PubMed Health⌕ Search

SEARCH · PubMed Health

Results for “RNA chaperone”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 37 records · Page 2Linked to original sources
PubMed2004

RNA chaperone activity of large ribosomal subunit proteins from Escherichia coli.

The ribosome is a highly dynamic ribonucleoprotein machine. During assembly and during translation the ribosomal RNAs must routinely be prevented from falling into kinetic folding traps. Stable occupation of these trapped states may be prevented by proteins with RNA chaperone activity. Here, ribosomal proteins from the large (50S) ribosome subunit of Escherichia coli were tested for RNA chaperone activity in an in vitro trans splicing assay. Nearly a third of the 34 large ribosomal subunit proteins displayed RNA chaperone activity. We discuss a possible role of this function during ribosome assembly and during translation.

Escherichia coli↗
PubMed2000

Escherichia coli CspA-family RNA chaperones are transcription antiterminators.

CspA, the major cold-shock protein of Escherichia coli, is an RNA chaperone, which is thought to facilitate translation at low temperature by destabilizing mRNA structures. Here we demonstrate that CspA, as well as homologous RNA chaperones CspE and CspC, are transcription antiterminators. In vitro, the addition of physiological concentrations of recombinant CspA, CspE, or CspC decreased transcription termination at several intrinsic terminators and also decreased transcription pausing. In vivo, overexpression of cloned CspC and CspE at 37 degrees C was sufficient to induce transcription of the metY-rpsO operon genes nusA, infB, rbfA, and pnp located downstream of multiple transcription terminators. Similar induction of downstream metY-rpsO operon genes was observed at cold shock, a condition to which the cell responds by massive overproduction of CspA. The products of nusA, infB, rbfA, and pnp-NusA, IF2, RbfA, and PNP-are known to be induced at cold shock. We propose that the cold-shock induction of nusA, infB, rbfA, and pnp occurs through transcription antitermination, which is mediated by CspA and other cold shock-induced Csp proteins.

Adaptation, Biological↗
PubMed2004

The splicing of yeast mitochondrial group I and group II introns requires a DEAD-box protein with RNA chaperone function.

Group I and II introns self-splice in vitro, but require proteins for efficient splicing in vivo, to stabilize the catalytically active RNA structure. Recent studies showed that the splicing of some Neurospora crassa mitochondrial group I introns additionally requires a DEAD-box protein, CYT-19, which acts as an RNA chaperone to resolve nonnative structures formed during RNA folding. Here we show that, in Saccharomyces cerevisiae mitochondria, a related DEAD-box protein, Mss116p, is required for the efficient splicing of all group I and II introns, some RNA end-processing reactions, and translation of a subset of mRNAs, and that all these defects can be partially or completely suppressed by the expression of CYT-19. Results for the aI2 group II intron indicate that Mss116p is needed after binding the intron-encoded maturase, likely for the disruption of stable but inactive RNA structures. Our results suggest that both group I and II introns are prone to kinetic traps in RNA folding in vivo and that the splicing of both types of introns may require DEAD-box proteins that function as RNA chaperones.

Amino Acid Motifs↗
PubMed2002

Folding of the td pre-RNA with the help of the RNA chaperone StpA.

The td group I intron is inserted in the reading frame of the thymidylate synthase gene, which is mainly devoid of structural elements. In vivo, translation of the pre-mRNA is required for efficient folding of the intron into its splicing-competent structure. The ribosome probably resolves exon-intron interactions that interfere with splicing. Uncoupling splicing from translation, by introducing a non-sense codon into the upstream exon, reduces the splicing efficiency of the mutant pre-mRNA. Alternatively to the ribosome, co-expression of genes that encode proteins with RNA chaperone activity promote folding of the td pre-mRNA in vivo. These proteins also efficiently accelerate folding of the td pre-mRNA in vitro. In order to understand the mechanism of action of RNA chaperones, we probed the impact of the RNA chaperone StpA on the structure of the td intron in vivo and compared it with that of the well characterized Cyt-18 protein, which is a group-I-intron-specific splicing factor. We found that the two proteins have opposite effects on the structure of the td intron. While StpA loosens the three-dimensional structure, Cyt-18 tightens it up. Furthermore, mutations that destabilize the intron structure render the mutants sensitive to StpA, whereas splicing of these mutants is rescued by Cyt-18. Our results provide direct evidence for protein-induced conformational changes within a catalytic RNA in vivo. Whereas StpA resolves tertiary contacts enabling the RNA to refold, Cyt-18 contributes to the stabilization of the native three-dimensional structure.

