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Sample records for adjacent rrna nucleotide

  1. Role of conserved nucleotides in building the 16 S rRNA binding site for ribosomal protein S15.

    PubMed

    Serganov, A; Bénard, L; Portier, C; Ennifar, E; Garber, M; Ehresmann, B; Ehresmann, C

    2001-01-26

    Ribosomal protein S15 recognizes a highly conserved target on 16 S rRNA, which consists of two distinct binding regions. Here, we used extensive site-directed mutagenesis on a Escherichia coli 16 S rRNA fragment containing the S15 binding site, to investigate the role of conserved nucleotides in protein recognition and to evaluate the relative contribution of the two sites. The effect of mutations on S15 recognition was studied by measuring the relative binding affinity, RNA probing and footprinting. The crystallographic structure of the Thermus thermophilus complex allowed molecular modelling of the E. coli complex and facilitated interpretation of biochemical data. Binding is essentially driven by site 1, which includes a three-way junction constrained by a conserved base triple and cross-strand stacking. Recognition is based mainly on shape complementarity, and the role of conserved nucleotides is to maintain a unique backbone geometry. The wild-type base triple is absolutely required for protein interaction, while changes in the conserved surrounding nucleotides are partially tolerated. Site 2, which provides functional groups in a conserved G-U/G-C motif, contributes only modestly to the stability of the interaction. Binding to this motif is dependent on binding at site 1 and is allowed only if the two sites are in the correct relative orientation. Non-conserved bulged nucleotides as well as a conserved purine interior loop, although not directly involved in recognition, are used to provide an appropriate flexibility between the two sites. In addition, correct binding at the two sites triggers conformational adjustments in the purine interior loop and in a distal region, which are known to be involved for subsequent binding of proteins S6 and S18. Thus, the role of site 1 is to anchor S15 to the rRNA, while binding at site 2 is aimed to induce a cascade of events required for subunit assembly. PMID:11162092

  2. Methylation of 23S rRNA Nucleotide G748 by RlmAII Methyltransferase Renders Streptococcus pneumoniae Telithromycin Susceptible

    PubMed Central

    Sato, Yoshiharu; Shoji, Tatsuma; Yamamoto, Tomoko

    2013-01-01

    Several posttranscriptional modifications of bacterial rRNAs are important in determining antibiotic resistance or sensitivity. In all Gram-positive bacteria, dimethylation of nucleotide A2058, located in domain V of 23S rRNA, by the dimethyltransferase Erm(B) results in low susceptibility and resistance to telithromycin (TEL). However, this is insufficient to produce high-level resistance to TEL in Streptococcus pneumoniae. Inactivation of the methyltransferase RlmAII, which methylates the N-1 position of nucleotide G748, located in hairpin 35 of domain II of 23S rRNA, results in increased resistance to TEL in erm(B)-carrying S. pneumoniae. Sixteen TEL-resistant mutants (MICs, 16 to 32 μg/ml) were obtained from a clinically isolated S. pneumoniae strain showing low TEL susceptibility (MIC, 2 μg/ml), with mutation resulting in constitutive dimethylation of A2058 because of nucleotide differences in the regulatory region of erm(B) mRNA. Primer extension analysis showed that the degree of methylation at G748 in all TEL-resistant mutants was significantly reduced by a mutation in the gene encoding RlmAII to create a stop codon or change an amino acid residue. Furthermore, RNA footprinting with dimethyl sulfate and a molecular modeling study suggested that methylation of G748 may contribute to the stable interaction of TEL with domain II of 23S rRNA, even after dimethylation of A2058 by Erm(B). This novel finding shows that methylation of G748 by RlmAII renders S. pneumoniae TEL susceptible. PMID:23716046

  3. The antibiotics micrococcin and thiostrepton interact directly with 23S rRNA nucleotides 1067A and 1095A.

    PubMed Central

    Rosendahl, G; Douthwaite, S

    1994-01-01

    The antibiotics thiostrepton and micrococcin bind to the GTPase region in domain II of 23S rRNA, and inhibit ribosomal A-site associated reactions. When bound to the ribosome, these antibiotics alter the accessibility of nucleotides 1067A and 1095A towards chemical reagents. Plasmid-coded Escherichia coli 23S rRNAs with single mutations at positions 1067 or 1095 were expressed in vivo. Mutant ribosomes are functional in protein synthesis, although those with transversion mutations function less effectively. Antibiotics were bound under conditions where wild-type and mutant ribosomes compete in the same reaction for drug molecules; binding was analysed by allele-specific footprinting. Transversion mutations at 1067 reduce thiostrepton binding more than 1000-fold. The 1067G substitution gives a more modest decrease in thiostrepton binding. The changes at 1095 slightly, but significantly, lower the affinity of ribosomes for thiostrepton, again with the G mutation having the smallest effect. Micrococcin binding to ribosomes is reduced to a far greater extent than thiostrepton by all the 1067 and 1095 mutations. Extrapolating these results to growing cells, mutation of nucleotide 1067A confers resistance towards micrococcin and thiostrepton, while substitutions at 1095A confer micrococcin resistance, and increase tolerance towards thiostrepton. These data support an rRNA tertiary structure model in which 1067A and 1095A lie in close proximity, and are key components in the drug binding site. None of the mutations alters either the higher order rRNA structure or the binding of r-proteins. We therefore conclude that thiostrepton and micrococcin interact directly with 1067A and 1095A. Images PMID:8127673

  4. Methanosarcina acetivorans 16S rRNA and transcription factor nucleotide fluctuation with implications in exobiology and pathology

    NASA Astrophysics Data System (ADS)

    Holden, Todd; Tremberger, G., Jr.; Cheung, E.; Subramaniam, R.; Sullivan, R.; Schneider, P.; Flamholz, A.; Marchese, P.; Hiciano, O.; Yao, H.; Lieberman, D.; Cheung, T.

    2008-08-01

    Cultures of the methane-producing archaea Methanosarcina, have recently been isolated from Alaskan sediments. It has been proposed that methanogens are strong candidates for exobiological life in extreme conditions. The spatial environmental gradients, such as those associated with the polygons on Mars' surface, could have been produced by past methanogenesis activity. The 16S rRNA gene has been used routinely to classify phenotypes. Using the fractal dimension of nucleotide fluctuation, a comparative study of the 16S rRNA nucleotide fluctuation in Methanosarcina acetivorans C2A, Deinococcus radiodurans, and E. coli was conducted. The results suggest that Methanosarcina acetivorans has the lowest fractal dimension, consistent with its ancestral position in evolution. Variation in fluctuation complexity was also detected in the transcription factors. The transcription factor B (TFB) was found to have a higher fractal dimension as compared to transcription factor E (TFE), consistent with the fact that a single TFB in Methanosarcina acetivorans can code three different TATA box proteins. The average nucleotide pair-wise free energy of the DNA repair genes was found to be highest for Methanosarcina acetivorans, suggesting a relatively weak bonding, which is consistent with its low prevalence in pathology. Multitasking capacity comparison of type-I and type-II topoisomerases has been shown to correlate with fractal dimension using the methicillin-resistant strain MRSA 252. The analysis suggests that gene adaptation in a changing chemical environment can be measured in terms of bioinformatics. Given that the radiation resistant Deinococcus radiodurans is a strong candidate for an extraterrestrial origin and that the cold temperature Psychrobacter cryohalolentis K5 can function in Siberian permafrost, the fractal dimension comparison in this study suggests that a chemical resistant methanogen could exist in extremely cold conditions (such as that which existed on early

  5. 23S rRNA nucleotides in the peptidyl transferase center are essential for tryptophanase operon induction.

    PubMed

    Yang, Rui; Cruz-Vera, Luis R; Yanofsky, Charles

    2009-06-01

    Distinct features of the ribosomal peptide exit tunnel are known to be essential for recognition of specific amino acids of a nascent peptidyl-tRNA. Thus, a tryptophan residue at position 12 of the peptidyl-tRNA TnaC-tRNA(Pro) leads to the creation of a free tryptophan binding site within the ribosome at which bound tryptophan inhibits normal ribosome functions. The ribosomal processes that are inhibited are hydrolysis of TnaC-tRNA(Pro) by release factor 2 and peptidyl transfer of TnaC of TnaC-tRNA(Pro) to puromycin. These events are normally performed in the ribosomal peptidyl transferase center. In the present study, changes of 23S rRNA nucleotides in the 2585 region of the peptidyl transferase center, G2583A and U2584C, were observed to reduce maximum induction of tna operon expression by tryptophan in vivo without affecting the concentration of tryptophan necessary to obtain 50% induction. The growth rate of strains with ribosomes with either of these changes was not altered appreciably. In vitro analyses with mutant ribosomes with these changes showed that tryptophan was not as efficient in protecting TnaC-tRNA(Pro) from puromycin action as wild-type ribosomes. However, added tryptophan did prevent sparsomycin action as it normally does with wild-type ribosomes. These findings suggest that these two mutational changes act by reducing the ability of ribosome-bound tryptophan to inhibit peptidyl transferase activity rather than by reducing the ability of the ribosome to bind tryptophan. Thus, the present study identifies specific nucleotides within the ribosomal peptidyl transferase center that appear to be essential for effective tryptophan induction of tna operon expression. PMID:19329641

  6. Guanine nucleotide metabolism in a mutant strain of Escherichia coli with a temperature sensitive lesion in rRNA synthesis.

    PubMed

    Harris, J S; Chaney, S G

    1978-12-21

    We have described a mutant of Escherichia coli (designated 2S142) which shows specific inhibition of rRNA synthesis at 42 degrees C. ppGpp levels increase at the restrictive temperature, as expected. However, when the cells are returned to 30 degrees C, rRNA synthesis resumes before ppGpp levels have returned to normal. Furthermore, when ppGpp levels are decreased by the addition of tetracycline or choramphenicol, rRNA synthesis does not resume at 42 degrees C. Also, a derivative of 2S142 with a temperature-sensitive G factor (which cannot synthesize either protein or ppGpp at 42 degrees C) shows identical kinetics of rRNA shut-off at 42 degrees C as 2S142. Thus, the elevated ppGpp levels in this mutant do not appear to be directly responsible for the cessation of rRNA synthesis at 42 degrees C. PMID:367439

  7. A genome walking strategy for the identification of nucleotide sequences adjacent to known regions.

    PubMed

    Wang, Hailong; Yao, Ting; Cai, Mei; Xiao, Xiuqing; Ding, Xuezhi; Xia, Liqiu

    2013-02-01

    To identify the transposon insertion sites in a soil actinomycete, Saccharopolyspora spinosa, a genome walking approach, termed SPTA-PCR, was developed. In SPTA-PCR, a simple procedure consisting of TA cloning and a high stringency PCR, following the single primer-mediated, randomly-primed PCR, can eliminate non-target DNA fragments and obtain target fragments specifically. Using SPTA-PCR, the DNA sequence adjacent to the highly conserved region of lectin coding gene in onion plant, Allium chinense, was also cloned. PMID:23108875

  8. Update on Pneumocystis carinii f. sp. hominis Typing Based on Nucleotide Sequence Variations in Internal Transcribed Spacer Regions of rRNA Genes

    PubMed Central

    Lee, Chao-Hung; Helweg-Larsen, Jannik; Tang, Xing; Jin, Shaoling; Li, Baozheng; Bartlett, Marilyn S.; Lu, Jang-Jih; Lundgren, Bettina; Lundgren, Jens D.; Olsson, Mats; Lucas, Sebastian B.; Roux, Patricia; Cargnel, Antonietta; Atzori, Chiara; Matos, Olga; Smith, James W.

