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Results for “Ribosome Profiling”

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At least 649 records · Page 36Linked to original sources

PubMed2002

Molecular characterization of Blastocystis isolates from birds by PCR with diagnostic primers and restriction fragment length polymorphism analysis of the small subunit ribosomal RNA gene.

Seven Blastocystis isolates from birds were genotyped by polymerase chain reaction (PCR) using diagnostic primers, and the homology among isolates was then confirmed by restriction fragment length polymorphism (RFLP) analysis of the small subunit ribosomal RNA gene. A zoonotic genotype, subtype 4, was detected in three isolates by PCR, but the RFLP profiles of these isolates differed from those of the subtype 4 reported previously from a human. None of the other four isolates were amplified with any subtype-specific primer sets, and the profiles of these isolates were different from any ribodemes of Blastocystis hominis reported previously. In the present study, the seven isolates from birds were classified into three genetically distinct genotypes. Therefore, birds appear to be infected with multiple genotypes. It is unclear whether the isolates examined in the present study are zoonotic or not. Therefore, it is important to determine the phylogenetic relationships between unknown genotypes from birds and the multiple genotypes of B. hominis.

Animals↗

PubMed2006

The world of subinhibitory antibiotic concentrations.

Although antibiotics have long been known to have multiple effects on bacterial cells at low concentrations, it is only with the advent of genome transcription analyses that these activities have been studied in detail at the level of cell metabolism. It has been shown that all antibiotics, regardless of their receptors and mode of action, exhibit the phenomenon of hormesis and provoke considerable transcription activation at low concentrations. These analyses should be of value in providing information on antibiotic side-effects, in bioactive natural product discovery and antibiotic mode-of-action studies.

Anti-Bacterial Agents↗

PubMed2006

Composition of the bacterial flora in tonsilloliths.

Tonsilloliths are a potential cause of oral malodor. In this study, microbial profiles and composition of tonsilloliths were determined using culture-independent molecular methods and scanning electron microscopy. 16S ribosomal RNA bacterial genes (16S rDNAs) isolated from tonsilloliths of 6 individuals were amplified by PCR and cloned into Escherichia coli. Partial 16S rDNA sequences of approximately 600 bases of cloned inserts were used to determine species identity by comparison with sequences of known species. Characteristics of bacteria on the surface and inside the tonsillolith were analyzed using scanning electron microscopy. Anaerobic bacteria detected in tonsilloliths belonged to the genera Eubacterium, Fusobacterium, Megasphaera, Porphyromonas, Prevotella, Selenomonas and Tannerella, all of which appear to be associated with production of volatile sulfur compounds. Electron microscopy revealed cocci and rods on the surface and rods predominating inside the tonsilloliths. These results support the tonsillolith as an origin of oral malodor.

Adult↗

PubMed2002

Subspecies variation in Greek strains of Chlamydophila abortus.

The Greek chlamydial strains FAS, FAG, VPG and LLG, isolated from aborted sheep or goat foetuses, had been previously characterized as divergent on the basis of mouse cross-protection experiments, with LLG and its homologous POS significantly different from the rest in inclusion morphology, polypeptide profiles and reactivity with monoclonal antibodies. To determine the genetic basis of their divergence the 16S-23S ribosomal intergenic spacer was analysed by RFLP analysis of PCR 16SF2/23R amplicons. Using the restriction enzymes BfaI, SfcI, HpaI, BclI, DdeI and AclI, the strains were classified as Chlamydophila abortus. However, digestion with RsaI made it possible to differentiate strains FAS, FAG and VPG from strains LLG and POS, generating DNA fragments of 530/55 and 585bp, respectively. By subsequent sequence analysis of the 23S domain I rRNA gene only strain FAS was identical to reference strain A22 of C. abortus. Strains FAG and VPG presented an identical nucleotide deviation at position 593 of signature sequences. Strains LLG and POS presented three identical nucleotide deviations at positions 156, 186 and 307. Variation within the domain I signature sequences for the examined abortion strains was < or =0.69%. In conclusion, substantial genetic and biological diversity among strains of C. abortus was demonstrated, suggesting that subspecies variation status for certain strains may be applicable. Our findings suggest that differentiation may be possible at a subspecies level by RFLP analysis.

