SUMMARY: We present the cpPredictor webserver that implements a novel template-based method for prediction of secondary structure of RNA. The method outperforms available prediction methods as it uses RNA structures of related molecules, either predicted or experimentally identified, as structural templates. The server aims at three major tasks: i) prediction of RNA secondary structures that are difficult to predict by available methods, ii) characterization of uncharacterized RNAs as compatible or incompatible with a chosen template structure and iii) an identification of the most relevant structure among different candidate structures of a single RNA ambiguously predicted by available methods. The web server is accompanied with a comprehensive documentation. AVAILABILITY AND IMPLEMENTATION: The web server is freely available at http://cppredictor.elixir-czech.cz/. The source code of the cpPredictor algorithm is freely available from the webserver under the Apache License, Version 2.0.

• MeSH
• algoritmy MeSH
• internet MeSH
• konformace nukleové kyseliny * MeSH
• RNA MeSH
• sekundární struktura proteinů MeSH
• sekvenční analýza RNA MeSH
• software * MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• RNA MeSH

BACKGROUND: Visualization of RNA secondary structures is a complex task, and, especially in the case of large RNA structures where the expected layout is largely habitual, the existing visualization tools often fail to produce suitable visualizations. This led us to the idea to use existing layouts as templates for the visualization of new RNAs similarly to how templates are used in homology-based structure prediction. RESULTS: This article introduces Traveler, a software tool enabling visualization of a target RNA secondary structure using an existing layout of a sufficiently similar RNA structure as a template. Traveler is based on an algorithm which converts the target and template structures into corresponding tree representations and utilizes tree edit distance coupled with layout modification operations to transform the template layout into the target one. Traveler thus accepts a pair of secondary structures and a template layout and outputs a layout for the target structure. CONCLUSIONS: Traveler is a command-line open source tool able to quickly generate layouts for even the largest RNA structures in the presence of a sufficiently similar layout. It is available at http://github.com/davidhoksza/traveler .

• Klíčová slova
• RNA secondary structure, Software tool, Template-based modeling, Visualization,
• MeSH
• algoritmy MeSH
• konformace nukleové kyseliny MeSH
• RNA chemie   MeSH
• software * MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• RNA MeSH

To maintain genome integrity, segmented double-stranded RNA viruses of the Reoviridae family must accurately select and package a complete set of up to a dozen distinct genomic RNAs. It is thought that the high fidelity segmented genome assembly involves multiple sequence-specific RNA-RNA interactions between single-stranded RNA segment precursors. These are mediated by virus-encoded non-structural proteins with RNA chaperone-like activities, such as rotavirus (RV) NSP2 and avian reovirus σNS. Here, we compared the abilities of NSP2 and σNS to mediate sequence-specific interactions between RV genomic segment precursors. Despite their similar activities, NSP2 successfully promotes inter-segment association, while σNS fails to do so. To understand the mechanisms underlying such selectivity in promoting inter-molecular duplex formation, we compared RNA-binding and helix-unwinding activities of both proteins. We demonstrate that octameric NSP2 binds structured RNAs with high affinity, resulting in efficient intramolecular RNA helix disruption. Hexameric σNS oligomerizes into an octamer that binds two RNAs, yet it exhibits only limited RNA-unwinding activity compared to NSP2. Thus, the formation of intersegment RNA-RNA interactions is governed by both helix-unwinding capacity of the chaperones and stability of RNA structure. We propose that this protein-mediated RNA selection mechanism may underpin the high fidelity assembly of multi-segmented RNA genomes in Reoviridae.

• MeSH
• genom virový genetika   MeSH
• konformace nukleové kyseliny MeSH
• molekulární chaperony chemie   genetika   metabolismus   MeSH
• molekulární modely MeSH
• proteiny vázající RNA chemie   genetika   metabolismus   MeSH
• ptačí orthoreovirus genetika   metabolismus   MeSH
• RNA virová chemie   genetika   metabolismus   MeSH
• sekundární struktura proteinů MeSH
• sekvence nukleotidů MeSH
• vazba proteinů MeSH
• virové nestrukturální proteiny chemie   genetika   metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• molekulární chaperony MeSH
• NS35 protein, rotavirus MeSH Prohlížeč
• proteiny vázající RNA MeSH
• RNA virová MeSH
• virové nestrukturální proteiny MeSH

RNA secondary (2D) structure visualization is an essential tool for understanding RNA function. R2DT is a software package designed to visualize RNA 2D structures in consistent, recognizable, and reproducible layouts. The latest release, R2DT 2.0, introduces multiple significant features, including the ability to display position-specific information, such as single nucleotide polymorphisms or SHAPE reactivities. It also offers a new template-free mode allowing visualization of RNAs without pre-existing templates, alongside a constrained folding mode and support for animated visualizations. Users can interactively modify R2DT diagrams, either manually or using natural language prompts, to generate new templates or create publication-quality images. Additionally, R2DT features faster performance, an expanded template library, and a growing collection of compatible tools and utilities. Already integrated into multiple biological databases, R2DT has evolved into a comprehensive platform for RNA 2D visualization, accessible at https://r2dt.bio.