    1998-01-01

    Pneumocystis carinii f. sp. hominis isolates from 207 clinical specimens from nine countries were typed based on nucleotide sequence variations in the internal transcribed spacer regions I and II (ITS1 and ITS2, respectively) of rRNA genes. The number of ITS1 nucleotides has been revised from the previously reported 157 bp to 161 bp. Likewise, the number of ITS2 nucleotides has been changed from 177 to 192 bp. The number of ITS1 sequence types has increased from 2 to 15, and that of ITS2 has increased from 3 to 14. The 15 ITS1 sequence types are designated types A through O, and the 14 ITS2 types are named types a through n. A total of 59 types of P. carinii f. sp. hominis were found in this study. PMID:9508304

  9. Targeting single-nucleotide polymorphisms in the 16S rRNA gene to detect and differentiate Legionella pneumophila and non-Legionella pneumophila species.

    PubMed

    Zhan, Xiao-Yong; Hu, Chao-Hui; Zhu, Qing-Yi

    2016-08-01

    A PCR-based method targeting single-nucleotide polymorphisms (SNPs) in the 16S rRNA gene was developed for differential identification of Legionella pneumophila and non-Legionella pneumophila. Based on the bioinformatics analysis for 176 Legionella 16S rRNA gene fragments of 56 different Legionella species, a set of SNPs, A(628)C(629) was found to be highly specific to L. pneumophila strains. A multiplex assay was designed that was able to distinguish sites with limited sequence heterogeneity between L. pneumophila and non-L. pneumophila in the targeted 16S rRNA gene. The assay amplified a 261-bp amplicon for Legionella spp. and a set of 203- and 97-bp amplicons only specific to L. pneumophila species. Among 49 ATCC strains and 284 Legionella isolates from environmental water and clinical samples, 100 % of L. pneumophila and non-L. pneumophila strains were correctly identified and differentiated by this assay. The assay presents a more rapid, sensitive and alternative method to the currently available PCR-sequencing detection and differentiation method. PMID:27112927

  10. Events during eucaryotic rRNA transcription initiation and elongation: Conversion from the closed to the open promoter complex requires nucleotide substrates

    SciTech Connect

    Bateman, E.; Paule, M.R.

    1988-05-01

    Chemical footprinting and topological analysis were carried out on the Acanthamoeba castellanii rRNA transcription initiation factor (TIF) and RNA polymerase I complexes with DNA during transcription initiation and elongation. The results show that the binding of TIF and polymerase to the promoter does not alter the supercoiling of the DNA template and the template does not become sensitive to modification by diethylpyro-carbonate, which can identify melted DNA regions. Thus, in contrast to bacterial RNA polymerase, the eucaryotic RNA polymerase I-promoter complex is in a closed configuration preceding addition of nucleotides in vitro. Initiation and 3'-O-methyl CTP-limited translocation by RNA polymerase I results in separation of the polymerase-TIF footprints, leaving the TIF footprint unaltered. In contrast, initiation and translocation result in a significant change in the conformation of the polymerase-DNA complex, culminating in an unwound DNA region of at least 10 base pairs.

  11. The nucleotide sequence of the large ribosomal RNA gene and the adjacent tRNA genes from rat mitochondria.

    PubMed Central

    Saccone, C; Cantatore, P; Gadaleta, G; Gallerani, R; Lanave, C; Pepe, G; Kroon, A M

    1981-01-01

    We have sequenced the Eco R(1) fragment D from rat mitochondrial DNA. It contains one third of the tRNA (Val) gene (the remaining part has been sequenced from the 3' end of the Eco R(1) fragment A) the complete gene for the large mt 16S rRNA, the tRNA (Leu) gene and the 5' end of an unidentified reading frame. The mt gene for the large rRNA from rat has been aligned with the homologous genes from mouse and human using graphic computer programs. Hypervariable regions at the center of the molecule and highly conserved regions toward the 3' end have been detected. The mt gene for tRNA Leu is of the conventional type and its primary structure is highly conserved among mammals. The mt gene for tRNA(Val) shows characteristics similar to those of other mt tRNA genes but the degree of homology is lower. Comparative studies confirm that AGA and AGG are read as stop codons in mammalian mitochondria. PMID:6913863

  12. Nucleotide sequence composition adjacent to intronic splice sites improves splicing efficiency via its effect on pre-mRNA local folding in fungi.

    PubMed

    Zafrir, Zohar; Tuller, Tamir

    2015-10-01

    RNA splicing is the central process of intron removal in eukaryotes known to regulate various cellular functions such as growth, development, and response to external signals. The canonical sequences indicating the splicing sites needed for intronic boundary recognition are well known. However, the roles and evolution of the local folding of intronic and exonic sequence features adjacent to splice sites has yet to be thoroughly studied. Here, focusing on four fungi (Saccharomyces cerevisiae, Schizosaccharomyces pombe, Aspergillus nidulans, and Candida albicans), we performed for the first time a comprehensive high-resolution study aimed at characterizing the encoding of intronic splicing efficiency in pre-mRNA transcripts and its effect on intron evolution. Our analysis supports the conjecture that pre-mRNA local folding strength at intronic boundaries is under selective pressure, as it significantly affects splicing efficiency. Specifically, we show that in the immediate region of 12-30 nucleotides (nt) surrounding the intronic donor site there is a preference for weak pre-mRNA folding; similarly, in the region of 15-33 nt surrounding the acceptor and branch sites there is a preference for weak pre-mRNA folding. We also show that in most cases there is a preference for strong pre-mRNA folding further away from intronic splice sites. In addition, we demonstrate that these signals are not associated with gene-specific functions, and they correlate with splicing efficiency measurements (r = 0.77, P = 2.98 × 10(-21)) and with expression levels of the corresponding genes (P = 1.24 × 10(-19)). We suggest that pre-mRNA folding strength in the above-mentioned regions has a direct effect on splicing efficiency by improving the recognition of intronic boundaries. These new discoveries are contributory steps toward a broader understanding of splicing regulation and intronic/transcript evolution. PMID:26246046

  13. Modified nucleotides m(2)G966/m(5)C967 of Escherichia coli 16S rRNA are required for attenuation of tryptophan operon.

    PubMed

    Prokhorova, Irina V; Osterman, Ilya A; Burakovsky, Dmitry E; Serebryakova, Marina V; Galyamina, Maria A; Pobeguts, Olga V; Altukhov, Ilya; Kovalchuk, Sergey; Alexeev, Dmitry G; Govorun, Vadim M; Bogdanov, Alexey A; Sergiev, Petr V; Dontsova, Olga A

    2013-01-01

    Ribosomes contain a number of modifications in rRNA, the function of which is unclear. Here we show--using proteomic analysis and dual fluorescence reporter in vivo assays--that m(2)G966 and m(5)C967 in 16S rRNA of Escherichia coli ribosomes are necessary for correct attenuation of tryptophan (trp) operon. Expression of trp operon is upregulated in the strain where RsmD and RsmB methyltransferases were deleted, which results in the lack of m(2)G966 and m(5)C967 modifications. The upregulation requires the trpL attenuator, but is independent of the promotor of trp operon, ribosome binding site of the trpE gene, which follows trp attenuator and even Trp codons in the trpL sequence. Suboptimal translation initiation efficiency in the rsmB/rsmD knockout strain is likely to cause a delay in translation relative to transcription which causes misregulation of attenuation control of trp operon. PMID:24241179

  14. Modified nucleotides m2G966/m5C967 of Escherichia coli 16S rRNA are required for attenuation of tryptophan operon

    NASA Astrophysics Data System (ADS)

    Prokhorova, Irina V.; Osterman, Ilya A.; Burakovsky, Dmitry E.; Serebryakova, Marina V.; Galyamina, Maria A.; Pobeguts, Olga V.; Altukhov, Ilya; Kovalchuk, Sergey; Alexeev, Dmitry G.; Govorun, Vadim M.; Bogdanov, Alexey A.; Sergiev, Petr V.; Dontsova, Olga A.

    2013-11-01

    Ribosomes contain a number of modifications in rRNA, the function of which is unclear. Here we show - using proteomic analysis and dual fluorescence reporter in vivo assays - that m2G966 and m5C967 in 16S rRNA of Escherichia coli ribosomes are necessary for correct attenuation of tryptophan (trp) operon. Expression of trp operon is upregulated in the strain where RsmD and RsmB methyltransferases were deleted, which results in the lack of m2G966 and m5C967 modifications. The upregulation requires the trpL attenuator, but is independent of the promotor of trp operon, ribosome binding site of the trpE gene, which follows trp attenuator and even Trp codons in the trpL sequence. Suboptimal translation initiation efficiency in the rsmB/rsmD knockout strain is likely to cause a delay in translation relative to transcription which causes misregulation of attenuation control of trp operon.

  15. The Homopolyadenylate and Adjacent Nucleotides at the 3′-Terminus of 30-40S RNA Subunits in the Genome of Murine Sarcoma-Leukemia Virus

    PubMed Central

    Rho, Hyune Mo; Green, Maurice

    1974-01-01

    Adenosine is the major 3′OH-terminal nucleoside of the 60-70S RNA genome of the murine sarcoma-leukemia virus, its 30-40S RNA subunits, and the poly(A) segments derived by RNase treatment of both RNA species, as determined by periodate oxidation-[3H]-borohydride reduction. The binding 30-40S RNA to oligo(dT)-cellulose suggests that most viral RNA subunits contain poly(A). The molecular weight of poly(A) derived from viral RNA by digestion with RNase and purified by affinity chromatography is 64,000-68,000, as determined by gel electrophoresis. From the size of poly(A) and the poly(A) content of viral RNA (1.6%), it is estimated that there is about one poly(A) segment for each viral 30-40S RNA subunit. The results of 3′-termini labeling with [3H]borohydride, in vivo labeling with [3H]adenosine, and base composition of [32P]poly(A) indicate that a homopoly(A) segment is located at the 3′-end of a 30-40S RNA subunit. The homogeneous poly(A) segments isolated from RNase T1 digests of 60-70S [32P]RNA consist of one cytidylate, one uridylate, and about 190 adenylate residues, while those isolated from RNase A digests consist exclusively of adenylate residues. These results indicate that -G(C,U)A190AOH is the 3′-terminal nucleotide sequence of the viral 30-40S RNA subunits. PMID:4366765

  16. RlmCD-mediated U747 methylation promotes efficient G748 methylation by methyltransferase RlmAII in 23S rRNA in Streptococcus pneumoniae; interplay between two rRNA methylations responsible for telithromycin susceptibility

    PubMed Central

    Shoji, Tatsuma; Takaya, Akiko; Sato, Yoshiharu; Kimura, Satoshi; Suzuki, Tsutomu; Yamamoto, Tomoko

    2015-01-01

    Adenine at position 752 in a loop of helix 35 from positions 745 to 752 in domain II of 23S rRNA is involved in binding to the ribosome of telithromycin (TEL), a member of ketolides. Methylation of guanine at position 748 by the intrinsic methyltransferase RlmAII enhances binding of telithromycin (TEL) to A752 in Streptococcus pneumoniae. We have found that another intrinsic methylation of the adjacent uridine at position 747 enhances G748 methylation by RlmAII, rendering TEL susceptibility. U747 and another nucleotide, U1939, were methylated by the dual-specific methyltransferase RlmCD encoded by SP_1029 in S. pneumoniae. Inactivation of RlmCD reduced N1-methylated level of G748 by RlmAII in vivo, leading to TEL resistance when the nucleotide A2058, located in domain V of 23S rRNA, was dimethylated by the dimethyltransferase Erm(B). In vitro methylation of rRNA showed that RlmAII activity was significantly enhanced by RlmCD-mediated pre-methylation of 23S rRNA. These results suggest that RlmCD-mediated U747 methylation promotes efficient G748 methylation by RlmAII, thereby facilitating TEL binding to the ribosome. PMID:26365244

  17. Comparison of the nucleotide sequences of 16S rRNA, 444 Ep-ank, and groESL heat shock operon genes in naturally occurring Ehrlichia equi and human granulocytic ehrlichiosis agent isolates from Northern California.