Abortion, Veterinary↗

PubMed1969

Daily rhythms in hepatic polysome profiles and tyrosine transaminase activity: role of dietary protein.

Hepatic polysome profiles vary in untreated rats as a function of time of day. The ratio of polysomes to total ribosomes increases from 50 to 73 per cent in darkness. There is also a daily rhythm in tyrosine transaminase activity which resembles but does not coincide with the polysome rhythm. Both rhythms are dependent on the cyclic ingestion of dietary protein, and disappear in rats given a protein-free diet.

Animals↗

PubMed1978

Two chromatographically separable forms of Escherichia coli elongation factor Tu.

Two forms of the elongation factor Tu from Escherichia coli have been separated by chromatography on DEAE-Sephadex A50. Obvious chromatographic artifacts have been ruled out by investigation of the elution profile of GDP (a component of the column buffer as well as a ligand of Tu) and by rechromatography of the two components, either separately to give the component peaks or together to give a double peak. The two components have been confirmed as Tu by the poly(uridylic)-dependent polyphenylalanine synthesis and by the distribution of the Tu protein as quantitated from sodium dodecyl sulfate/polyacrylamide gel electrophoresis. Complexes with the elongation factors Ts and G have also been ruled out by activity profiles and by quantitation of the protein distribution, again on gels. The distribution of the two forms between ribosomal and supernatant fractions has been examined: one is bound preferentially to the ribosomal fraction and the other is found in the supernatant fraction. The possible significance of this is discussed.

Bacterial Proteins↗

PubMed2005

Immunopurification of polyribosomal complexes of Arabidopsis for global analysis of gene expression.

Immunoaffinity purification of polyribosomes (polysomes) from crude leaf extracts of Arabidopsis (Arabidopsis thaliana) was achieved with transgenic genotypes that overexpress a translational fusion of a ribosomal protein (RP) with a His(6)-FLAG dual epitope tag. In plants with a cauliflower mosaic virus 35S:HF-RPL18 transgene immunopurification with anti-FLAG agarose beads yielded 60-Svedberg ribosomal subunits, intact 80-Svedberg monosomes and polysomes. Sucrose density gradient fractionation of the purified complexes demonstrated that the distribution of polysome size was similar in crude cell extracts and the purified complexes. The immunopurified complexes included putative cytosolic RPs of Arabidopsis and ribosome-associated proteins, as well as full-length transcripts of high and low abundance. Whole-genome profiling using long DNA oligonucleotide-based microarrays provided a high level of reproducibility between polysomal mRNA samples immunopurified from two independent biological replicates (r approximately 0.90). Comparison of immunopurified and total cellular RNA samples revealed that for most of the genes, the mRNAs were associated with the epitope-tagged polysomal complexes, with an average relative level of association of 62.06% +/- 4.39%. The results demonstrate that the immunopurification of polysomes can be a valuable tool for the quantification of mRNAs present in translation complexes in plant cells. This technology can be extended to evaluation of mRNA populations at the cell- or tissue-specific level by regulation of the tagged RP with distinct promoters.

Arabidopsis↗

PubMed1975

Activity of polyribosomes from the muscle of normal and dystrophic mice in cell-free amino-acid incorporation.

Polyribosomes sedimenting in the manner characteristic of those from embryonic chick muscle, as described by Heywood et al. in 1967 (Proc. Natl Acad. Sci. U.S.A. 57,1002--1009) were reproducibly obtained from normal mouse muscle by homogenization of the muscle with a Dounce homogenizer. The polyribosome profiles of dystrophic muscle were qualitatively similar to those of normal muscle except that the relative amount of ribosomes in polyribosome complexes was smaller (44% +/- 3S.E.) in dystrophic muscle than in normal muscle (67% +/- 4S.E.). In spite of this difference, polyribosomes from dystrophic muscle incorporated amino acids in vitro at a faster rate and produced a larger amount of polypeptide at the end of the reaction than polyribosomes from normal muscle.

