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A phage RNA-binding protein binds to a non-cognate structured RNA and stabilizes its core structure.

ABSTRACT: Recent studies suggest that some RNA-binding proteins facilitate the folding of non-cognate RNAs. Here, we report that bacteriophage MS2 coat protein (MS2 CP) bound and promoted the catalytic activity of Candida group I ribozyme. Cloning of the MS2-bound RNA segments showed that this protein primarily interacts with the P5ab-P5 structure. Ultraviolet cross-linking and the T1 footprinting assay further showed that MS2 binding stabilized tertiary interactions, including the conserved L9-P5 interaction, and led to a more compact core structure. This mechanism is similar to that of the yeast mitochondrial tyrosyl-tRNA synthetase on other group I introns, suggesting that different RNA-binding proteins may use common mechanisms to support RNA structures.

SUBMITTER: Xie MH

PROVIDER: S-EPMC7117394 | biostudies-literature | 2009 Jan

REPOSITORIES: biostudies-literature

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A phage RNA-binding protein binds to a non-cognate structured RNA and stabilizes its core structure.

Xie Mao-Hua MH Wu Qi-Jia QJ Jiang Yan-Fei YF Bao Penghui P Zhang Yi Y

Biochemical and biophysical research communications 20081108 2

Recent studies suggest that some RNA-binding proteins facilitate the folding of non-cognate RNAs. Here, we report that bacteriophage MS2 coat protein (MS2 CP) bound and promoted the catalytic activity of Candida group I ribozyme. Cloning of the MS2-bound RNA segments showed that this protein primarily interacts with the P5ab-P5 structure. Ultraviolet cross-linking and the T1 footprinting assay further showed that MS2 binding stabilized tertiary interactions, including the conserved L9-P5 interac ...[more]

PMID: 19000653

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Project description:BackgroundNSC260594, a quinolinium derivative from the NCI diversity set II compound library, was previously identified in a target-based assay as an inhibitor of the interaction between the HIV-1 (ψ) stem-loop 3 (SL3) RNA and Gag. This compound was shown to exhibit potent antiviral activity. Here, the effects of this compound on individual stages of the viral lifecycle were examined by qRT-PCR, ELISA and Western blot, to see if its actions were specific to the viral packaging stage. The structural effects of NSC260594 binding to the HIV-1 gRNA were also examined by SHAPE and dimerization assays.ResultsTreatment of cells with NSC260594 did not reduce the number of integration events of incoming virus, and treatment of virus producing cells did not affect the level of intracellular Gag protein or viral particle release as determined by immunoblot. However, NSC260594 reduced the incorporation of gRNA into virions by up to 82%, without affecting levels of gRNA inside the cell. This reduction in packaging correlated closely with the reduction in infectivity of the released viral particles. To establish the structural effects of NSC260594 on the HIV-1 gRNA, we performed SHAPE analyses to pinpoint RNA structural changes. NSC260594 had a stabilizing effect on the wild type RNA that was not confined to SL3, but that was propagated across the structure. A packaging mutant lacking SL3 did not show this effect.ConclusionsNSC260594 acts as a specific inhibitor of HIV-1 RNA packaging. No other viral functions are affected. Its action involves preventing the interaction of Gag with SL3 by stabilizing this small RNA stem-loop which then leads to stabilization of the global packaging signal region (psi or ψ). This confirms data, previously only shown in analyses of isolated SL3 oligonucleotides, that SL3 is structurally labile in the presence of Gag and that this is critical for the complete psi region to be able to adopt different conformations. Since replication is otherwise unaffected by NSC260594 the flexibility of SL3 appears to be a unique requirement for genome encapsidation and identifies this process as a highly specific drug target. This study is proof of principle that development of a new class of antiretroviral drugs that specifically target viral packaging by binding to the viral genomic RNA is achievable.

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A phage RNA-binding protein binds to a non-cognate structured RNA and stabilizes its core structure.

Publications

A phage RNA-binding protein binds to a non-cognate structured RNA and stabilizes its core structure.

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