    PubMed

    Chae, J S; Foley, J E; Dumler, J S; Madigan, J E

    2000-04-01

    We examined 11 naturally occurring isolates of Ehrlichia equi in horses and two human granulocytic ehrlichiosis agent isolates in California for sequence diversity in three genes. Ehrlichia equi isolates were from Sierra (n = 6), Mendocino (n = 3), Sonoma (n = 1), and Marin (n = 1) counties, and human granulocytic ehrlichiosis (HGE) agent isolates were obtained from Humboldt county. PCR with specific primers for 16S rRNA, 444 Ep-ank and groESL heat shock operon genes successfully produced amplicons for all 13 clinical samples. The 444 Ep-ank gene of the HGE agent and E. equi isolates from northern California is different from the eastern U.S. isolates BDS and USG3. The translated amino acid sequence of the groESL heat shock operon gene fragment is identical among E. equi, the HGE agent, and E. phagocytophila, with the exception of the northern Californian equine CASOLJ isolate. Microheterogeneity was observed in the 16S rRNA gene sequences of HGE agent and E. equi isolates from northern California. These results suggest that E. equi and the HGE agent found in California are similar or identical but may differ from the isolates of equine and human origin found in the eastern United States. PMID:10747108

  18. Promoter of the Mycoplasma pneumoniae rRNA operon.

    PubMed Central

    Hyman, H C; Gafny, R; Glaser, G; Razin, S

    1988-01-01

    RNA transcripts starting from the 5' end of the single Mycoplasma pneumoniae rRNA operon were analyzed by several methods. By primer extension analysis a start site was found 62 nucleotides upstream from the start site of the 16S rRNA. This site was preceded by a putative Pribnow box; however, a defined -35 recognition region was absent. The cloned rRNA operon was transcribed in vitro by using purified RNA polymerase of Escherichia coli. A single start site could be demonstrated within a few nucleotides of the start site found by primer extension analysis of M. pneumoniae transcripts. When fragments from the cloned operon were used as hybridization probes, S1 nuclease mapping yielded a single transcript extending approximately 193 nucleotides upstream from the 16S rRNA start site. The region surrounding this endpoint did not resemble any known promoter sequence. Dot blot hybridization of M. pneumoniae RNA to three oligonucleotides consisting of nucleotides -5 to -21, -38 to -54, and -112 to -132 (from the start of the 16S rRNA gene) indicated that most rRNA transcripts were processed at the stem site preceding the 16S rRNA gene. The majority of the longer precursor transcripts, extending beyond this point, did not extend further upstream to an oligonucleotide consisting of nucleotides -112 to -132. It was concluded that transcription of the rRNA operon of M. pneumoniae is initiated by a single promoter. The nucleotide sequence of the region is presented. Images PMID:2838465

  19. Case of localized recombination in 23S rRNA genes from divergent bradyrhizobium lineages associated with neotropical legumes.

    PubMed

    Parker, M A

    2001-05-01

    Enzyme electrophoresis and rRNA sequencing were used to analyze relationships of Bradyrhizobium sp. nodule bacteria from four papilionoid legumes (Clitoria javitensis, Erythrina costaricensis, Rhynchosia pyramidalis, and Desmodium axillare) growing on Barro Colorado Island (BCI), Panama. Bacteria with identical multilocus allele profiles were commonly found in association with two or more legume genera. Among the 16 multilocus genotypes (electrophoretic types [ETs]) detected, six ETs formed a closely related cluster that included isolates from all four legume taxa. Bacteria from two other BCI legumes (Platypodium and Machaerium) sampled in a previous study were also identical to certain ETs in this group. Isolates from different legume genera that had the same ET had identical nucleotide sequences for both a 5' portion of the 23S rRNA and the nearly full-length 16S rRNA genes. These results suggest that Bradyrhizobium genotypes with low host specificity may be prevalent in this tropical forest. Parsimony analysis of 16S rRNA sequence variation indicated that most isolates were related to Bradyrhizobium japonicum USDA 110, although one ET sampled from C. javitensis had a 16S rRNA gene highly similar to that of Bradyrhizobium elkanii USDA 76. However, this isolate displayed a mosaic structure within the 5' 23S rRNA region: one 84-bp segment was identical to that of BCI isolate Pe1-3 (a close relative of B. japonicum USDA 110, based on 16S rRNA data), while an adjacent 288-bp segment matched that of B. elkanii USDA 76. This mosaic structure is one of the first observations suggesting recombination in nature between Bradyrhizobium isolates related to B. japonicum versus B. elkanii. PMID:11319084

  20. [Unusual motifs of the nucleotide sequence adjacent to the putative transcription initiation site in the rDNA intergenic spacer of diploid wheat Triticum urartu Thum. ex Gandil, T. boeoticum Boiss, and T. monococcum L].

    PubMed

    Akhunov, E D; Chemeris, A V; Vakhitov, V A

    1997-11-01

    In the intergenic spacer (IGS) of rDNA of diploid wheats Triticum urartu, T. boeoticum, and T. monococcum, the uncommon motives adjacent to the site of transcription initiation (TIS) are revealed. They are located in the region from -6 to +1 relative to the putative TIS and are not encountered in cereals studied earlier. In T. urartu and T. boeoticum, the motif TACTATG has been revealed, in T. monococcum--TATTATG, while diploid Aegilops speltoides has the motif TATAGTA, typical of the remaining cereal species. The TIS-surrounding rDNA IGS region of diploid wheats was compared to the correspondent known rDNA IGS regions of different plant and animal species. PMID:9480224

  1. Automated Identification of Nucleotide Sequences

    NASA Technical Reports Server (NTRS)

    Osman, Shariff; Venkateswaran, Kasthuri; Fox, George; Zhu, Dian-Hui

    2007-01-01

    STITCH is a computer program that processes raw nucleotide-sequence data to automatically remove unwanted vector information, perform reverse-complement comparison, stitch shorter sequences together to make longer ones to which the shorter ones presumably belong, and search against the user s choice of private and Internet-accessible public 16S rRNA databases. ["16S rRNA" denotes a ribosomal ribonucleic acid (rRNA) sequence that is common to all organisms.] In STITCH, a template 16S rRNA sequence is used to position forward and reverse reads. STITCH then automatically searches known 16S rRNA sequences in the user s chosen database(s) to find the sequence most similar to (the sequence that lies at the smallest edit distance from) each spliced sequence. The result of processing by STITCH is the identification of the most similar well-described bacterium. Whereas previously commercially available software for analyzing genetic sequences operates on one sequence at a time, STITCH can manipulate multiple sequences simultaneously to perform the aforementioned operations. A typical analysis of several dozen sequences (length of the order of 103 base pairs) by use of STITCH is completed in a few minutes, whereas such an analysis performed by use of prior software takes hours or days.

  2. rRNA fragmentation induced by a yeast killer toxin.

    PubMed

    Kast, Alene; Klassen, Roland; Meinhardt, Friedhelm

    2014-02-01

    Virus like dsDNA elements (VLE) in yeast were previously shown to encode the killer toxins PaT and zymocin, which target distinct tRNA species via specific anticodon nuclease (ACNase) activities. Here, we characterize a third member of the VLE-encoded toxins, PiT from Pichia inositovora, and identify PiOrf4 as the cytotoxic subunit by conditional expression in Saccharomyces cerevisiae. In contrast to the tRNA targeting toxins, however, neither a change of the wobble uridine modification status by introduction of elp3 or trm9 mutations nor tRNA overexpression rescued from PiOrf4 toxicity. Consistent with a distinct RNA target, expression of PiOrf4 causes specific fragmentation of the 25S and 18S rRNA. A stable cleavage product comprising the first ∼ 130 nucleotides of the 18S rRNA was purified and characterized by linker ligation and subsequent reverse transcription; 3'-termini were mapped to nucleotide 131 and 132 of the 18S rRNA sequence, a region showing some similarity to the anticodon loop of tRNA(Glu)(UUC), the zymocin target. PiOrf4 residues Glu9 and His214, corresponding to catalytic sites Glu9 and His209 in the ACNase subunit of zymocin are essential for in vivo toxicity and rRNA fragmentation, raising the possibility of functionally conserved RNase modules in both proteins. PMID:24308908

  3. Adjacent segment disease.

    PubMed

    Virk, Sohrab S; Niedermeier, Steven; Yu, Elizabeth; Khan, Safdar N

    2014-08-01

    EDUCATIONAL OBJECTIVES As a result of reading this article, physicians should be able to: 1. Understand the forces that predispose adjacent cervical segments to degeneration. 2. Understand the challenges of radiographic evaluation in the diagnosis of cervical and lumbar adjacent segment disease. 3. Describe the changes in biomechanical forces applied to adjacent segments of lumbar vertebrae with fusion. 4. Know the risk factors for adjacent segment disease in spinal fusion. Adjacent segment disease (ASD) is a broad term encompassing many complications of spinal fusion, including listhesis, instability, herniated nucleus pulposus, stenosis, hypertrophic facet arthritis, scoliosis, and vertebral compression fracture. The area of the cervical spine where most fusions occur (C3-C7) is adjacent to a highly mobile upper cervical region, and this contributes to the biomechanical stress put on the adjacent cervical segments postfusion. Studies have shown that after fusion surgery, there is increased load on adjacent segments. Definitive treatment of ASD is a topic of continuing research, but in general, treatment choices are dictated by patient age and degree of debilitation. Investigators have also studied the risk factors associated with spinal fusion that may predispose certain patients to ASD postfusion, and these data are invaluable for properly counseling patients considering spinal fusion surgery. Biomechanical studies have confirmed the added stress on adjacent segments in the cervical and lumbar spine. The diagnosis of cervical ASD is complicated given the imprecise correlation of radiographic and clinical findings. Although radiological and clinical diagnoses do not always correlate, radiographs and clinical examination dictate how a patient with prolonged pain is treated. Options for both cervical and lumbar spine ASD include fusion and/or decompression. Current studies are encouraging regarding the adoption of arthroplasty in spinal surgery, but more long

  4. Strain identification and 5S rRNA gene characterization of the hyperthermophilic archaebacterium Sulfolobus acidocaldarius.