Base Sequence↗
PubMed2000

Copy-choice recombination by reverse transcriptases: reshuffling of genetic markers mediated by RNA chaperones.

Copy-choice recombination efficiently reshuffles genetic markers in retroviruses. In vivo, the folding of the genomic RNA is controlled by the nucleocapsid protein (NC). We show that binding of NC onto the acceptor RNA molecule is sufficient to enhance recombination, providing evidence for a mechanism where the structure of the acceptor template determines the template switch. NC as well as another RNA chaperone (StpA) converts recombination into a widespread process no longer restricted to rare hot spots, an effect maximized when both the NC and the reverse transcriptase come from HIV-1. These data suggest that RNA chaperones confer a higher genetic flexibility to retroviruses.

Genetic Markers↗
PubMed2006

The RNA chaperone Hfq is essential for the virulence of Salmonella typhimurium.

The RNA chaperone, Hfq, plays a diverse role in bacterial physiology beyond its original role as a host factor required for replication of Qbeta RNA bacteriophage. In this study, we show that Hfq is involved in the expression and secretion of virulence factors in the facultative intracellular pathogen, Salmonella typhimurium. A Salmonella hfq deletion strain is highly attenuated in mice after both oral and intraperitoneal infection, and shows a severe defect in invasion of epithelial cells and a growth defect in both epithelial cells and macrophages in vitro. Surprisingly, we find that these phenotypes are largely independent of the previously reported requirement of Hfq for expression of the stationary phase sigma factor, RpoS. Our results implicate Hfq as a key regulator of multiple aspects of virulence including regulation of motility and outer membrane protein (OmpD) expression in addition to invasion and intracellular growth. These pleiotropic effects are suggested to involve a network of regulatory small non-coding RNAs, placing Hfq at the centre of post-transcriptional regulation of virulence gene expression in Salmonella. In addition, the hfq mutation appears to cause a chronic activation of the RpoE-mediated envelope stress response which is likely due to a misregulation of membrane protein expression.

Bacterial Outer Membrane Proteins↗
PubMed2003

RNA chaperone activity of the Sm-like Hfq protein.

The Escherichia coli Sm-like host factor I (Hfq) protein is thought to function in post-transcriptional regulation by modulating the function of small regulatory RNAs. Hfq also interferes with ribosome binding on E. coli ompA messenger RNA, indicating that Hfq also interacts with mRNAs. In this study, we have used stimulation of group I intron splicing in vivo and a modified in vitro toeprinting assay to determine whether Hfq acts as an RNA chaperone. Hfq was able to rescue an RNA 'folding trap' in a splicing defective T4 bacteriophage td gene in vivo. Enzymatic analysis showed that Hfq affects the accessibility of the ompA start codon, as well as other bases within the ribosome-binding site, explaining its negative effect on ribosome binding. We also show that the Hfq-induced structural changes in ompA mRNA are maintained after proteolytic digestion of the protein, which classifies Hfq as an RNA chaperone.

Base Sequence↗
PubMed2002

RNA chaperone StpA loosens interactions of the tertiary structure in the td group I intron in vivo.

Efficient splicing of the td group I intron in vivo is dependent on the ribosome. In the absence of translation, the pre-mRNA is trapped in nonnative-splicing-incompetent conformations. Alternatively, folding of the pre-mRNA can be promoted by the RNA chaperone StpA or by the group I intron-specific splicing factor Cyt-18. To understand the mechanism of action of RNA chaperones, we probed the impact of StpA on the structure of the td intron in vivo. Our data suggest that StpA loosens tertiary interactions. The most prominent structural change was the opening of the base triples, which are involved in the correct orientation of the two major intron core domains. In line with the destabilizing activity of StpA, splicing of mutant introns with a reduced structural stability is sensitive to StpA. In contrast, Cyt-18 strengthens tertiary contacts, thereby rescuing splicing of structurally compromised td mutants in vivo. Our data provide direct evidence for protein-induced conformational changes within catalytic RNA in vivo. Whereas StpA resolves tertiary contacts enabling the RNA to refold, Cyt-18 contributes to the overall compactness of the td intron in vivo.

Amino Acyl-tRNA Synthetases↗
PubMed2006

Characterization of the RNA chaperone activity of hantavirus nucleocapsid protein.