• MeSH
• jednonukleotidový polymorfismus MeSH
• konformace nukleové kyseliny * MeSH
• počítačová grafika MeSH
• RNA * chemie   MeSH
• sbalování RNA MeSH
• software * MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• RNA * MeSH

Non-coding RNAs (ncRNA) are essential for all life, and their functions often depend on their secondary (2D) and tertiary structure. Despite the abundance of software for the visualisation of ncRNAs, few automatically generate consistent and recognisable 2D layouts, which makes it challenging for users to construct, compare and analyse structures. Here, we present R2DT, a method for predicting and visualising a wide range of RNA structures in standardised layouts. R2DT is based on a library of 3,647 templates representing the majority of known structured RNAs. R2DT has been applied to ncRNA sequences from the RNAcentral database and produced >13 million diagrams, creating the world's largest RNA 2D structure dataset. The software is amenable to community expansion, and is freely available at https://github.com/rnacentral/R2DT and a web server is found at https://rnacentral.org/r2dt .

• MeSH
• databáze nukleových kyselin MeSH
• konformace nukleové kyseliny MeSH
• nekódující RNA chemie   MeSH
• reprodukovatelnost výsledků MeSH
• RNA chemie   MeSH
• sekvenční analýza RNA MeSH
• software MeSH
• výpočetní biologie metody   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Intramural MeSH
• Research Support, U.S. Gov't, Non-P.H.S. MeSH
• Názvy látek
• nekódující RNA MeSH
• RNA MeSH

Viral RNA dependent polymerases (vRdPs) are present in all RNA viruses; unfortunately, their sequence similarity is too low for phylogenetic studies. Nevertheless, vRdP protein structures are remarkably conserved. In this study, we used the structural similarity of vRdPs to reconstruct their evolutionary history. The major strength of this work is in unifying sequence and structural data into a single quantitative phylogenetic analysis, using powerful a Bayesian approach. The resulting phylogram of vRdPs demonstrates that RNA-dependent DNA polymerases (RdDPs) of viruses within Retroviridae family cluster in a clearly separated group of vRdPs, while RNA-dependent RNA polymerases (RdRPs) of dsRNA and +ssRNA viruses are mixed together. This evidence supports the hypothesis that RdRPs replicating +ssRNA viruses evolved multiple times from RdRPs replicating +dsRNA viruses, and vice versa. Moreover, our phylogram may be presented as a scheme for RNA virus evolution. The results are in concordance with the actual concept of RNA virus evolution. Finally, the methods used in our work provide a new direction for studying ancient virus evolution.

• MeSH
• druhová specificita MeSH
• fylogeneze MeSH
• molekulární evoluce * MeSH
• molekulární modely MeSH
• molekulární sekvence - údaje MeSH
• RNA-dependentní RNA-polymerasa chemie   genetika   MeSH
• RNA-viry klasifikace   enzymologie   genetika   MeSH
• sekundární struktura proteinů MeSH
• sekvence aminokyselin MeSH
• sekvenční homologie aminokyselin MeSH
• terciární struktura proteinů * MeSH
• vazebná místa genetika   MeSH
• virové proteiny chemie   genetika   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• RNA-dependentní RNA-polymerasa MeSH
• virové proteiny MeSH

Methods of artificial evolution such as SELEX and in vitro selection have made it possible to isolate RNA and DNA motifs with a wide range of functions from large random sequence libraries. Once the primary sequence of a functional motif is known, the sequence space around it can be comprehensively explored using a combination of random mutagenesis and selection. However, methods to explore the sequence space of a secondary structure are not as well characterized. Here we address this question by describing a method to construct libraries in a single synthesis which are enriched for sequences with the potential to form a specific secondary structure, such as that of an aptamer, ribozyme, or deoxyribozyme. Although interactions such as base pairs cannot be encoded in a library using conventional DNA synthesizers, it is possible to modulate the probability that two positions will have the potential to pair by biasing the nucleotide composition at these positions. Here we show how to maximize this probability for each of the possible ways to encode a pair (in this study defined as A-U or U-A or C-G or G-C or G.U or U.G). We then use these optimized coding schemes to calculate the number of different variants of model stems and secondary structures expected to occur in a library for a series of structures in which the number of pairs and the extent of conservation of unpaired positions is systematically varied. Our calculations reveal a tradeoff between maximizing the probability of forming a pair and maximizing the number of possible variants of a desired secondary structure that can occur in the library. They also indicate that the optimal coding strategy for a library depends on the complexity of the motif being characterized. Because this approach provides a simple way to generate libraries enriched for sequences with the potential to form a specific secondary structure, we anticipate that it should be useful for the optimization and structural characterization of functional nucleic acid motifs.