    PubMed Central

    Durovic, P; Kutay, U; Schleper, C; Dennis, P P

    1994-01-01

    A commonly used laboratory Sulfolobus strain has been unambiguously identified as Sulfolobus acidocaldarius DSM639. The 5S rRNA gene from this strain was cloned and sequenced. It differs at 17 of 124 positions from the identical 5S rRNA sequences from Sulfolobus solfataricus and a strain apparently misidentified as S. acidocaldarius. Analysis of the transcripts from the 5S rRNA gene failed to identify any precursor extending a significant distance beyond the 5' or 3' boundary of the 5S rRNA-coding sequence. This result suggests that the primary transcript of the 5S rRNA gene corresponds in length (within 1 or 2 nucleotides) to the mature 5S rRNA sequence found in 50S ribosomal subunits. Images PMID:8288546

  5. Lessons from an evolving rRNA: 16S and 23S rRNA structures from a comparative perspective

    NASA Technical Reports Server (NTRS)

    Gutell, R. R.; Larsen, N.; Woese, C. R.

    1994-01-01

    The 16S and 23S rRNA higher-order structures inferred from comparative analysis are now quite refined. The models presented here differ from their immediate predecessors only in minor detail. Thus, it is safe to assert that all of the standard secondary-structure elements in (prokaryotic) rRNAs have been identified, with approximately 90% of the individual base pairs in each molecule having independent comparative support, and that at least some of the tertiary interactions have been revealed. It is interesting to compare the rRNAs in this respect with tRNA, whose higher-order structure is known in detail from its crystal structure (36) (Table 2). It can be seen that rRNAs have as great a fraction of their sequence in established secondary-structure elements as does tRNA. However, the fact that the former show a much lower fraction of identified tertiary interactions and a greater fraction of unpaired nucleotides than the latter implies that many of the rRNA tertiary interactions remain to be located. (Alternatively, the ribosome might involve protein-rRNA rather than intramolecular rRNA interactions to stabilize three-dimensional structure.) Experimental studies on rRNA are consistent to a first approximation with the structures proposed here, confirming the basic assumption of comparative analysis, i.e., that bases whose compositions strictly covary are physically interacting. In the exhaustive study of Moazed et al. (45) on protection of the bases in the small-subunit rRNA against chemical modification, the vast majority of bases inferred to pair by covariation are found to be protected from chemical modification, both in isolated small-subunit rRNA and in the 30S subunit. The majority of the tertiary interactions are reflected in the chemical protection data as well (45). On the other hand, many of the bases not shown as paired in Fig. 1 are accessible to chemical attack (45). However, in this case a sizeable fraction of them are also protected against chemical

  6. Dinoflagellate 17S rRNA sequence inferred from the gene sequence: Evolutionary implications.

    PubMed

    Herzog, M; Maroteaux, L

    1986-11-01

    We present the complete sequence of the nuclear-encoded small-ribosomal-subunit RNA inferred from the cloned gene sequence of the dinoflagellate Prorocentrum micans. The dinoflagellate 17S rRNA sequence of 1798 nucleotides is contained in a family of 200 tandemly repeated genes per haploid genome. A tentative model of the secondary structure of P. micans 17S rRNA is presented. This sequence is compared with the small-ribosomal-subunit rRNA of Xenopus laevis (Animalia), Saccharomyces cerevisiae (Fungi), Zea mays (Planta), Dictyostelium discoideum (Protoctista), and Halobacterium volcanii (Monera). Although the secondary structure of the dinoflagellate 17S rRNA presents most of the eukaryotic characteristics, it contains sufficient archaeobacterial-like structural features to reinforce the view that dinoflagellates branch off very early from the eukaryotic lineage. PMID:16578795

  7. Dinoflagellate 17S rRNA sequence inferred from the gene sequence: Evolutionary implications

    PubMed Central

    Herzog, Michel; Maroteaux, Luc

    1986-01-01

    We present the complete sequence of the nuclear-encoded small-ribosomal-subunit RNA inferred from the cloned gene sequence of the dinoflagellate Prorocentrum micans. The dinoflagellate 17S rRNA sequence of 1798 nucleotides is contained in a family of 200 tandemly repeated genes per haploid genome. A tentative model of the secondary structure of P. micans 17S rRNA is presented. This sequence is compared with the small-ribosomal-subunit rRNA of Xenopus laevis (Animalia), Saccharomyces cerevisiae (Fungi), Zea mays (Planta), Dictyostelium discoideum (Protoctista), and Halobacterium volcanii (Monera). Although the secondary structure of the dinoflagellate 17S rRNA presents most of the eukaryotic characteristics, it contains sufficient archaeobacterial-like structural features to reinforce the view that dinoflagellates branch off very early from the eukaryotic lineage. PMID:16578795

  8. Thermus thermophilus 16S rRNA is transcribed from an isolated transcription unit.

    PubMed Central

    Hartmann, R K; Erdmann, V A

    1989-01-01

    A cloned 16S rRNA gene from the extreme thermophilic eubacterium Thermus thermophilus HB8 was used to characterize the in vivo expression of the 16S rRNA genes in this organism by nuclease S1 mapping. The gene represents an isolated transcription unit encoding solely 16S rRNA. Under exponential growth conditions, transcription was initiated at a single promoter, which represents the structural equivalent of Escherichia coli rrn P2 promoters. The promoter-leader region was very similar to the E. coli rrn P2 promoter-leader segment that is responsible for antitermination. The T. thermophilus leader region was approximately 85 nucleotides shorter than its E. coli P2 counterpart. Potential processing intermediates were correlated with a proposed secondary structure of T. thermophilus pre-16S rRNA. Images PMID:2722737

  9. Primary and secondary structure of 5.8S rRNA from the silkgland of Bombyx mori.

    PubMed Central

    Fujiwara, H; Kawata, Y; Ishikawa, H

    1982-01-01

    Nucleotide sequence of 5.8S rRNA of the silkworm, Bombyx mori has been determined by gel sequencing methods. The 5.8S rRNA was the longest so far reported, with the 5'-terminal sequence several nucleotides longer than those of the other organisms. Upon constructing the secondary structure in accordance with the "burp gun" model (12), the Bombyx 5.8S rRNA formed a wide-open "muzzle" due to several unpaired bases at the ends. The overall structure also appeared less stable with less G . C pairs and more unpaired bases than that of the HeLa 5.8S rRNA. These structural features may be essential for those 5.8S rRNAs which interact with 28S rRNAs containing the hidden break to form a stable complex. PMID:7088713

  10. Sequence heterogeneity in the two 16S rRNA genes of Phormium yellow leaf phytoplasma.

    PubMed Central

    Liefting, L W; Andersen, M T; Beever, R E; Gardner, R C; Forster, R L

    1996-01-01

    Phormium yellow leaf (PYL) phytoplasma causes a lethal disease of the monocotyledon, New Zealand flax (Phormium tenax). The 16S rRNA genes of PYL phytoplasma were amplified from infected flax by PCR and cloned, and the nucleotide sequences were determined. DNA sequencing and Southern hybridization analysis of genomic DNA indicated the presence of two copies of the 16S rRNA gene. The two 16S rRNA genes exhibited sequence heterogeneity in 4 nucleotide positions and could be distinguished by the restriction enzymes BpmI and BsrI. This is the first record in which sequence heterogeneity in the 16S rRNA genes of a phytoplasma has been determined by sequence analysis. A phylogenetic tree based on 16S rRNA gene sequences showed that PYL phytoplasma is most closely related to the stolbur and German grapevine yellows phytoplasmas, which form the stolbur subgroup of the aster yellows group. This phylogenetic position of PYL phytoplasma was supported by 16S/23S spacer region sequence data. PMID:8795200

  11. Chicken rRNA Gene Cluster Structure

    PubMed Central

    Dyomin, Alexander G.; Koshel, Elena I.; Kiselev, Artem M.; Saifitdinova, Alsu F.; Galkina, Svetlana A.; Fukagawa, Tatsuo; Kostareva, Anna A.

    2016-01-01

    Ribosomal RNA (rRNA) genes, whose activity results in nucleolus formation, constitute an extremely important part of genome. Despite the extensive exploration into avian genomes, no complete description of avian rRNA gene primary structure has been offered so far. We publish a complete chicken rRNA gene cluster sequence here, including 5’ETS (1836 bp), 18S rRNA gene (1823 bp), ITS1 (2530 bp), 5.8S rRNA gene (157 bp), ITS2 (733 bp), 28S rRNA gene (4441 bp) and 3’ETS (343 bp). The rRNA gene cluster sequence of 11863 bp was assembled from raw reads and deposited to GenBank under KT445934 accession number. The assembly was validated through in situ fluorescent hybridization analysis on chicken metaphase chromosomes using computed and synthesized specific probes, as well as through the reference assembly against de novo assembled rRNA gene cluster sequence using sequenced fragments of BAC-clone containing chicken NOR (nucleolus organizer region). The results have confirmed the chicken rRNA gene cluster validity. PMID:27299357

  12. 5S rRNA sequences from four marine invertebrates and implications for base pairing models of metazoan sequences.

    PubMed

    Walker, W F; Doolittle, W F

    1983-08-11

    The nucleotide sequences of 5S rRNAs from the starfish Asterias vulgaris, the squid Illex illecebrosus, the sipunculid Phascolopsis gouldii and the jellyfish Aurelia aurita were determined. The sequence from Asterias lends support for one of two previous base pairing models for helix E in metazoan sequences. The Aurelia sequence differs by five nucleotides from that previously reported and does not violate the consensus secondary structure model for eukaryotic 5S rRNA. PMID:6136024

  13. Correlated Evolution of Nucleotide Positions within Splice Sites in Mammals

    PubMed Central

    Denisov, Stepan; Bazykin, Georgii; Favorov, Alexander; Mironov, Andrey; Gelfand, Mikhail

    2015-01-01

    Splice sites (SSs)—short nucleotide sequences flanking introns—are under selection for spliceosome binding, and adhere to consensus sequences. However, non-consensus nucleotides, many of which probably reduce SS performance, are frequent. Little is known about the mechanisms maintaining such apparently suboptimal SSs. Here, we study the correlations between strengths of nucleotides occupying different positions of the same SS. Such correlations may arise due to epistatic interactions between positions (i.e., a situation when the fitness effect of a nucleotide in one position depends on the nucleotide in another position), their evolutionary history, or to other reasons. Within both the intronic and the exonic parts of donor SSs, nucleotides that increase (decrease) SS strength tend to co-occur with other nucleotides increasing (respectively, decreasing) it, consistent with positive epistasis. Between the intronic and exonic parts of donor SSs, the correlations of nucleotide strengths tend to be negative, consistent with negative epistasis. In the course of evolution, substitutions at a donor SS tend to decrease the strength of its exonic part, and either increase or do not change the strength of its intronic part. In acceptor SSs, the situation is more complicated; the correlations between adjacent positions appear to be driven mainly by avoidance of the AG dinucleotide which may cause aberrant splicing. In summary, both the content and the evolution of SSs is shaped by a complex network of interdependences between adjacent nucleotides that respond to a range of sometimes conflicting selective constraints. PMID:26642327