Hantaviruses are tripartite negative-sense RNA viruses and members of the Bunyaviridae family. The nucleocapsid (N) protein, encoded by the smallest of the three genome segments (S), has nonspecific RNA chaperone activity. This activity results in transient dissociation of misfolded RNA structures, may be required for facilitating correct higher-order RNA structure, and may function in viral genome replication. We carried out a series of experiments to further characterize the ability of N to dissociate RNA duplexes. As might be expected, N dissociated RNA duplexes but not DNA duplexes or RNA-DNA heteroduplexes. The RNA-destabilizing activity of N is ATP independent, has a pH optimum of 7.5, and has an Mg(2+) concentration optimum of 1 to 2 mM. N protein is unable to unwind the RNA duplexes that are completely double stranded. However, in the presence of an adjoining single-stranded region, helix unwinding takes place in the 3'-to-5' direction through an unknown mechanism. The N protein trimer specifically recognizes and unwinds the terminal panhandle structure in the viral RNA and remains associated with unwound 5' terminus. We suggest that hantaviral nucleocapsid protein has an active role in hantaviral replication by working cooperatively with viral RNA polymerase. After specific recognition of the panhandle structure by N protein, the unwound 5' terminus likely remains transiently bound to N protein, creating an opportunity for the viral polymerase to initiate transcription at the accessible 3' terminus.

Adenosine Triphosphate↗
PubMed2002

A DEAD-box protein functions as an ATP-dependent RNA chaperone in group I intron splicing.

The Neurospora crassa CYT-18 protein, the mitochondrial tyrosyl-tRNA synthetase, functions in splicing group I introns by inducing formation of the catalytically active RNA structure. Here, we identified a DEAD-box protein (CYT-19) that functions in concert with CYT-18 to promote group I intron splicing in vivo and vitro. CYT-19 does not bind specifically to group I intron RNAs and instead functions as an ATP-dependent RNA chaperone to destabilize nonnative RNA structures that constitute kinetic traps in the CYT-18-assisted RNA-folding pathway. Our results demonstrate that a DExH/D-box protein has a specific, physiologically relevant chaperone function in the folding of a natural RNA substrate.

Adenosine Triphosphatases↗
PubMed2006

Nonspecific binding to structured RNA and preferential unwinding of an exposed helix by the CYT-19 protein, a DEAD-box RNA chaperone.

We explore the interactions of CYT-19, a DExD/H-box protein that functions in folding of group I RNAs, with a well characterized misfolded species of the Tetrahymena ribozyme. Consistent with its function, CYT-19 accelerates refolding of the misfolded RNA to its native state. Unexpectedly, CYT-19 performs another reaction much more efficiently; it unwinds the 6-bp P1 duplex formed between the ribozyme and its oligonucleotide substrate. Furthermore, CYT-19 performs this reaction 50-fold more efficiently than it unwinds the same duplex free in solution, suggesting that it forms additional interactions with the ribozyme, most likely using a distinct RNA binding site from the one responsible for unwinding. This site can apparently bind double-stranded RNA, as attachment of a simple duplex adjacent to P1 recapitulates much of the activation provided by the ribozyme. Unwinding the native P1 duplex does not accelerate refolding of the misfolded ribozyme, implying that CYT-19 can disrupt multiple contacts on the RNA, consistent with its function in folding of multiple RNAs. Further experiments showed that the P1 duplex unwinding activity is virtually the same whether the ribozyme is misfolded or native but is abrogated by formation of tertiary contacts between the P1 duplex and the body of the ribozyme. Together these results suggest a mechanism for CYT-19 and other general DExD/H-box RNA chaperones in which the proteins bind to structured RNAs and efficiently unwind loosely associated duplexes, which biases the proteins to disrupt nonnative base pairs and gives the liberated strands an opportunity to refold.

Animals↗
PubMed2005

Evidence for an RNA chaperone function of polypyrimidine tract-binding protein in picornavirus translation.

The cellular polypyrimidine tract-binding protein (PTB) is recruited by the genomic RNAs of picornaviruses to stimulate translation initiation at their internal ribosome entry site (IRES) elements. We investigated the contribution of the individual RNA recognition motif (RRM) domains of PTB to its interaction with the IRES of foot-and-mouth disease virus (FMDV). Using a native gel system, we found that PTB is a monomer, confirming recent reports that challenged the previous view that PTB is a dimer. Mapping the spatial orientation of PTB relative to the bound IRES RNA, we found that the two C-terminal RRM domains III and IV of PTB bind in an oriented way to the IRES. Domain III contacts the IRES stem-loop 2, while domain IV contacts the separate IRES 3' region. PTB domain I appears not to be involved directly in RNA binding, but domain II stabilizes the RNA binding conferred by domains III and IV. A PTB protein containing only these two C-terminal PTB domains is sufficient to enhance the entry of initiation factor eIF4G to the IRES and stimulate IRES activity, and the long-lived PTB-IRES interaction stabilized by domain II is not a prerequisite for this function. Thus, PTB most likely acts as an RNA chaperone to stabilize IRES structure and, in that way, augment IRES activity.

