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At least 361 records · Page 20Linked to original sources
PubMed1999

Mapping sequences active in homologous RNA recombination in brome mosaic virus: prediction of recombination hot spots.

The mechanism of homologous recombination has been studied previously in brome mosaic virus (BMV), a tricomponent, positive-stranded RNA virus of plants, by using artificial sequences (reviewed by J. J. BujarskiP. D. Nagy (1996). Semin. Virol. 7, 363-372). Here we extend these studies over BMV-derived sequences to obtain clues on prediction of homologous recombination hot spots. First, mismatch mutations, which reduced the AU content, were introduced into the common 60-nt recombination hot-spot sequence, either in the RNA2 or in both RNA2RNA3 components. This decreased the frequency of targeted homologous RNA2/RNA3 recombinationchanged the distribution of junction sites. Second, several short BMV RNA1- or RNA2-derived sequences were introduced into the RNA3 component, homologous recombination activity of these sequences was compared with that observed for previously characterized artificial sequences. Third, sequences at homologous recombinant junctions were compared among a large number of targetednontargeted recombinants. All these studies revealed several factors important for homologous recombination including the length of sequence identity, the extent of sequence identity, the AU content of the common sequences, the relative position of the AU-rich segment vs a GC-rich segment,the presence of GC-rich sequences. Based on this novel model, we suggest that recombination hot spots can be predicted by means of RNA sequence analysis. In addition, we show that recombination can occur between positivenegative strands of BMV RNAs. This provides further clues toward the mechanism of recombination processes in BMV.

Base Sequence↗
PubMed1994

Characterization of the "hepatic" asialoglycoprotein receptor in rat late-stage spermatids and epididymal sperm.

Northern blot analysis of rat testicular (Te) poly(A)+RNA reveals that a transcript homologous to the major form of the asialoglycoprotein receptor (ASGP-R), designated RHL-1, is expressed as early as one week postnatally and that steady-state levels are approx. 8-times higher in the Te of an 8-week-old rat (sexually mature) as compared to an 84-week-old rat (aged). Partial cDNAs encoding RHL-1 and the minor form of the ASGP-R, designated RHL-2/3, have been cloned from two rat Te/epididymal (Ep) cDNA libraries and rat Te poly(A)+RNA. Sequence analysis of the Te/Ep RHL-1 cDNA and the Te/Ep RHL-2/3 cDNA indicates that these cDNAs are identical to the forms expressed in rat liver. Western blot analysis demonstrates the presence of a 49-kDa Te/Ep RHL-1-related protein band and a 54-kDa Te/Ep RHL-2/3-related protein band in both rat Te membrane fractions (MF) and rat Ep sperm MF. The RHL-1-related protein has been localized to late-stage Te spermatids at the time of release from the seminiferous tubules and to Ep sperm in the region of the sperm tail, referred to as the middle piece. Taken collectively, these data indicate that the authentic RHL-1 and RHL-2/3 genes of the ASGP-R are expressed in late-stage spermatids; however, the Te/Ep RHL-1-related protein differs in size from the hepatic RHL-1 polypeptide, possibly indicating a specific function of the RHL-1-related protein in spermatogenesis.

Age Factors↗
PubMed1980

The purification and properties of chicken liver RNase: An enzyme which is useful in distinguishing between cytidylic and uridylic acid residues.

A heat-stable endoribonuclease isolated from chicken liver has been purified to homogeneity as evidenced by the presence of a single protein band upon polyacrylamide gel electrophoresis. The enzyme can, in limit digests of 5 S rRNA and 5.8 S rRNA, dinstinguish between cytidylic and uridylic acids bonds at a ratio of 61:1 and, therefore, may be useful in RNA sequence analysis. The means by which the enzyme hydrolyzes substrate is unusual in that kinetic data do not support a simple formation and breakdown of an enzyme . substrate complex. Rather, the existence of a second complex, consisting of 2 mol of substrate and one of enzyme, derived from the initial enzyme . substrate complex, is postulated. In common with the other endonucleases, enzyme activity is inhibited by free poly(A) or tracts of the polypurine present at the 3'-terminus of RNA. Reversal of inhibition and restoration of activity may be achieved by the addition of low concentrations of spermidine to reaction mixtures.