  14. Correlated Evolution of Nucleotide Positions within Splice Sites in Mammals.

    PubMed

    Denisov, Stepan; Bazykin, Georgii; Favorov, Alexander; Mironov, Andrey; Gelfand, Mikhail

    2015-01-01

    Splice sites (SSs)--short nucleotide sequences flanking introns--are under selection for spliceosome binding, and adhere to consensus sequences. However, non-consensus nucleotides, many of which probably reduce SS performance, are frequent. Little is known about the mechanisms maintaining such apparently suboptimal SSs. Here, we study the correlations between strengths of nucleotides occupying different positions of the same SS. Such correlations may arise due to epistatic interactions between positions (i.e., a situation when the fitness effect of a nucleotide in one position depends on the nucleotide in another position), their evolutionary history, or to other reasons. Within both the intronic and the exonic parts of donor SSs, nucleotides that increase (decrease) SS strength tend to co-occur with other nucleotides increasing (respectively, decreasing) it, consistent with positive epistasis. Between the intronic and exonic parts of donor SSs, the correlations of nucleotide strengths tend to be negative, consistent with negative epistasis. In the course of evolution, substitutions at a donor SS tend to decrease the strength of its exonic part, and either increase or do not change the strength of its intronic part. In acceptor SSs, the situation is more complicated; the correlations between adjacent positions appear to be driven mainly by avoidance of the AG dinucleotide which may cause aberrant splicing. In summary, both the content and the evolution of SSs is shaped by a complex network of interdependences between adjacent nucleotides that respond to a range of sometimes conflicting selective constraints. PMID:26642327

  15. Modified Method of rRNA Structure Analysis Reveals Novel Characteristics of Box C/D RNA Analogues.

    PubMed

    Filippova, J A; Stepanov, G A; Semenov, D V; Koval, O A; Kuligina, E V; Rabinov, I V; Richter, V A

    2015-01-01

    Ribosomal RNA (rRNA) maturation is a complex process that involves chemical modifications of the bases or sugar residues of specific nucleotides. One of the most abundant types of rRNA modifications, ribose 2'-O-methylation, is guided by ribonucleoprotein complexes containing small nucleolar box C/D RNAs. Since the majority of 2'-O-methylated nucleotides are located in the most conserved regions of rRNA that comprise functionally important centers of the ribosome, an alteration in a 2'-O-methylation profile can affect ribosome assembly and function. One of the key approaches for localization of 2'-O-methylated nucleotides in long RNAs is a method based on the termination of reverse transcription. The current study presents an adaptation of this method for the use of fluorescently labeled primers and analysis of termination products by capillary gel electrophoresis on an automated genetic analyzer. The developed approach allowed us to analyze the influence of the synthetic analogues of box C/D RNAs on post-transcriptional modifications of human 28S rRNA in MCF-7 cells. It has been established that the transfection of MCF-7 cells with a box C/D RNA analogue leads to an enhanced modification level of certain native sites of 2'-O-methylation in the target rRNA. The observed effect of synthetic RNAs on the 2'-O-methylation of rRNA in human cells demonstrates a path towards targeted regulation of rRNA post-transcriptional maturation. The described approach can be applied in the development of novel diagnostic methods for detecting diseases in humans. PMID:26085946

  16. Interactions between 23S rRNA and tRNA in the ribosomal E site.

    PubMed Central

    Bocchetta, M; Xiong, L; Shah, S; Mankin, A S

    2001-01-01

    Interactions between tRNA or its analogs and 23S rRNA in the large ribosomal subunit were analyzed by RNA footprinting and by modification-interference selection. In the E site, tRNA protected bases G2112, A2392, and C2394 of 23S rRNA. Truncated tRNA, lacking the anticodon stem-loop, protected A2392 and C2394, but not G2112, and tRNA derivatives with a shortened 3' end protected only G2112, but not A2392 or C2394. Modification interference revealed C2394 as the only accessible nucleotide in 23S rRNA whose modification interferes with binding of tRNA in the large ribosomal subunit E site. The results suggest a direct contact between A76 of tRNA A76 and C2394 of 23S rRNA. Protections at G2112 may reflect interaction of this 23S rRNA region with the tRNA central fold. PMID:11214181

  17. Molecular Evolution of Mycoplasma capricolum subsp. capripneumoniae Strains, Based on Polymorphisms in the 16S rRNA Genes

    PubMed Central

    Pettersson, Bertil; Bölske, Göran; Thiaucourt, François; Uhlén, Mathias; Johansson, Karl-Erik

    1998-01-01

    Mycoplasma capricolum subsp. capripneumoniae belongs to the so-called Mycoplasma mycoides cluster and is the causal agent of contagious caprine pleuropneumonia (CCPP). All members of the M. mycoides cluster have two rRNA operons. The sequences of the 16S rRNA genes of both rRNA operons from 20 strains of M. capricolum subsp. capripneumoniae of different geographical origins in Africa and Asia were determined. Nucleotide differences which were present in only one of the two operons (polymorphisms) were detected in 24 positions. The polymorphisms were not randomly distributed in the 16S rRNA genes, and some of them were found in regions of low evolutionary variability. Interestingly, 11 polymorphisms were found in all the M. capricolum subsp. capripneumoniae strains, thus defining a putative ancestor. A sequence length difference between the 16S rRNA genes in a poly(A) region and 12 additional polymorphisms were found in only one or some of the strains. A phylogenetic tree was constructed by comparative analysis of the polymorphisms, and this tree revealed two distinct lines of descent. The nucleotide substitution rate of strains within line II was up to 50% higher than within line I. A tree was also constructed from individual operonal 16S rRNA sequences, and the sequences of the two operons were found to form two distinct clades. The topologies of both clades were strikingly similar, which supports the use of 16S rRNA sequence data from homologous operons for phylogenetic studies. The strain-specific polymorphism patterns of the 16S rRNA genes of M. capricolum subsp. capripneumoniae may be used as epidemiological markers for CCPP. PMID:9573185

  18. Plant Cyclic Nucleotide Signalling

    PubMed Central

    Martinez-Atienza, Juliana; Van Ingelgem, Carl; Roef, Luc

    2007-01-01

    The presence of the cyclic nucleotides 3′,5′-cyclic adenyl monophosphate (cAMP) and 3′,5′-cyclic guanyl monophosphate (cGMP) in plants is now generally accepted. In addition, cAMP and cGMP have been implicated in the regulation of important plant processes such as stomatal functioning, monovalent and divalent cation fluxes, chloroplast development, gibberellic acid signalling, pathogen response and gene transcription. However, very little is known regarding the components of cyclic nucleotide signalling in plants. In this addendum, the evidence for specific mechanisms of plant cyclic nucleotide signalling is evaluated and discussed. PMID:19704553

  19. Evolving nucleotide binding surfaces

    NASA Technical Reports Server (NTRS)

    Kieber-Emmons, T.; Rein, R.

    1981-01-01

    An analysis is presented of the stability and nature of binding of a nucleotide to several known dehydrogenases. The employed approach includes calculation of hydrophobic stabilization of the binding motif and its intermolecular interaction with the ligand. The evolutionary changes of the binding motif are studied by calculating the Euclidean deviation of the respective dehydrogenases. Attention is given to the possible structural elements involved in the origin of nucleotide recognition by non-coded primordial polypeptides.

  20. Mutations of mitochondrial 12S rRNA in gastric carcinoma and their significance

    PubMed Central

    Han, Cheng-Bo; Ma, Jia-Ming; Xin, Yan; Mao, Xiao-Yun; Zhao, Yu-Jie; Wu, Dong-Ying; Zhang, Su-Min; Zhang, Yu-Kui

    2005-01-01

    AIM: To detect the variations of mitochondrial 12S rRNA in patients with gastric carcinoma, and to study their significance and the relationship between these variations and the genesis of gastric carcinoma. METHODS: PCR amplified mitochondrial 12S rRNA of 44 samples including 22 from gastric carcinoma tissues and 22 from adjacent normal tissues, was detected by direct DNA sequencing. Then laser capture microdissection technique (LCM) was used to separate the cancerous cells and dysplasia cells with specific mutations. Denaturing high performance liquid chromatography (DHPLC) plus allele-specific PCR (AS-PCR), nest-PCR and polyacrylamide gel electrophoresis (PAGE) were used to further evaluate this mutant property and quantitative difference of mutant type between cancerous and dysplasia cells. Finally, RNAdraw biosoft was used to analyze the RNA secondary structure of mutant-type 12S rRNA. RESULTS: Compared with Mitomap database, some new variations were found, among which np652 G insertion and np716 T-G transversion were found only in cancerous tissues. There was a statistic difference in the frequency of 12S rRNA variation between intestinal type (12/17, 70.59%) and diffusive type (5/17, 29.41%) of gastric carcinoma (P<0.05). DHPLC analysis showed that 12S rRNA np652 G insertion and np716 T-G transversion were heteroplasmic mutations. The frequency of 12S rRNA variation in cancerous cells was higher than that in dysplasia cells (P<0.01). 12S rRNA np652 G insertion showed obviously negative effects on the stability of 12S rRNA secondary structure, while others such as T-G transversion did not. CONCLUSION: The mutations of mitochondrial 12S rRNA may be associated with the occurrence of intestinal-type gastric carcinoma. Most variations exist both in gastric carcinomas and in normal tissues, and they might not be the characteristics of tumors. However, np652 G insertion and np716 T-G transversion may possess some molecular significance in gastric carcinogenesis

  1. Comparative structural analysis of eubacterial 5S rRNA by oxidation of adenines in the N-1 position.

    PubMed Central

    Pieler, T; Schreiber, A; Erdmann, V A

    1984-01-01

    Adenines in free 5S rRNA from Escherichia coli, Bacillus stearothermophilus and Thermus thermophilus have been oxidized at their N-1 position using monoperphthalic acid. The determination of the number of adenine 1-N-oxides was on the basis of UV spectroscopic data of the intact molecule. Identification of the most readily accessible nucleotides by sequencing gel analysis reveals that they are located in conserved positions within loops, exposed hairpin loops and single-base bulge loops. Implications for the structure and function of 5S rRNA will be discussed on the basis of this comparative analysis. Images PMID:6201825

  2. Technologically important extremophile 16S rRNA sequence Shannon entropy and fractal property comparison with long term dormant microbes

    NASA Astrophysics Data System (ADS)

    Holden, Todd; Gadura, N.; Dehipawala, S.; Cheung, E.; Tuffour, M.; Schneider, P.; Tremberger, G., Jr.; Lieberman, D.; Cheung, T.

