR1as and circHIPK3, the highly expressed and most extensively studied circRNAs, were among the selected circRNAs (17,23,24).…”

Section: Circrna Levels Are Associated With Dm Severitymentioning

confidence: 99%

Global increase in circRNA levels in myotonic dystrophy

Czubak

Taylor

Piasecka

et al. 2018

Preprint

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Splicing aberrations induced as a consequence of the sequestration of MBNL splicing factors on the DMPK transcript, which contains expanded CUG repeats, present a major pathomechanism of myotonic dystrophy type 1 (DM1). As MBNLs may also be important factors involved in the biogenesis of circular RNAs (circRNAs), we hypothesized that the level of circRNAs would be decreased in DM1. To test this hypothesis, we selected twenty well-validated circRNAs and analyzed their levels in several experimental systems (e.g., cell lines, DM muscle tissues, and a mouse model of DM1) using droplet digital PCR assays.We also explored the global level of circRNAs using two RNA-Seq datasets of DM1 muscle samples. Contrary to our original hypothesis, our results consistently showed a global increase in circRNA levels in DM1 and we identified numerous circRNAs that were increased in DM1. We also identified many genes (including muscle-specific genes) giving rise to numerous (>10) circRNAs. Thus, this study is the first to show an increase in global circRNA levels in DM1. We also provided preliminary results showing the association of circRNA level with muscle weakness and alternative splicing changes that are biomarkers of DM1 severity. Author SummaryRecently, a great deal of interest has been focused on a new class of RNA molecules called circular RNAs (circRNAs). To date, thousands of circRNAs have been found in different human cells/tissues. Although the function of circRNAs remains mostly unknown, circRNAs have emerged as an important component of the RNA-RNA and RNA-protein interactome.Thus, intensive efforts are being made to fully understand the biology and function of circRNAs, especially their role in human diseases. As an important role in the biogenesis of circRNA may be played by MBNL splicing factors, in this study we used DM1 (to a lesser extent, DM2) as a natural model in which the level of MBNLs is decreased. In contrast to the expected effect, our results consistently showed a global increase in circRNA levels in DM1.As a consequence, whole genome transcriptome analysis revealed dozens of circRNAs with significantly altered (mostly increased) levels in DM1. Furthermore, we observed that the circRNA levels were in many cases strongly associated with DM1 severity.

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“…The production of the mbl circRNA is regulated by Mbl protein itself, in an autoregulatory feedback loop. In vertebrates it appears that MBNL1 does not regulate production of its own circRNA in neuronal cells, but the effect in muscle has not been determined [19,45]. Our discovery that defects in an RNA export factor, Nxt1, can generate a muscular dystrophy phenotype in Drosophila suggest that investigation of the RNA export pathway in context of the disease is warranted.…”

Section: Discussionmentioning

confidence: 99%