Animals↗
PubMed1989

Specific oligodeoxynucleotide probes obtained through RNA sequencing.

By combining several established techniques we developed a method to test the specificity of mixed oligodeoxynucleotide hybridization probes and to provide the information for the design of long nondegenerate, and therefore more specific probes. Mixed oligodeoxynucleotide probes derived from known peptide sequences are first used to initiate primer extension reactions with poly(A)+RNA as template in the presence of three dNTPs and one ddNTP to generate cDNA transcripts of defined lengths. Comparing the lengths of the cDNA transcripts with the possible nucleic acid sequence coding for the known oligopeptide indicates whether the oligodeoxynucleotide mix hybridizes predominantly to the RNA of interest. In a second step, the oligodeoxynucleotide mix with the highest specificity is used for indirect RNA sequence analysis. This confirms the specificity of the probe and provides information to design a long, highly specific oligodeoxynucleotide probe for the gene of interest. This simple two-step-procedure helps to circumvent the time-consuming procedures of subcloning and sequencing of cross-hybridizing fragments.

Base Sequence↗
PubMed2026

Peripheral pain threshold, glycaemic status, and LAMP3 genetic variation: A community-based analysis.

Diabetic polyneuropathy is a common complication of diabetes, yet substantial inter-individual variation in peripheral pain perception suggests underlying genetic influences. This population-based study investigated clinical, metabolic, and genetic determinants of pain threshold using intraepidermal electrical stimulation in 906 participants from the Iwaki Health Promotion Project 2017. Genome-wide association analysis identified 12 loci showing suggestive associations, among which a missense variant in LAMP3 (rs482912) was prioritized as a biologically plausible candidate. Phenotype-stratified analyses showed that individuals carrying the CT or CC genotypes had lower PINT indices than those with the TT genotype, indicating reduced pain thresholds. Notably, the CC genotype retained an association with lower pain threshold using intraepidermal electrical stimulation under conditions of metabolic stress, including impaired glucose tolerance, elevated HbA1c, and obesity, whereas this association was attenuated in the presence of hypertension. Single-cell RNA sequencing analysis of human skin revealed that LAMP3-positive mature dendritic cells, enriched in immunoregulatory molecules, exhibited transcriptional enrichment of inflammatory, antigen-presenting, and nociception-related pathways, including NF-κB, JAK-STAT, cytokine signaling, and neuroimmune sensitization cascades. Autopsy-based skin analysis further demonstrated genotype-associated differences in dermal LAMP3-positive cell infiltration and CD8-positive T-cell abundance, while CD4-positive T-cell abundance and intraepidermal nerve fiber density remained unchanged across genotypes. Taken together, these findings suggest a potential association between LAMP3 variation and individual differences in peripheral pain threshold and provide biological context supporting a role for neuroimmune interactions in early sensory modulation under metabolic stress. Given the suggestive genetic evidence and indirect mechanistic data, these observations should be interpreted as exploratory and hypothesis-generating.

Humans↗
PubMed1984

Transfer RNA splicing in Saccharomyces cerevisiae: defining the substrates.

The primary sequences of all the tRNA precursors which contain intervening sequences and which accumulate in the Saccharomyces cerevisiae rnal mutant are presented. A combination of DNA and RNA sequence analysis has led to elucidation of the primary sequence of four hitherto uncharacterized precursors. The location of the intervening sequence has in all cases been unambiguously determined by analysis of the intermediates in the splicing reaction. Secondary structures based upon the tRNA cloverleaf are shown for all the tRNA precursors and discussed with respect to common recognition by the yeast splicing endonuclease.

Base Sequence↗
PubMed2006

RNAKinetics: a web server that models secondary structure kinetics of an elongating RNA.

The RNAKinetics server (http://www.ig-msk.ru/RNA/kinetics) is a web interface for the newly developed RNAKinetics software. The software models the dynamics of RNA secondary structure by the means of kinetic analysis of folding transitions of a growing RNA molecule. The result of the modeling is a kinetic ensemble, i.e. a collection of RNA structures that are endowed with probabilities, which depend on time. This approach gives comprehensive probabilistic description of RNA folding pathways, revealing important kinetic details that are not captured by the traditional structure prediction methods. The access to the RNAKinetics server is free.