Amino Acids↗

PubMed2004

Application of molecular fingerprinting for qualitative assessment of small-intestinal bacterial diversity in dogs.

The aims of this study were to evaluate the use of molecular fingerprinting for assessment of bacterial diversity in canine duodenal juice and to evaluate the variation in the small intestinal microflora at repeated sampling. Two groups of dogs were used. Duodenal juice was collected from eight dogs euthanized for an unrelated project (group 1). Duodenal juice was also collected endoscopically from six dogs at weekly intervals for a total of 3 weeks (group 2). The variable V6-V8 region of bacterial 16S ribosomal DNA was amplified, and PCR amplicons separated by denaturing gradient gel electrophoresis (DGGE). The reproducibility of DGGE profiles and variations in bacterial diversity between dogs were evaluated by comparing similarity indices (Dice's coefficient; 100% represents complete identity) of DGGE profiles from group 1 dogs. Weekly variations in the flora of the small intestine were evaluated by comparison of DGGE profiles from different time points within the same individuals in group 2. The mean (+/- standard deviation) similarity of DGGE profiles of duodenal juice between the dogs in group 1 was 38.3 +/- 15.7% (range, 12.5 to 76.65%). There was a significantly higher variation in DGGE profiles between different dogs than between duplicates obtained from the same dog (P < 0.0001). DGGE profiles from samples collected at different time points varied within individuals, possibly due to variation over time or slight variation in sampling location. DGGE profiles indicate that dogs have a highly diverse microflora of the small intestine, with marked differences between individual dogs.

Animals↗

PubMed2003

Production of ribosome components in effector CD4+ T cells is accelerated by TCR stimulation and coordinated by ERK-MAPK.

Effector CD4+ T cells rapidly activate high-level cytokine expression following TCR stimulation. Consistent with accelerated protein production in these cells, global mRNA profiles revealed that, after cytokines, the most impressive cluster of activated genes encode rRNA-maturation factors. Activation of these genes was ERK-MAPK dependent, accompanied by increased rRNA transcription and faster maturation kinetics, and much greater in effector CD4+ T cells than in naive cells. Ribosomal protein subunit (RPS) synthesis was also ERK-MAPK dependent and increased to match rRNA production, but without evident increase in RPS mRNA. Instead, stimulation promoted polysome loading of RPS mRNA via cis-acting, 5'-terminal oligopyrimidines. These results demonstrate how, in response to extracellular signals, effector CD4+ T cells coordinately increase multiple ribosomal components to accommodate burgeoning cytokine production.

Animals↗

PubMed1985

Quantitative assessment of the polysome profile of the livers of mice treated with tetracycline or doxycycline.

The influence of tetracycline and doxycycline (10-100 micrograms/g i.v.) on the aggregational state of ribosomes from mouse liver was tested. Both drugs caused a disaggregation of the ribosomes as evidenced by a rise of the monosomes + disomes/polysomes ratio. Tetracycline was much more potent than doxycycline, the minimum effective doses for tetracycline being 10 micrograms/g i.v. as compared to 100 micrograms/g for doxycycline. The results show that tetracycline but not doxycycline at therapeutic dose range may interfere with the protein synthesis of the liver.

Animals↗

PubMed1992

Characterization and functional expression in Escherichia coli of the sodium/proton/glutamate symport proteins of Bacillus stearothermophilus and Bacillus caldotenax.