Aphthovirus↗
PubMed1994

Escherichia coli proteins, including ribosomal protein S12, facilitate in vitro splicing of phage T4 introns by acting as RNA chaperones.

To address the effect of host proteins on the self-splicing properties of the group I introns of bacteriophage T4, we have purified an activity from Escherichia coli extracts that facilitates both trans- and cis-splicing of the T4 introns in vitro. The activity is attributable to a number of proteins, several of which are ribosomal proteins. Although these proteins have variable abilities to stimulate splicing, ribosomal protein S12 is the most effective. The activity mitigates the negative effects on splicing of the large internal open reading frames (ORFs) common to the T4 introns. In contrast to proteins shown previously to facilitate group I splicing, S12 does not bind strongly or specifically to the intron. Rather, S12 binds RNA with broad specificity and can also facilitate the action of a hammerhead ribozyme. Addition of S12 to unreactive trans-splicing precursors promoted splicing, suggesting that S12 can resolve misfolded RNAs. Furthermore, incubation with S12 followed by its proteolytic removal prior to the initiation of the splicing reaction still resulted in splicing enhancement. These results suggest that this protein facilitates splicing by acting as an RNA chaperone, promoting the assembly of the catalytically active tertiary structure of ribozymes.

Amino Acid Sequence↗
PubMed1997

CspA, the major cold-shock protein of Escherichia coli, is an RNA chaperone.

CspA, the major cold-shock protein of Escherichia coli, is dramatically induced during the cold-shock response. The amino acid sequence of CspA shows 43% identity to the "cold-shock domain" of the eukaryotic Y-box protein family, which interacts with RNA and DNA to regulate their functions. Here, we demonstrate that CspA binds to RNA as a chaperone. First, CspA cooperatively binds to heat-denatured single-stranded RNA if it is larger than 74 bases, causing a supershift in gel electrophoresis. A minimal concentration of CspA at 2.7 x 10(-5) M is absolutely required for this cooperative binding, which is sufficiently lower than the estimated cellular concentration of CspA (10(-4) M) in cold-shocked cells. No specific RNA sequences for CspA binding were identified, indicating that it has a broad sequence specificity for its binding. When the 142-base 5'-untranslated region of the cspA mRNA was used as a substrate for ribonucleases A and T1, the addition of CspA significantly stimulated RNA hydrolysis by preventing the formation of RNase-resistant bands due to stable secondary structures in the 5'-untranslated region. These results indicate that binding of CspA to RNA destabilizes RNA secondary structures to make them susceptible to ribonucleases. We propose that CspA functions as an RNA chaperone to prevent the formation of secondary structures in RNA molecules at low temperature. Such a function may be crucial for efficient translation of mRNAs at low temperatures and may also have an effect on transcription.

Bacterial Proteins↗
PubMed2001

A collapsed state functions to self-chaperone RNA folding into a native ribonucleoprotein complex.

Most large RNAs achieve their active, native structures only as complexes with one or more cofactor proteins. By varying the Mg(2+) concentration, the catalytic core of the bI5 group I intron RNA can be manipulated into one of three states, expanded, collapsed or native, or into balanced equilibria between these states. Under near-physiological conditions, the bI5 RNA folds rapidly to a collapsed but non-native state. Hydroxyl radical footprinting demonstrates that assembly with the CBP2 protein cofactor chases the RNA from the collapsed state to the native state. In contrast, CBP2 also binds to the RNA in the expanded state to form many non-native interactions. This structural picture is reinforced by functional splicing experiments showing that RNA in an expanded state forms a non-productive, kinetically trapped complex with CBP2. Thus, rapid folding to the collapsed state functions to self-chaperone bI5 RNA folding by preventing premature interaction with its protein cofactor. This productive, self-chaperoning role for RNA collapsed states may be especially important to avert misassembly of large multi-component RNA-protein machines in the cell.