Base Sequence↗
PubMed2025

The impact of m6A methyltransferase METTL3 on airway remodeling in bronchial asthma.

BACKGROUND: Methyltransferase-like 3 (METTL3) is known to play a role in asthma airway remodeling and cell proliferation. Adenylate kinase 4 (AK4) regulates the proliferation of pulmonary artery smooth muscle cells and exerts its effects through the protein kinase B (AKT) pathway. However, the role of METTL3 and AK4-AKT in bronchial smooth muscle cells remains unclear. METHODS: Systemic METTL3 knockout mice and a mouse model of asthma were established. Airway remodeling was assessed using pulmonary function tests, histopathological staining, and Western blot analysis. RNA sequencing (RNA-seq) was performed to detect changes in gene expression following METTL3 knockdown. The 5-ethynyl-2'-deoxyuridine (EdU) assay was used to evaluate cell proliferation. Finally, the expression levels of relevant proteins were validated by Western blotting. RESULTS: Compared with the control model group, the METTL3 knockout group showed significantly reduced inflammatory cell infiltration, decreased collagen fiber deposition, and attenuated airway smooth muscle hyperplasia. RNA-seq revealed that the expression of numerous proliferation-related genes, including AK4, was upregulated following METTL3 knockdown. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis indicated that these genes were primarily enriched in the phosphatidylinositol 3-kinase (PI3K)/AKT signaling pathway. The EdU assay demonstrated that METTL3 knockdown inhibited cell proliferation. Western blot validation showed increased AK4 expression in lung tissues of the METTL3 knockout group compared to the control group, while phosphorylated AKT (p-AKT) levels were reduced. CONCLUSIONS: METTL3 knockout inhibits airway smooth muscle hyperplasia and alleviates airway remodeling in asthma. This effect may be mediated through the regulation of AK4 and the AKT signaling pathway.

Airway Remodeling↗
PubMedDate not supplied

Statistical analysis of RNA backbone.

Local conformation is an important determinant of RNA catalysis and binding. The analysis of RNA conformation is particularly difficult due to the large number of degrees of freedom (torsion angles) per residue. Proteins, by comparison, have many fewer degrees of freedom per residue. In this work, we use and extend classical tools from statistics and signal processing to search for clusters in RNA conformational space. Results are reported both for scalar analysis, where each torsion angle is separately studied, and for vectorial analysis, where several angles are simultaneously clustered. Adapting techniques from vector quantization and clustering to the RNA structure, we find torsion angle clusters and RNA conformational motifs. We validate the technique using well-known conformational motifs, showing that the simultaneous study of the total torsion angle space leads to results consistent with known motifs reported in the literature and also to the finding of new ones.

Base Sequence↗
PubMed1992

The two similarly expressed genes encoding U3 snRNA in Schizosaccharomyces pombe lack introns.

Both genes encoding U3 small nuclear RNA (snRNA) from the budding yeast Saccharomyces cerevisiae were recently shown to be interrupted by introns of the type removed by the pre-mRNA splicing machinery. We previously described one of the two U3 genes from the fission yeast Schizosaccharomyces pombe. In the present work, the second S. pombe U3 coding sequence was identified, and direct RNA sequence analysis was used to show that neither the U3A nor the U3B gene from this organism contains an intervening sequence. Our data also demonstrate that, as expected, the two RNAs exhibit great primary- and secondary-structure conservation. These similarities are not likely to be the result of a recent gene duplication or conversion event, because the DNA sequences flanking the U3A and U3B genes have diverged substantially. A notable exception is a 19-bp block, centered 36 nucleotides upstream from the transcriptional start site, in which the two loci match in 15 positions; this motif may represent an RNA polymerase II upstream regulatory element, because related sequences are found preceding fission yeast U1, U2, U4, and U5 snRNA genes. The significance of a short conserved sequence just downstream of the U3A and U3B genes is unknown, as it is not found 3' to other snRNA coding sequences in S. pombe. The 5' one-third of U3B RNA can be folded into a dual hairpin structure, as we previously proposed for Schizosaccharomyces pombe U3A and for other lower eukaryotic U3 homologues. Quantitation of fission yeast U3A and U3B indicates that, in contrast to snR17A and B in Saccharomyces cerevisiae, these RNAs accumulate to similar levels.

