PMC:1459172 / 28668-31391 JSONTXT

{"project":"2_test","denotations":[{"id":"16722605-15716109-1692569","span":{"begin":199,"end":201},"obj":"15716109"},{"id":"16722605-15457527-1692570","span":{"begin":383,"end":385},"obj":"15457527"},{"id":"16722605-11222752-1692571","span":{"begin":389,"end":391},"obj":"11222752"},{"id":"16722605-15014042-1692572","span":{"begin":1900,"end":1902},"obj":"15014042"}],"text":"Applications\nIntermolecular binding of RNA molecules is important in a broad spectrum of cases, ranging from mRNA accessibility to siRNA or miRNA binding, RNA probe design, or designing RNA openers [50]. An important question that arises repeatedly is to explain differences in RNA-RNA binding between seemingly very similar or even identical binding sites. As demonstrated e.g. in [22,29,51,52], different RNA secondary structure of the target molecule can have dramatic effects on binding affinities even if the sequence of the binding site is identical.\nSince the comparison of base pairing patterns is a crucial step in such investigations we provide a tool for graphically comparing two dot plots, see Fig. 4. It is written in Perl-Tk and takes two dot plot files and, optionally, an alignment file as input. The differences between the two dot plots are displayed in color-code, the dot plot is zoomable and the identity and probability(-difference) of a base pair is displayed when a box is clicked.\nFigure 4 Difference dot Plot of native and mutated secondary structure of a 3 GU mutation of the CXCR4 siRNA gene. The red part on the right hand side shows the base pairing probability of the 5' part of the micro RNA, which is 80% higher in the native structure. This is an alternative explanation for the missing function of the mutant. Because of the mutations, the stack a little to the left gets more stable, and the probability of binding of the 5' end of the siRNA is reduced significantly.The color of the dots encodes the difference of the pair probabilities in the two molecules such positive (red) squares denote pairs more more probable in the second molecule (see color bar). The area of the dots is proportional to the larger of the two pair probabilities. As a simple example for the applicability of RNAcofold, we re-evaluate here parts of a recent study by Doench and Sharp [53]. In this work, the influence of GU base pairs on the effectivity of translation attenuation by miRNAs is assayed by mutating binding sites and comparing attenuation effectivity to wild type binding sites\nIntroducing three GU base pairs into the mRNA/miRNA duplex did, with only minor changes to the binding energy, almost completely destroy the functionality of the binding site. While Doench and Sharp concluded that miRNA binding sites are not functional because of the GU base pairs, testing the dimer with RNAcofold shows that there is also a significant difference in the cofolding structure that might account for the activity difference without invoking sequence specificities: Because of the secondary structure of the target, the binding at the 5' end of the miRNA is much weaker than in the wild type, Fig. 4."}