Animals↗
PubMed2004

Artemin is an RNA-binding protein with high thermal stability and potential RNA chaperone activity.

Encysted embryos of the crustacean, Artemia franciscana, are among the most stress-resistant of all multicellular eukaryotes, due in part to massive amounts of p26, a small heat shock protein, that acts as a molecular chaperone. These embryos contain equally large amounts of another protein called artemin, of previously unknown function, that we report on here. Its thermal stability allows large-scale purification in about a day, using ammonium sulfate fractionation and incubation at 70 degrees C for 7 min, followed by gel filtration. The latter yields an artemin-RNA complex from which the pure protein, apo-artemin, was obtained by anion-exchange chromatography. We evaluated the possibility that artemin acts as a molecular chaperone for proteins, but obtained no evidence for that in vitro. The association of RNA with apo-artemin occurs at high temperatures and, although it is not yet clear whether artemin has a specific role as an RNA chaperone, it does bind non-polyadenylated RNAs which are then translated in vitro. Artemin-RNA is thermostable, some molecules resisting destruction after 30 min at 90 degrees C. The first order rate constant for denaturation and aggregation of artemin-RNA at 85 degrees C is 8.5 x 10(-3)min(-1), which compares well with other thermostable proteins of similar size ( approximately 500 kDa) such as the ferritins with which artemin has amino acid sequence similarity. The amount of artemin extracted from embryos that had been stored dry, under laboratory conditions, since 1951 is comparable to the amount in contemporary embryos, indicating its stability in situ, and supporting the in vitro heating studies.

Amino Acid Sequence↗
PubMed2004

Extracellular and circulating redox- and metalloregulated eRNA and eRNP: copper ion-structured RNA cytokines (angiotropin ribokines) and bioaptamer targets imparting RNA chaperone and novel biofunctions to S100-EF-hand and disease-associated proteins.

Bioassays for cellular differentiation and tissue morphogenesis were used to design methods for isolation of bioactive redox- and metalloregulated nucleic acids and copper ion complexes with proteins from extracellular, circulating, wound, and supernatant fluids of cultured cells. In extracellular biospheres, diversities of nucleic acids were found to be secreted by cells upon activation. They may reflect nucleic acid biolibraries with molecular imprints of cellular history. After removal of protein components, eRNA prototypes exuded by activated cells were sequenced. They are small, endogenous, highly modified and edited, redox- and metalloregulated 5'-end phosphorylated extracellular eRNA (approximately 2-200 bases) with cellular, enzymic, and bioaptamer functions. Fenton-type OH* radical redox reactions may form modified nucleotides in RNA as wobbles eRNA per se, or as copper ion-complex with protein (e.g., S100A12-EF-hand protein, angiotropin-related protein, calgranulin-C, hippocampal neurite differentiation factor) are shown to be bioactive in vivo and in vitro as cytokines (ribokines) and as nonmitogenic angiomorphogens for endothelial cell differentiation in the formation of organoid supracellular capillary structures. As bioaptamers, copper ion-structured eRNA imparts novel biofunctions to proteins that they do not have on their own. The origin of extracellular RNA and intermediate precursors (up to 500 bases) was traced to intracellular parent nucleic acids. Intermediate precursors with and without partial homology were found. This suggests that bioaptamers are not directly retranslatable gene products. Metalloregulated eRNA bioaptamer function was investigated by domains (e.g. 5'...CUG...3' hairpin loop) for folding, bioactivity, and binding of protein with copper, calcium, and alkali metal ion affinity. Vice versa, metalloregulated nucleic acid-binding domains (K3H, R3H) in proteins were identified. Interaction of protein and eRNA docking potentials were visualized by 3D-rapid prototyping of accurate molecular image models based on crystallographic or NMR data. For S100A12-homologous proteins, receptor- and metalloregulated RNA chaperone-shaped protein assemblies were investigated. They suggest insight into signaling cascades as to how eRNA transmits its cytokine (ribokine) bioinformation from the extracellular RNA biosphere into cells. Proteomics of the extracellular RNA biosphere demonstrate the presence of nucleic acid-binding domain homologies in defense-, aging-, and disease-associated neuronal and other proteins as targets for RNA orphans. By structural relationships found to transmissible processes, proteinaceous transfer ("infectivity") and feedback of bioinformation beyond the central dogma of molecular biology are considered in terms of metalloregulated RNA bioaptamer function, nucleic acid-binding domains, and protein conformation.

Amino Acid Sequence↗