Dataset Information

Frameshifting RNA pseudoknots: structure and mechanism.

ABSTRACT: Programmed ribosomal frameshifting (PRF) is one of the multiple translational recoding processes that fundamentally alters triplet decoding of the messenger RNA by the elongating ribosome. The ability of the ribosome to change translational reading frames in the -1 direction (-1 PRF) is employed by many positive strand RNA viruses, including economically important plant viruses and many human pathogens, such as retroviruses, e.g., HIV-1, and coronaviruses, e.g., the causative agent of severe acute respiratory syndrome (SARS), in order to properly express their genomes. -1 PRF is programmed by a bipartite signal embedded in the mRNA and includes a heptanucleotide "slip site" over which the paused ribosome "backs up" by one nucleotide, and a downstream stimulatory element, either an RNA pseudoknot or a very stable RNA stem-loop. These two elements are separated by six to eight nucleotides, a distance that places the 5' edge of the downstream stimulatory element in direct contact with the mRNA entry channel of the 30S ribosomal subunit. The precise mechanism by which the downstream RNA stimulates -1 PRF by the translocating ribosome remains unclear. This review summarizes the recent structural and biophysical studies of RNA pseudoknots and places this work in the context of our evolving mechanistic understanding of translation elongation. Support for the hypothesis that the downstream stimulatory element provides a kinetic barrier to the ribosome-mediated unfolding is discussed.

SUBMITTER: Giedroc DP

PROVIDER: S-EPMC2670756 | biostudies-literature | 2009 Feb

REPOSITORIES: biostudies-literature

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Frameshifting RNA pseudoknots: structure and mechanism.

Giedroc David P DP Cornish Peter V PV

Virus research 20080725 2

Programmed ribosomal frameshifting (PRF) is one of the multiple translational recoding processes that fundamentally alters triplet decoding of the messenger RNA by the elongating ribosome. The ability of the ribosome to change translational reading frames in the -1 direction (-1 PRF) is employed by many positive strand RNA viruses, including economically important plant viruses and many human pathogens, such as retroviruses, e.g., HIV-1, and coronaviruses, e.g., the causative agent of severe acu ...[more]

PMID: 18621088

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Project description:Mutational and NMR methods were used to investigate features of sequence, structure, and dynamics that are associated with the ability of a pseudoknot to stimulate a -1 frameshift. In vitro frameshift assays were performed on retroviral gag-pro frameshift-stimulating pseudoknots and their derivatives, a pseudoknot from the gene 32 mRNA of bacteriophage T2 that is not naturally associated with frameshifting, and hybrids of these pseudoknots. Results show that the gag-pro pseudoknot from human endogenous retrovirus-K10 (HERV) stimulates a -1 frameshift with an efficiency similar to that of the closely related retrovirus MMTV. The bacteriophage T2 mRNA pseudoknot was found to be a poor stimulator of frameshifting, supporting a hypothesis that the retroviral pseudoknots have distinctive properties that make them efficient frameshift stimulators. A hybrid, designed by combining features of the bacteriophage and retroviral pseudoknots, was found to stimulate frameshifting while retaining significant structural similarity to the nonframeshifting bacteriophage pseudoknot. Mutational analyses of the retroviral and hybrid pseudoknots were used to evaluate the effects of an unpaired (wedged) adenosine at the junction of the pseudoknot stems, changing the base pairs near the junction of the two stems, and changing the identity of the loop 2 nucleotide nearest the junction of the stems. Pseudoknots both with and without the wedged adenosine can stimulate frameshifting, though the identities of the nucleotides near the stem1/stem2 junction do influence efficiency. NMR data showed that the bacteriophage and hybrid pseudoknots are similar in their local structure at the junction of the stems, indicating that pseudoknots that are similar in this structural feature can differ radically in their ability to stimulate frameshifting. NMR methods were used to compare the internal motions of the bacteriophage T2 pseudoknot and representative frameshifting pseudoknots. The stems of the investigated pseudoknots are similarly well ordered on the time scales to which nitrogen-15 relaxation data are sensitive; however, solvent exchange rates for protons at the junction of the two stems of the nonframeshifting bacteriophage pseudoknot are significantly slower than the analogous protons in the representative frameshifting pseudoknots.

Project description:MotivationThe function of an RNA molecule is not only linked to its native structure, which is usually taken to be the ground state of its folding landscape, but also in many cases crucially depends on the details of the folding pathways such as stable folding intermediates or the timing of the folding process itself. To model and understand these processes, it is necessary to go beyond ground state structures. The study of rugged RNA folding landscapes holds the key to answer these questions. Efficient coarse-graining methods are required to reduce the intractably vast energy landscapes into condensed representations such as barrier trees or basin hopping graphs : BHG) that convey an approximate but comprehensive picture of the folding kinetics. So far, exact and heuristic coarse-graining methods have been mostly restricted to the pseudoknot-free secondary structures. Pseudoknots, which are common motifs and have been repeatedly hypothesized to play an important role in guiding folding trajectories, were usually excluded.ResultsWe generalize the BHG framework to include pseudoknotted RNA structures and systematically study the differences in predicted folding behavior depending on whether pseudoknotted structures are allowed to occur as folding intermediates or not. We observe that RNAs with pseudoknotted ground state structures tend to have more pseudoknotted folding intermediates than RNAs with pseudoknot-free ground state structures. The occurrence and influence of pseudoknotted intermediates on the folding pathway, however, appear to depend very strongly on the individual RNAs so that no general rule can be inferred.Availability and implementationThe algorithms described here are implemented in C++ as standalone programs. Its source code and Supplemental material can be freely downloaded from http://www.tbi.univie.ac.at/bhg.html.Contactqin@bioinf.uni-leipzig.deSupplementary informationSupplementary data are available at Bioinformatics online.

Dataset Information

Frameshifting RNA pseudoknots: structure and mechanism.

Publications

Frameshifting RNA pseudoknots: structure and mechanism.

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