    2011-10-01

    Technologically important extremophiles including oil eating microbes, uranium and rocket fuel perchlorate reduction microbes, electron producing microbes and electrode electrons feeding microbes were compared in terms of their 16S rRNA sequences, a standard targeted sequence in comparative phylogeny studies. Microbes that were reported to have survived a prolonged dormant duration were also studied. Examples included the recently discovered microbe that survives after 34,000 years in a salty environment while feeding off organic compounds from other trapped dead microbes. Shannon entropy of the 16S rRNA nucleotide composition and fractal dimension of the nucleotide sequence in terms of its atomic number fluctuation analyses suggest a selected range for these extremophiles as compared to other microbes; consistent with the experience of relatively mild evolutionary pressure. However, most of the microbes that have been reported to survive in prolonged dormant duration carry sequences with fractal dimension between 1.995 and 2.005 (N = 10 out of 13). Similar results are observed for halophiles, red-shifted chlorophyll and radiation resistant microbes. The results suggest that prolonged dormant duration, in analogous to high salty or radiation environment, would select high fractal 16S rRNA sequences. Path analysis in structural equation modeling supports a causal relation between entropy and fractal dimension for the studied 16S rRNA sequences (N = 7). Candidate choices for high fractal 16S rRNA microbes could offer protection for prolonged spaceflights. BioBrick gene network manipulation could include extremophile 16S rRNA sequences in synthetic biology and shed more light on exobiology and future colonization in shielded spaceflights. Whether the high fractal 16S rRNA sequences contain an asteroidlike extra-terrestrial source could be speculative but interesting.

  3. The phylogeny of intestinal porcine spirochetes (Serpulina species) based on sequence analysis of the 16S rRNA gene.

    PubMed Central

    Pettersson, B; Fellström, C; Andersson, A; Uhlén, M; Gunnarsson, A; Johansson, K E

    1996-01-01

    Four type or reference strains and twenty-two field strains of intestinal spirochetes isolated from Swedish pig herds were subjected to phylogenetic analysis based on 16S rRNA sequences. Almost complete (>95%) 16S rRNA sequences were obtained by solid-phase DNA sequencing of in vitro-amplified rRNA genes. The genotypic patterns were compared with a previously proposed biochemical classification scheme, comprising beta-hemolysis, indole production, hippurate hydrolysis, and alpha-galactosidase, alpha-glucosidase, and beta-glucosidase activities. Comparison of the small-subunit rRNA sequences showed that the strains of the genus Serpulina were closely related. Phylogenetic trees were constructed, and three clusters were observed. This was also confirmed by signature nucleotide analysis of the serpulinas. The indole-producing strains, including the strains of S. hyodysenteriae and some weakly beta-hemolytic Serpulina strains, formed one cluster. A second cluster comprised weakly beta-hemolytic strains that showed beta-galactosidase activity but lacked indole production and hippurate-hydrolyzing capacity. The second cluster contained two subclusters with similar phenotypic profiles. A third cluster involved strains that possessed a hippurate-hydrolyzing capacity which was distinct from that of the former two clusters, because of 17 unique nucleotide positions of the 16S rRNA gene. Interestingly, the strains of this third cluster were found likely to have a 16S rRNA structure in the V2 region of the molecule different from that of the serpulinas belonging to the other clusters. As a consequence of these findings, we propose that the intestinal spirochetes of this phenotype (i.e., P43/6/78-like strains) should be regarded as a separate Serpulina species. Furthermore, this cluster was found to be by far the most homogeneous one. In conclusion, the biochemical classification of porcine intestinal spirochetes was comparable to that by phylogenetic analysis based on 16S rRNA

  4. Prokaryotic nucleotide excision repair.

    PubMed

    Kisker, Caroline; Kuper, Jochen; Van Houten, Bennett

    2013-03-01

    Nucleotide excision repair (NER) has allowed bacteria to flourish in many different niches around the globe that inflict harsh environmental damage to their genetic material. NER is remarkable because of its diverse substrate repertoire, which differs greatly in chemical composition and structure. Recent advances in structural biology and single-molecule studies have given great insight into the structure and function of NER components. This ensemble of proteins orchestrates faithful removal of toxic DNA lesions through a multistep process. The damaged nucleotide is recognized by dynamic probing of the DNA structure that is then verified and marked for dual incisions followed by excision of the damage and surrounding nucleotides. The opposite DNA strand serves as a template for repair, which is completed after resynthesis and ligation. PMID:23457260

  5. The nucleotide sequence of Beneckea harveyi 5S rRNA. [bioluminescent marine bacterium

    NASA Technical Reports Server (NTRS)

    Luehrsen, K. R.; Fox, G. E.

    1981-01-01

    The primary sequence of the 5S ribosomal RNA isolated from the free-living bioluminescent marine bacterium Beneckea harveyi is reported and discussed in regard to indications of phylogenetic relationships with the bacteria Escherichia coli and Photobacterium phosphoreum. Sequences were determined for oligonucleotide products generated by digestion with ribonuclease T1, pancreatic ribonuclease and ribonuclease T2. The presence of heterogeneity is indicated for two sites. The B. harveyi sequence can be arranged into the same four helix secondary structures as E. coli and other prokaryotic 5S rRNAs. Examination of the 5S-RNS sequences of the three bacteria indicates that B. harveyi and P. phosphoreum are specifically related and share a common ancestor which diverged from an ancestor of E. coli at a somewhat earlier time, consistent with previous studies.

  6. Control by Phytochrome of Cytoplasmic Precursor rRNA Synthesis in the Cotyledons of Mustard Seedlings 1

    PubMed Central

    Thien, Wilfried; Schopfer, Peter

    1982-01-01

    The influence of phytochrome (high irradiance reaction; operationally, continuous far red light) on the incorporation of [3H]uridine into the cytoplasmic 2.5 megadalton precursor rRNA in the cotyledons of mustard (Sinapis alba L.) seedlings has been investigated. After irradiating 36-hour-old etiolated seedlings with 12 hours of far red light, the rate of incorporation is stimulated about 2-fold, leading to 50% labeling of the precursor rRNA pool about 15 minutes after the tracer has reached the nucleotide precursor pool. In the dark control, there is a significantly smaller pool of precursor rRNA which is half-saturated with label only after about 27 minutes. Since neither the specific radioactivity of the UTP pool nor the processing of the precursor rRNA demonstrate a corresponding light-dependent change, it is concluded that phytochrome mediates an increase of the transcription of the rRNA genes. This gene activation accounts for the increased accumulation of mature cytoplasmic rRNA during the course of photomorphogenesis of the cotyledons. PMID:16662362

  7. Renibacterium salmoninarum isolates from different sources possess two highly conserved copies of the rRNA operon .

    PubMed

    Grayson, T H; Alexander, S M; Cooper, L F; Gilpin, M L

    2000-07-01

    The nucleotide sequences of the rRNA genes and the 5' flanking region were determined for R. salmoninarum ATCC 33209T from overlapping products generated by PCR amplification from the genomic DNA. Comparison of the sequences with rRNA genes from a variety of bacteria demonstrated the close relatedness between R. salmoninarum and the high G+C group of the actinobacteria, in particular, Arthrobacter species. A regulatory element within the 5' leader of the rRNA operon was identical to an element, CL2, described for mycobacteria. PCR, DNA sequence analysis, and DNA hybridisation were performed to examine variation between isolates from diverse sources which represented the four 16S-23S rRNA intergenic spacer sequevars previously described for R. salmoninarum. Two 23S-5S rRNA intergenic spacer sequevars of identical length were found. DNA hybridisation using probes complementary to 23S rDNA and 16S rDNA identified two rRNA operons which were identical or nearly identical amongst 40 isolates sourced from a variety of countries. PMID:11016696

  8. Phylogenetic analysis of the genera Thiobacillus and Thiomicrospira by 5S rRNA sequences.

    PubMed Central

    Lane, D J; Stahl, D A; Olsen, G J; Heller, D J; Pace, N R

    1985-01-01

    5S rRNA nucleotide sequences from Thiobacillus neapolitanus, Thiobacillus ferrooxidans, Thiobacillus thiooxidans, Thiobacillus intermedius, Thiobacillus perometabolis, Thiobacillus thioparus, Thiobacillus versutus, Thiobacillus novellus, Thiobacillus acidophilus, Thiomicrospira pelophila, Thiomicrospira sp. strain L-12, and Acidiphilium cryptum were determined. A phylogenetic tree, based upon comparison of these and other related 5S rRNA sequences, is presented. The results place the thiobacilli, Thiomicrospira spp., and Acidiphilium spp. in the "purple photosynthetic" bacterial grouping which also includes the enteric, vibrio, pseudomonad, and other familiar eubacterial groups in addition to the purple photosynthetic bacteria. The genus Thiobacillus is not an evolutionarily coherent grouping but rather spans the full breadth of the purple photosynthetic bacteria. PMID:3924899

  9. Nucleotide diversity in gorillas.

    PubMed Central

    Yu, Ning; Jensen-Seaman, Michael I; Chemnick, Leona; Ryder, Oliver; Li, Wen-Hsiung

    2004-01-01

    Comparison of the levels of nucleotide diversity in humans and apes may provide valuable information for inferring the demographic history of these species, the effect of social structure on genetic diversity, patterns of past migration, and signatures of past selection events. Previous DNA sequence data from both the mitochondrial and the nuclear genomes suggested a much higher level of nucleotide diversity in the African apes than in humans. Noting that the nuclear DNA data from the apes were very limited, we previously conducted a DNA polymorphism study in humans and another in chimpanzees and bonobos, using 50 DNA segments randomly chosen from the noncoding, nonrepetitive parts of the human genome. The data revealed that the nucleotide diversity (pi) in bonobos (0.077%) is actually lower than that in humans (0.087%) and that pi in chimpanzees (0.134%) is only 50% higher than that in humans. In the present study we sequenced the same 50 segments in 15 western lowland gorillas and estimated pi to be 0.158%. This is the highest value among the African apes but is only about two times higher than that in humans. Interestingly, available mtDNA sequence data also suggest a twofold higher nucleotide diversity in gorillas than in humans, but suggest a threefold higher nucleotide diversity in chimpanzees than in humans. The higher mtDNA diversity in chimpanzees might be due to the unique pattern in the evolution of chimpanzee mtDNA. From the nuclear DNA pi values, we estimated that the long-term effective population sizes of humans, bonobos, chimpanzees, and gorillas are, respectively, 10,400, 12,300, 21,300, and 25,200. PMID:15082556

  10. The human 18S rRNA base methyltransferases DIMT1L and WBSCR22-TRMT112 but not rRNA modification are required for ribosome biogenesis

    PubMed Central

    Zorbas, Christiane; Nicolas, Emilien; Wacheul, Ludivine; Huvelle, Emmeline; Heurgué-Hamard, Valérie; Lafontaine, Denis L. J.

    2015-01-01

    At the heart of the ribosome lie rRNAs, whose catalytic function in translation is subtly modulated by posttranscriptional modifications. In the small ribosomal subunit of budding yeast, on the 18S rRNA, two adjacent adenosines (A1781/A1782) are N6-dimethylated by Dim1 near the decoding site, and one guanosine (G1575) is N7-methylated by Bud23-Trm112 at a ridge between the P- and E-site tRNAs. Here we establish human DIMT1L and WBSCR22-TRMT112 as the functional homologues of yeast Dim1 and Bud23-Trm112. We report that these enzymes are required for distinct pre-rRNA processing reactions leading to synthesis of 18S rRNA, and we demonstrate that in human cells, as in budding yeast, ribosome biogenesis requires the presence of the modification enzyme rather than its RNA-modifying catalytic activity. We conclude that a quality control mechanism has been conserved from yeast to human by which binding of a methyltransferase to nascent pre-rRNAs is a prerequisite to processing, so that all cleaved RNAs are committed to faithful modification. We further report that 18S rRNA dimethylation is nuclear in human cells, in contrast to yeast, where it is cytoplasmic. Yeast and human ribosome biogenesis thus have both conserved and distinctive features. PMID:25851604

  11. Variable rRNA gene copies in extreme halobacteria

    SciTech Connect

    Sanz, J.L.; Marin, I.; Ramirez, L.; Amils, R. ); Abad, J.P.; Smith, C.L. )

    1988-08-25

    Using PFG electrophoresis techniques, the authors have examined the organization of rRNA gene in halobacterium species. The results show that the organization of rRNA genes among closely related halobacteria is quite heterogeneous. This contrasts with the high degree of conservation of rRNA sequence. The possible mechanism of such rRNA gene amplification and its evolutionary implications are discussed.