The genes encoding the Na+/H+/L-glutamate symport proteins of the thermophilic organisms Bacillus stearothermophilus (gltTBs) and Bacillus caldotenax (gltTBc) were cloned by complementation of Escherichia coli JC5412 for growth on glutamate as sole source of carbon, energy and nitrogen. The nucleotide sequences of the gltTBs and gltTBc genes were determined. In both cases the translated sequences corresponded with proteins of 421 amino acid residues (96.7% amino acid identity between GltTBs and GltTBc). Putative promoter, terminator and ribosome-binding-site sequences were found in the flanking regions. These expression signals were functional in E. coli. The hydropathy profiles indicate that the proteins are hydrophobic and could form 12 membrane-spanning regions. The Na+/H+ coupled L-glutamate symport proteins GltTBs and GltTBc are homologous to the strictly H+ coupled L-glutamate transport protein of E. coli K-12 (overall 57.2% identity). Functional expression of glutamate transport activity was demonstrated by uptake of glutamate in whole cells and membrane vesicles. In accordance with previous observations (de Vrij et al., 1989; Heyne et al., 1991), glutamate uptake was driven by the electrochemical gradients of sodium ions and protons.

Amino Acid Sequence↗

PubMed2002

Identification and characterization of genes abnormally expressed in wing-deficient mutant (flügellos) of the silkworm, Bombyx mori.

The wing-deficient mutant, flügellos (fl), of the silkworm lacks four wings in the pupa and the adult, due to aberrant wing morphogenesis during metamorphosis. To elucidate the mechanisms of wing-specific deficiencies in the fl mutant, we used mRNA differential display and identified five genes abnormally expressed in the fl wing discs. Northern blot and RT-PCR analyses revealed that four genes were overexpressed, but the fifth one was not transcribed in the fl wing discs. The expression level of ribosome-associated protein p40 in the fl wing discs was elevated approximately 10 times compared to the wild-type (WT) discs. Another overexpressed gene CB10 encodes a novel wing-specific protein with a putative zinc-finger motif. Overexpression of two components of extracellular matrix, cuticle protein 18 (BMCP18) and a fibrillin-like protein AD10, may result in the abnormal wing morphogenesis in the fl mutant. In contrast, a novel member of multifunctional Ca2+-binding protein annexins, designated as annexin b13 (Anx b13), was expressed dominantly in the wing discs of WT but completely repressed in the fl tissues. Strong expression of Anx b13 in wing discs during the fourth and fifth instar indicates that ANX B13 plays an important role in wing morphogenesis.

Amino Acid Sequence↗

PubMed2005

Genotypic and phenotypic characterization of a biofilm-forming Serratia plymuthica isolate from a raw vegetable processing line.

Recently, we isolated from a raw vegetable processing line a Serratia strain with strong biofilm-forming capacity and which produced N-acyl-L-homoserine lactones (AHLs). Within the Enterobacteriaceae, strains of the genus Serratia are a frequent cause of human nosocomial infections; in addition, biofilm formation is often associated with persistent infections. In the current report, we describe the detailed characterization of the isolate using a variety of genotypic and phenotypic criteria. Although the strain was identified as Serratia plymuthica on the basis of its small subunit ribosomal RNA (16S rRNA) gene sequence, it differed from the S. plymuthica type strain in production of pigment and antibacterial compounds, and in AHL production profile. Nevertheless, the identification as S. plymuthica could be confirmed by gyrB phylogeny and DNA:DNA hybridization.

4-Butyrolactone↗

PubMed1996

Yeast proteins related to the p40/laminin receptor precursor are essential components of the 40 S ribosomal subunit.

We report here the isolation of two genes from the yeast, Saccharomyces cerevisiae, that encode proteins closely related to mammalian p40/laminin receptor precursors (LRPs). The yeast genes, designated YST1 and YST2, encode proteins with over 95% amino acid sequence identity with one another and over 60% identity with the human p40/laminin receptor precursor. The Yst/p40/37-LRP proteins are also more distantly related to the S2 family of ribosomal proteins. Analysis of the distribution of Yst1 tagged with the c-myc epitope revealed that the Yst proteins are components of the 40 S ribosomal subunit. Disruption of either YST1 or YST2 causes a significant reduction in growth rate, while disruption of both genes is lethal. Compared to wild type, polysome profiles in strains lacking either YST1 or YST2 show a pronounced shift from larger to smaller polysomes. This shift is accompanied by a substantial increase in free 60 S subunits and reduced levels of 40 S subunits. We conclude that the Yst proteins are required for translation and contribute to the assembly and/or stability of the 40 S ribosomal subunit.