• Klíčová slova
• DNA, RNA, SELEX, aptamer, artificial evolution, deoxyribozyme, in vitro selection, nucleic acids, ribozyme, secondary structure, synthetic biology,
• MeSH
• aptamery nukleotidové genetika   MeSH
• DNA katalytická genetika   MeSH
• genová knihovna * MeSH
• konformace nukleové kyseliny MeSH
• mutageneze MeSH
• nukleotidové motivy genetika   MeSH
• obrácené repetice genetika   MeSH
• párování bází MeSH
• pravděpodobnost MeSH
• řízená evoluce molekul metody   MeSH
• RNA katalytická genetika   MeSH
• syntetická biologie metody   MeSH
• techniky in vitro MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• aptamery nukleotidové MeSH
• DNA katalytická MeSH
• RNA katalytická MeSH

While understanding the structure of RNA molecules is vital for deciphering their functions, determining RNA structures experimentally is exceptionally hard. At the same time, extant approaches to computational RNA structure prediction have limited applicability and reliability. In this paper we provide a method to solve a simpler yet still biologically relevant problem: prediction of secondary RNA structure using structure of different molecules as a template. Our method identifies conserved and unconserved subsequences within an RNA molecule. For conserved subsequences, the template structure is directly transferred into the generated structure and combined with de-novo predicted structure for the unconserved subsequences with low evolutionary conservation. The method also determines, when the generated structure is unreliable. The method is validated using experimentally identified structures. The accuracy of the method exceeds that of classical prediction algorithms and constrained prediction methods. This is demonstrated by comparison using large number of heterogeneous RNAs. The presented method is fast and robust, and useful for various applications requiring knowledge of secondary structures of individual RNA sequences.

• Klíčová slova
• RNA, homology, prediction, secondary structure, template structure,
• Publikační typ
• časopisecké články MeSH

The crystal structure of the N-terminal domain of the RNA polymerase δ subunit (Nδ) from Bacillus subtilis solved at a resolution of 2.0Å is compared with the NMR structure determined previously. The molecule crystallizes in the space group C222(1) with a dimer in the asymmetric unit. Importantly, the X-ray structure exhibits significant differences from the lowest energy NMR structure. In addition to the overall structure differences, structurally important β sheets found in the NMR structure are not present in the crystal structure. We systematically investigated the cause of the discrepancies between the NMR and X-ray structures of Nδ, addressing the pH dependence, presence of metal ions, and crystal packing forces. We convincingly showed that the crystal packing forces, together with the presence of Ni(2+) ions, are the main reason for such a difference. In summary, the study illustrates that the two structural approaches may give unequal results, which need to be interpreted with care to obtain reliable structural information in terms of biological relevance.

• Klíčová slova
• N-terminal domain, Nuclear magnetic resonance, Protein crystallography, RNA polymerase, δ-Subunit,
• MeSH
• Bacillus subtilis enzymologie   MeSH
• DNA řízené RNA-polymerasy chemie   MeSH
• koncentrace vodíkových iontů MeSH
• konformace proteinů * MeSH
• krystalografie rentgenová metody   MeSH
• nukleární magnetická rezonance biomolekulární metody   MeSH
• sekundární struktura proteinů MeSH
• sekvence aminokyselin MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• DNA řízené RNA-polymerasy MeSH

Telomerase RNA (TR) conformation determines its function as a template for telomere synthesis and as a scaffold for the assembly of the telomerase nucleoprotein complex. Experimental analyses of TR secondary structure using DMS-Map Seq and SHAPE-Map Seq techniques show its CLOSED conformation as the consensus structure where the template region cannot perform its function. Our data show that the apparent discrepancy between experimental results and predicted TR functional conformation, mostly ignored in published studies, can be explained using data analysis based on single-molecule structure prediction from individual sequencing reads by the recently established DaVinci method. This method results in several clusters of secondary structures reflecting the structural dynamics of TR, possibly related to its multiple functional states. Interestingly, the presumed active (OPEN) conformation of TR corresponds to a minor fraction of TR under in vivo conditions. Therefore, structural polymorphism and dynamic TR transitions between CLOSED and OPEN conformations may be involved in telomerase activity regulation as a switch that functions independently of total TR transcript levels.

• Klíčová slova
• DMS-Map Seq, RNA secondary structure, SHAPE-Map Seq, single molecule analysis, telomerase RNA,
• MeSH
• konformace nukleové kyseliny MeSH
• mechy * MeSH
• RNA * chemie   genetika   MeSH
• telomerasa * chemie   genetika   MeSH
• zobrazení jednotlivé molekuly MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• RNA * MeSH
• telomerasa * MeSH
• telomerase RNA MeSH Prohlížeč