  12. Evolution of functional six-nucleotide DNA.

    PubMed

    Zhang, Liqin; Yang, Zunyi; Sefah, Kwame; Bradley, Kevin M; Hoshika, Shuichi; Kim, Myong-Jung; Kim, Hyo-Joong; Zhu, Guizhi; Jiménez, Elizabeth; Cansiz, Sena; Teng, I-Ting; Champanhac, Carole; McLendon, Christopher; Liu, Chen; Zhang, Wen; Gerloff, Dietlind L; Huang, Zhen; Tan, Weihong; Benner, Steven A

    2015-06-01

    Axiomatically, the density of information stored in DNA, with just four nucleotides (GACT), is higher than in a binary code, but less than it might be if synthetic biologists succeed in adding independently replicating nucleotides to genetic systems. Such addition could also add functional groups not found in natural DNA, but useful for molecular performance. Here, we consider two new nucleotides (Z and P, 6-amino-5-nitro-3-(1'-β-D-2'-deoxyribo-furanosyl)-2(1H)-pyridone and 2-amino-8-(1'-β-D-2'-deoxyribofuranosyl)-imidazo[1,2-a]-1,3,5-triazin-4(8H)-one). These are designed to pair via complete Watson-Crick geometry. These were added to a library of oligonucleotides used in a laboratory in vitro evolution (LIVE) experiment; the GACTZP library was challenged to deliver molecules that bind selectively to liver cancer cells, but not to untransformed liver cells. Unlike in classical in vitro selection, low levels of mutation allow this system to evolve to create binding molecules not necessarily present in the original library. Over a dozen binding species were recovered. The best had Z and/or P in their sequences. Several had multiple, nearby, and adjacent Zs and Ps. Only the weaker binders contained no Z or P at all. This suggests that this system explored much of the sequence space available to this genetic system and that GACTZP libraries are richer reservoirs of functionality than standard libraries. PMID:25966323

  13. Nucleotide cleaving agents and method

    DOEpatents

    Que, Jr., Lawrence; Hanson, Richard S.; Schnaith, Leah M. T.

    2000-01-01

    The present invention provides a unique series of nucleotide cleaving agents and a method for cleaving a nucleotide sequence, whether single-stranded or double-stranded DNA or RNA, using and a cationic metal complex having at least one polydentate ligand to cleave the nucleotide sequence phosphate backbone to yield a hydroxyl end and a phosphate end.

  14. Modified nucleotides in T1 RNase oligonucleotides of 18S ribosomal RNA of the Novikoff hepatoma.

    PubMed

    Choi, Y C; Busch, H

    1978-06-27

    The primary structure of 18S rRNA of the Novikoff hepatoma cells was investigated. Regardless of whether the primary sequence of 18S rRNA is finally determined by RNA sequencing methods or DNA sequencing methods, it is important to identify numbers and types of the modified nucleotides and accordingly the present study was designed to localize the modified regions in T1 RNase derived oligonucleotide. Modified nucleotides found in 66 different oligonucleotide sequences included 2 m62A, 1 m6A, 1 m7G, 1m1cap3psi, 7 Cm, 13 Am, 9 Gm, 11 Um, and 38 psi residues. A number of these modified nucleotides are now placed in defined sequences of T1 RNase oligonucleotides which are now being searched for in larger fragments derived from partial T1 RNase digests of 18S rRNA. Improved homochromatography fingerprinting (Choi et al. (1976) Cancer Res. 36, 4301) of T1 RNase derived oligonucleotides provided a distinctive pattern for 18S rRNA of Novikoff hepatoma ascites cells. The 116 spots obtained by homochromatography contain 176 oligonucleotide sequences. PMID:209819

  15. Nucleotide sequence of an exceptionally long 5.8S ribosomal RNA from Crithidia fasciculata.

    PubMed

    Schnare, M N; Gray, M W

    1982-03-25

    In Crithidia fasciculata, a trypanosomatid protozoan, the large ribosomal subunit contains five small RNA species (e, f, g, i, j) in addition to 5S rRNA [Gray, M.W. (1981) Mol. Cell. Biol. 1, 347-357]. The complete primary sequence of species i is shown here to be pAACGUGUmCGCGAUGGAUGACUUGGCUUCCUAUCUCGUUGA ... AGAmACGCAGUAAAGUGCGAUAAGUGGUApsiCAAUUGmCAGAAUCAUUCAAUUACCGAAUCUUUGAACGAAACGG ... CGCAUGGGAGAAGCUCUUUUGAGUCAUCCCCGUGCAUGCCAUAUUCUCCAmGUGUCGAA(C)OH. This sequence establishes that species i is a 5.8S rRNA, despite its exceptional length (171-172 nucleotides). The extra nucleotides in C. fasciculata 5.8S rRNA are located in a region whose primary sequence and length are highly variable among 5.8S rRNAs, but which is capable of forming a stable hairpin loop structure (the "G+C-rich hairpin"). The sequence of C. fasciculata 5.8S rRNA is no more closely related to that of another protozoan, Acanthamoeba castellanii, than it is to representative 5.8S rRNA sequences from the other eukaryotic kingdoms, emphasizing the deep phylogenetic divisions that seem to exist within the Kingdom Protista. PMID:7079176

  16. 46 CFR 148.445 - Adjacent spaces.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 46 Shipping 5 2013-10-01 2013-10-01 false Adjacent spaces. 148.445 Section 148.445 Shipping COAST... THAT REQUIRE SPECIAL HANDLING Additional Special Requirements § 148.445 Adjacent spaces. When... following requirements must be met: (a) Each space adjacent to a cargo hold must be ventilated by...

  17. 46 CFR 148.445 - Adjacent spaces.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 46 Shipping 5 2014-10-01 2014-10-01 false Adjacent spaces. 148.445 Section 148.445 Shipping COAST... THAT REQUIRE SPECIAL HANDLING Additional Special Requirements § 148.445 Adjacent spaces. When... following requirements must be met: (a) Each space adjacent to a cargo hold must be ventilated by...

  18. 46 CFR 148.445 - Adjacent spaces.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... 46 Shipping 5 2012-10-01 2012-10-01 false Adjacent spaces. 148.445 Section 148.445 Shipping COAST... THAT REQUIRE SPECIAL HANDLING Additional Special Requirements § 148.445 Adjacent spaces. When... following requirements must be met: (a) Each space adjacent to a cargo hold must be ventilated by...

  19. 46 CFR 148.445 - Adjacent spaces.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 46 Shipping 5 2011-10-01 2011-10-01 false Adjacent spaces. 148.445 Section 148.445 Shipping COAST... THAT REQUIRE SPECIAL HANDLING Additional Special Requirements § 148.445 Adjacent spaces. When... following requirements must be met: (a) Each space adjacent to a cargo hold must be ventilated by...

  20. Molecular systematics of hystricognath rodents: evidence from the mitochondrial 12S rRNA gene.

    PubMed

    Nedbal, M A; Allard, M W; Honeycutt, R L

    1994-09-01

    Nucleotide sequence variation among 22 representatives of 14 families of hystricognathid rodents was examined using an 814-bp region of the mitochondrial 12S ribosomal RNA (rRNA) gene composing domains I-III. The purpose of this study was twofold. First, the phylogenetic relationships among Old World phiomorph (primarily African) and New World caviomorph (primarily South American) families were investigated, with a special emphasis on testing hypotheses pertaining to the origin of New World families and the identification of major monophyletic groups. Second, divergence times derived from molecular data were compared to those suggested by the fossil record. The resultant 12S rRNA gene phylogeny, analyzed separately and in combination with other morphological and molecular data, supported a monophyletic Caviomorpha. This finding is counter to the idea of a multiple origin for the South American families. The most strongly supported relationships within the Caviomorpha were a monophyletic Octodontoidea (containing five families) and the placement of New World porcupines (family Erethizontidae) as the most divergent family. Although comparisons to other data were more equivocal, the most parsimonious 12S rRNA trees also supported a monophyletic Phiomorpha that could be subdivided into two major groups, a clade containing the Thryonomyoidea (Thryonomyidae and Petromuridae) plus Bathyergidae and the more divergent Hystricidae (Old World porcupines). No significant differences in rates of 12S rRNA gene divergence were observed for hystricognathids in comparison to other rodent groups. Although time since divergence estimates were influenced by the fossil dates chosen to calibrate absolute rates, the overall divergence times derived from both transversions only and Kimura corrected distances and calibrations using two independent dates revealed a divergence time between Old and New World groups dating in the Eocene. PMID:7820285

  1. Methylation sites in Escherichia coli ribosomal RNA: localization and identification of four new sites of methylation in 23S rRNA.

    PubMed

    Smith, J E; Cooperman, B S; Mitchell, P

    1992-11-10

    Four previously undetermined sites of methylation are mapped in Escherichia coli 23S rRNA employing a novel combination of methods. First, using a double-isotope approach, the total number of methyl groups in 23S rRNA was determined to be 14.9 +/- 1.6. Second, hybridization of methyl-labeled rRNA to complementary DNA restriction fragments and PAGE analysis were used to purify RNA-DNA heteroduplexes and to quantify methyl groups within specific 23S rRNA fragments. Third, the methylated nucleosides in these fragments were identified and quantified using HPLC, confirming the presence of 14 methylation sites in 23S rRNA, four more than had been previously identified. In contrast, a similar set of analyses conducted on 16S rRNA gave evidence for 10 sites of methylation, at all approximate locations consistent with published 16S methylated nucleoside identities and locations. Selected regions of the 23S rRNA molecule containing previously unidentified methylated nucleosides were released by site-directed cleavage with ribonuclease H and isolated by PAGE. Sites of methylation within the RNA fragments were determined by classical oligonucleotide analyses. The four newly identified methylation sites in 23S rRNA are m2G-1835, m5C-1962, m6A-2503, and m2G at one of positions 2445-2447. Together with previously described sites of modification, these new sites form a group that is clustered in a current model for the three-dimensional organization of the 23S rRNA in the 50S ribosomal subunit, at a locus congruent with nucleotides previously implicated in ribosomal function. PMID:1384701

  2. Dietary nucleotides protect against alcoholic liver injury by attenuating inflammation and regulating gut microbiota in rats.