Amino Acid Sequence↗

PubMed2004

Transcription profiling of cyclic AMP signaling in Candida albicans.

We used transcription profiling in Candida albicans to investigate cellular regulation involving cAMP. We found that many genes require the adenylyl cyclase Cdc35p for proper expression. These include genes encoding ribosomal subunit proteins and RNA polymerase subunit proteins, suggesting that growth could be controlled in part by cAMP-mediated modulation of gene expression. Other genes influenced by loss of adenylyl cyclase are involved in metabolism, the cell wall, and stress response and include a group of genes of unknown function that are unique to C. albicans. The profiles generated by loss of the adenylyl cyclase regulator Ras1p and a downstream effector Efg1p were also examined. The loss of Ras1p function disturbs the expression of a subset of the genes regulated by adenylyl cyclase, suggesting both that the primary role of Ras1p in transcriptional regulation involves its influence on the function of Cdc35p and that there are Ras1p independent roles for Cdc35p. The transcription factor Efg1p is also needed for the expression of many genes; however, these genes are distinct from those modulated by Cdc35p with the exception of a class of hyphal-specific genes. Therefore transcription profiling establishes that cAMP plays a key role in the overall regulation of gene expression in C. albicans, and enhances our detailed understanding of the circuitry controlling this regulation.

Candida albicans↗

PubMed1999

Molecular variation in Leymus species and populations.

Icelandic populations of European lymegrass [Leymus arenarius (L.) Hochst.] were examined using amplified fragment length polymorphism (AFLP) and restriction fragment length polymorphism (RFLP) of the major ribosomal genes (18S-5.8S-26S rDNA), in comparison with Alaskan populations of its closely related species L. mollis (Trin.) Pilger. The AFLP profiles emerged as two distinct entities, clearly separating the two species, and based on species-specific bands it was simple to distinguish these two morphologically similar species. The rDNA-RFLPs also differentiated the species. Within species, the Icelandic L. arenarius was more homogeneous than the Alaskan L. mollis, and its variation was dispersed over geographically different populations, suggesting a common gene pool. The variation among the Alaskan L. mollis was more extensive and its interrupted pattern may be the result of gene introgression at subspecies level. Within a 40-year-old population of L. mollis established in Iceland from Alaskan material, the molecular profiles separated old and new genotypes. Both AFLP and rDNA revealed the new genotypes to be extremely similar. This rapid change in allele frequency is thought to be the result of adaptation to a new environment.

Alaska↗

PubMed2002

Identification of differentially expressed genes in human uterine leiomyomas using differential display.

UNLABELLED: In searching of differentially expressed genes in human uterine leiomyomas, differential display was used with twelve pairs of primers to compare human uterine leiomyomas with matched myometrium. False positives were eliminated by reverse Northern analysis. Positives were confirmed by Northern blot analysis. RESULTS: Four of 69 cDNA fragments (3 up-regulated named L1, L2 and L3 and 1 down-regulated named M1 in leiomyoma) were confirmed by Northern analysis. Sequence comparison and Northern analysis proved that L1 is exactly the human ribosomal protein S19. It was present ubiquitously in 13 tissues tested but in various levels and even in different size. L1 was highly expressed in parotidean cystadenocarcinoma, pancreatic cancer and breast cancer examined. No mutations have been found in human uterine leiomyomas (n=6). CONCLUSIONS: hRPS19 overexpression might be a universal signal in rapid cell growth tissues.

DNA, Complementary↗