Base Sequence↗
PubMed1985

Processing of precursor tRNAs in Drosophila. Processing of the 3' end involves an endonucleolytic cleavage and occurs after 5' end maturation.

Transfer RNA biosynthesis is a complex process which includes size trimming and nucleotide modification of an initial tRNA precursor. We have examined the temporal order and the nature of tRNA processing events in a Drosophila in vitro transcription/processing system using Drosophila tRNA genes as templates. RNA sequence analysis of processing products indicates that processing at both 5' and 3' ends occurs by endonucleolytic cleavage. The time course of processing of an initial tRNA precursor to mature tRNA reveals that trimming at the 5' end precedes 3' end maturation.

Animals↗
PubMed1983

Transcription of herpes simplex virus genes in vivo: overlap of a late promoter with the 3' end of the early thymidine kinase gene.

We identified in herpes simplex virus type 1-infected cells six cytoplasmic transcripts which were complementary to BamHI restriction endonuclease fragment Q. Two transcripts appeared in major amounts compared with the other four. One major transcript of about 1.4 kilobases was the mRNA for the viral thymidine kinase, was synthesized at intermediate times, and was classified as a beta transcript. The other major transcript was synthesized at late times and was classified as a gamma transcript. This late transcript was about 3 kilonucleotides long and was transcribed in the same direction as the gene for thymidine kinase. The 5' end of this late RNA was located by RNA sequence analysis and was 23 nucleotides downstream from the polyadenylation site for the thymidine kinase mRNA. This finding led to the conclusion that the control region for the 3-kilobase gamma transcript is contained within the 3' untranslated region of the thymidine kinase transcript.

Base Sequence↗
PubMed2005

Identification of plant microRNA homologs.

MicroRNAs (miRNAs) are a recently discovered class of non-coding RNAs that regulate gene and protein expression in plants and animals. MiRNAs have so far been identified mostly by specific cloning of small RNA molecules, complemented by computational methods. We present a computational identification approach that is able to identify candidate miRNA homologs in any set of sequences, given a query miRNA. The approach is based on a sequence similarity search step followed by a set of structural filters.

Algorithms↗
PubMed1991

Atypical structure of the 23S ribosomal RNA molecule in certain oral bacteria.

Ribosomal RNA (rRNA) isolated from Wolinella recta and seven related bacteria was examined by agarose gel electrophoresis. The 23S rRNA molecule could not be detected in W. recta, Wolinella curva, Bacteroides gracilis, or Bacteroides ureolyticus. In place of the 23S molecule, there were three smaller molecules of approximately 1700, 650, and 600 bases designated 23S alpha, 23S beta, and 23S delta, respectively. An intact 23S rRNA molecule could be isolated from Wolinella succinogenes, Campylobacter concisus, and Campylobacter sputorum. The cleavage sites of the W. recta 23S rRNA molecule were located by direct RNA sequence analysis and were found to be in similar locations, nucleotides 546 and 1180, as cleavage sites described in other prokaryotes. The presence or absence of the 23S rRNA molecule may be a useful marker for these micro-organisms.

Amino Acid Sequence↗
PubMed1998

Molecular analysis of the FHIT gene in human prostate cancer.

A variety of studies suggest that the FHIT gene, which encompasses the fragile site at 3p14.2, is a candidate tumor suppressor gene in several forms of human cancer. To determine whether the FHIT gene is altered in prostate carcinomas, we examined 15 prostate tumors, four normal prostate tissue specimens and RNA from a pool of 62 normal human prostate tissues (Clontech) for the integrity of FHIT transcripts, using a robust single-stage PCR and a nested PCR method. In each case a major FHIT-specific full-length product was observed. Additional aberrantly sized products, which were more numerous in the nested PCR strategy, were present at a far lower level than the full-length transcripts in 14 of 15 tumors, three of four normal human prostate tissues and in the pooled normal prostate RNA. Sequence analysis revealed that these aberrant products corresponded to alternatively spliced FHIT transcripts, which were neither more numerous nor more prominent in the tumors than in the normal prostate specimens. Deletion at the FHIT locus was also evaluated by using three intragenic polymorphic markers (D13S1481, D3S1300, and D3S1234). Allelic loss was observed in two tumors, but these genomic alterations did not correspond to the aberrant FHIT transcripts. DNA analysis, furthermore, suggested that the tumor heterogeneity was not a likely explanation for presence of normal and alternatively spliced FHIT transcripts in the prostate tumors. In conclusion, we detected neither frequent loss of heterozygosity nor tumor specific transcripts of the FHIT gene in human prostate cancer.

Acid Anhydride Hydrolases↗
PubMed1989

Chloroplast-like transfer RNA genes expressed in wheat mitochondria.

In the course of a systematic survey of wheat mitochondrial tRNA genes, we have sequenced chloroplast-like serine (trnS-GGA), phenylalanine (trnF-GAA) and cysteine (trnC-GCA) tRNA genes and their flanking regions. These genes are remnants of 'promiscuous' chloroplast DNA that has been incorporated into wheat mtDNA in the course of its evolution. Each gene differs by one or a few nucleotides from the authentic chloroplast homolog previously characterized in wheat or other plants, and each could potentially encode a functional tRNA whose secondary structure shows no deviations from the generalized model. To determine whether these chloroplast-like tRNA genes are actually expressed, wheat mitochondrial tRNAs were resolved by a series of polyacrylamide gel electrophoreses, after being specifically end-labeled in vitro by 3'-CCA addition mediated by wheat tRNA nucleotidyltransferase. Subsequent direct RNA sequence analysis identified prominent tRNA species corresponding to the mitochondrial and not the chloroplast trnS, trnF and trnC genes. This analysis also revealed chloroplast-like elongator methionine, asparagine and tryptophan tRNAs. Our results suggest that at least some chloroplast-like tRNA genes in wheat mtDNA are transcribed, with transcripts undergoing processing, post-transcriptional modification and 3'-CCA addition, to produce mature tRNAs that may participate in mitochondrial protein synthesis.

Base Sequence↗
PubMed2004

RALEE--RNA ALignment editor in Emacs.

UNLABELLED: Production of high quality multiple sequence alignments of structured RNAs relies on an iterative combination of manual editing and structure prediction. An essential feature of an RNA alignment editor is the facility to mark-up the alignment based on how it matches a given secondary structure prediction, but few available alignment editors offer such a feature. The RALEE (RNA ALignment Editor in Emacs) tool provides a simple environment for RNA multiple sequence alignment editing, including structure-specific colour schemes, utilizing helper applications for structure prediction and many more conventional editing functions. This is accomplished by extending the commonly used text editor, Emacs, which is available for Linux, most UNIX systems, Windows and Mac OS. AVAILABILITY: The ELISP source code for RALEE is freely available from http://www.sanger.ac.uk/Users/sgj/ralee/ along with documentation and examples. CONTACT: sgj@sanger.ac.uk

Algorithms↗
PubMed1991

A human autoantibody specific for a unique conserved region of 28 S ribosomal RNA inhibits the interaction of elongation factors 1 alpha and 2 with ribosomes.

An autoantibody reactive with a conserved sequence of 28 S rRNA (anti-28 S) was identified in serum from a patient with systemic lupus erythematosus. Anti-28 S protected a unique 59-nucleotide fragment synthesized in vitro against RNase T1 digestion. RNA sequence analysis revealed that it corresponded to residues 1944-2002 in human 28 S rRNA and 1767-1825 in mouse 28 S rRNA. These sequences are identical and highly conserved throughout all known eukaryotic 28 S rRNAs. In addition, this fragment is homologous to residues 1052-1110 of Escherichia coli 23 S rRNA that lies within the GTP hydrolysis center of the 50 S ribosomal subunit. Anti-28 S and its Fab fragments strongly inhibited poly(U)-directed polyphenylalanine synthesis, but had no effect on ribosomal peptidyltransferase activity. This effect resulted from inhibition of the binding of elongation factors EF-1 alpha and EF-2 to ribosomes and of the associated GTP hydrolysis. The inhibitory effect was almost completely suppressed by preincubation of anti-28 S with 28 S rRNA or in vitro synthesized RNA fragments containing the immunoreactive region. These results show that the immunoreactive conserved region of 28 S rRNA participates in the interaction of ribosomes with the two elongation factors in protein synthesis.

Animals↗