    PubMed

    Cai, Xiaxia; Bao, Lei; Wang, Nan; Ren, Jinwei; Chen, Qihe; Xu, Meihong; Li, Di; Mao, Ruixue; Li, Yong

    2016-06-15

    Nucleotides have been reported to be effective in attenuating liver damage and regulating gut microbiota. However, the protective effect of nucleotides against alcoholic liver injury remains unknown. The present study aims to investigate whether nucleotides ameliorate alcoholic liver injury and explores the possible mechanism. Male Wistar rats were given alcohol, equivalent distilled water or an isocaloric amount of dextrose intragastrically twice daily for up to 6 weeks respectively. Two subgroups of alcohol-treated rats were fed with a nucleotide-supplemented AIN-93G rodent diet. Serum enzymes, inflammatory cytokines and microbiota composition of the caecum content were evaluated. We found that nucleotides could significantly decrease serum alanine aminotransferase and aspartate aminotransferase, plasma lipopolysaccharide and inflammatory cytokine levels. Sequencing of 16S rRNA genes revealed that nucleotide-treated rats showed a higher abundance of Firmicutes and a lower abundance of Bacteroidetes than alcohol-treated rats. Moreover, nucleotide treatment inhibited the protein expression of toll-like receptor 4, CD14 and repressed the phosphorylation of inhibitor kappa Bα and nuclear factor-κB p65 in the liver. These results suggested that nucleotides suppressed the inflammatory response and regulated gut microbiota in alcoholic liver injury. The partial inhibition of lipopolysaccharide - toll-like receptor 4-nuclear factor-κB p65 signaling in the liver may be attributed to this mechanism. PMID:27247978

  3. Evolution of Functional Six-Nucleotide DNA

    PubMed Central

    Zhang, Liqin; Yang, Zunyi; Sefah, Kwame; Bradley, Kevin M.; Hoshika, Shuichi; Kim, Myong-Jung; Kim, Hyo-Joong; Zhu1, Guizhi; Jiménez, Elizabeth; Cansiz, Sena; Teng, I-Ting; Champanhac, Carole; McLendon, Christopher; Liu, Chen; Zhang, Wen; Gerloff, Dietlind L.

    2015-01-01

    Axiomatically, the density of information stored in DNA, with just four nucleotides (GACT), is higher than in a binary code, but less than it might be if synthetic biologists succeed in adding independently replicating nucleotides to genetic systems. Such addition could also add additional functional groups, not found in natural DNA but useful for molecular performance. Here, we consider two new nucleotides (Z and P, 6-amino-5-nitro-3-(1′-β-D-2′-deoxyribo-furanosyl)-2(1H)-pyridone and 2-amino-8-(1′-β-D-2′-deoxyribofuranosyl)-imidazo[1,2-a]-1,3,5-triazin-4(8H)-one). These are designed to pair via strict Watson-Crick geometry. These were added to lies in a ibrarlaboratory in vitro evolution (LIVE) experiment; the GACTZP library was challenged to deliver molecules that bind selectively to liver cancer cells, but not to untransformed liver cells. Unlike in classical in vitro selection systems, low levels of mutation allow this system to evolve to create binding molecules not necessarily present in the original library. Over a dozen binding species were recovered. The best had Z and/or P in their sequences. Several had multiple, nearby, and adjacent Z’s and P’s. Only the weaker binders contained no Z or P at all. This suggests that this system explored much of the sequence space available to this genetic system, and that GACTZP libraries are richer reservoir of functionality than standard libraries. PMID:25966323

  4. International interlaboratory study comparing single organism 16S rRNA gene sequencing data: Beyond consensus sequence comparisons.

    PubMed

    Olson, Nathan D; Lund, Steven P; Zook, Justin M; Rojas-Cornejo, Fabiola; Beck, Brian; Foy, Carole; Huggett, Jim; Whale, Alexandra S; Sui, Zhiwei; Baoutina, Anna; Dobeson, Michael; Partis, Lina; Morrow, Jayne B

    2015-03-01

    This study presents the results from an interlaboratory sequencing study for which we developed a novel high-resolution method for comparing data from different sequencing platforms for a multi-copy, paralogous gene. The combination of PCR amplification and 16S ribosomal RNA gene (16S rRNA) sequencing has revolutionized bacteriology by enabling rapid identification, frequently without the need for culture. To assess variability between laboratories in sequencing 16S rRNA, six laboratories sequenced the gene encoding the 16S rRNA from Escherichia coli O157:H7 strain EDL933 and Listeria monocytogenes serovar 4b strain NCTC11994. Participants performed sequencing methods and protocols available in their laboratories: Sanger sequencing, Roche 454 pyrosequencing(®), or Ion Torrent PGM(®). The sequencing data were evaluated on three levels: (1) identity of biologically conserved position, (2) ratio of 16S rRNA gene copies featuring identified variants, and (3) the collection of variant combinations in a set of 16S rRNA gene copies. The same set of biologically conserved positions was identified for each sequencing method. Analytical methods using Bayesian and maximum likelihood statistics were developed to estimate variant copy ratios, which describe the ratio of nucleotides at each identified biologically variable position, as well as the likely set of variant combinations present in 16S rRNA gene copies. Our results indicate that estimated variant copy ratios at biologically variable positions were only reproducible for high throughput sequencing methods. Furthermore, the likely variant combination set was only reproducible with increased sequencing depth and longer read lengths. We also demonstrate novel methods for evaluating variable positions when comparing multi-copy gene sequence data from multiple laboratories generated using multiple sequencing technologies. PMID:27077030

  5. Intra-Genomic Heterogeneity in 16S rRNA Genes in Strictly Anaerobic Clinical Isolates from Periodontal Abscesses

    PubMed Central

    Chen, Jiazhen; Miao, Xinyu; Xu, Meng; He, Junlin; Xie, Yi; Wu, Xingwen; Chen, Gang; Yu, Liying; Zhang, Wenhong

    2015-01-01

    Background Members of the genera Prevotella, Veillonella and Fusobacterium are the predominant culturable obligate anaerobic bacteria isolated from periodontal abscesses. When determining the cumulative number of clinical anaerobic isolates from periodontal abscesses, ambiguous or overlapping signals were frequently encountered in 16S rRNA gene sequencing chromatograms, resulting in ambiguous identifications. With the exception of the genus Veillonella, the high intra-chromosomal heterogeneity of rrs genes has not been reported. Methods The 16S rRNA genes of 138 clinical, strictly anaerobic isolates and one reference strain were directly sequenced, and the chromatograms were carefully examined. Gene cloning was performed for 22 typical isolates with doublet sequencing signals for the 16S rRNA genes, and four copies of the rrs-ITS genes of 9 Prevotella intermedia isolates were separately amplified by PCR, sequenced and compared. Five conserved housekeeping genes, hsp60, recA, dnaJ, gyrB1 and rpoB from 89 clinical isolates of Prevotella were also amplified by PCR and sequenced for identification and phylogenetic analysis along with 18 Prevotella reference strains. Results Heterogeneity of 16S rRNA genes was apparent in clinical, strictly anaerobic oral bacteria, particularly in the genera Prevotella and Veillonella. One hundred out of 138 anaerobic strains (72%) had intragenomic nucleotide polymorphisms (SNPs) in multiple locations, and 13 strains (9.4%) had intragenomic insertions or deletions in the 16S rRNA gene. In the genera Prevotella and Veillonella, 75% (67/89) and 100% (19/19) of the strains had SNPs in the 16S rRNA gene, respectively. Gene cloning and separate amplifications of four copies of the rrs-ITS genes confirmed that 2 to 4 heterogeneous 16S rRNA copies existed. Conclusion Sequence alignment of five housekeeping genes revealed that intra-species nucleotide similarities were very high in the genera Prevotella, ranging from 94.3–100%. However, the

  6. Functional importance of individual rRNA 2′-O-ribose methylations revealed by high-resolution phenotyping

    PubMed Central

    Esguerra, Jonathan; Warringer, Jonas; Blomberg, Anders

    2008-01-01

    Ribosomal RNAs contain numerous modifications at specific nucleotides. Despite their evolutionary conservation, the functional role of individual 2′-O-ribose methylations in rRNA is not known. A distinct family of small nucleolar RNAs, box C/D snoRNAs, guides the methylating complex to specific rRNA sites. Using a high-resolution phenotyping approach, we characterized 20 box C/D snoRNA gene deletions for altered growth dynamics under a wide array of environmental perturbations, encompassing intraribosomal antibiotics, inhibitors of specific cellular features, as well as general stressors. Ribosome-specific antibiotics generated phenotypes indicating different and long-ranging structural effects of rRNA methylations on the ribosome. For all studied box C/D snoRNA mutants we uncovered phenotypes to extraribosomal growth inhibitors, most frequently reflected in alteration in growth lag (adaptation time). A number of strains were highly pleiotropic and displayed a great number of sensitive phenotypes, e.g., deletion mutants of snR70 and snR71, which both have clear human homologues, and deletion mutants of snR65 and snR68. Our data indicate that individual rRNA ribose methylations can play either distinct or general roles in the workings of the ribosome. PMID:18256246

  7. Nucleotide sequence of 5S ribosomal RNA from Aspergillus nidulans and Neurospora crassa.

    PubMed Central

    Piechulla, B; Hahn, U; McLaughlin, L W; Küntzel, H

    1981-01-01

    The nucleotide sequences of 5S rRNA molecules isolated from the cytosol and the mitochondria of the ascomycetes A. nidulans and N. crassa were determined by partial chemical cleavage of 3'-terminally labelled RNA. The sequence identity of the cytosolic and mitochondrial RNA preparations confirms the absence of mitochondrion-specific 5S rRNA in these fungi. The sequences of the two organisms differ in 35 positions, and each sequence differs from yeast 5S rRNA in 44 positions. Both molecules contain the sequence GCUC in place of GAAC or GAUY found in all other 5S rRNAs, indicating that this region is not universally involved in base-pairing to the invariant GTpsiC sequence of tRNAs. Images PMID:6453331

  8. Identification of new 18S rRNA strains of Babesia canis isolated from dogs with subclinical babesiosis.

    PubMed

    Łyp, P; Adaszek, Ł; Furmaga, B; Winiarczyk, S

    2015-01-01

    In this study, we used PCR to detect and characterize B. canis from naturally infected dogs in Poland with subclinical babesiosis by amplifying and sequencing a portion of the 18S ribosomal RNA (rRNA) gene. Venous blood samples were collected from ten dogs with subclinical babesiosis. A 559-bp fragment of the B. canis 18S rRNA gene was amplified by PCR. Sequencing of the PCR products led to the identification of a new variant of Babesia canis, differing from the previously detected protozoa genotypes (18S rRNA-A and 18S rRNA-B) with nucleotide substitutions in positions 150 and 151 of the tested gene fragment. The results indicate the emergence within the Polish territory of a new, previously unencountered Babesia canis genotype responsible for the development of subclinical babesiosis. PMID:26618590

  9. Evolution of multicellular animals as deduced from 5S rRNA sequences: a possible early emergence of the Mesozoa.

    PubMed

    Ohama, T; Kumazaki, T; Hori, H; Osawa, S

    1984-06-25

    The nucleotide sequences of 5S rRNA from a mesozoan Dicyema misakiense and three metazoan species, i.e., an acorn-worm Saccoglossus kowalevskii, a moss-animal Bugula neritina, and an octopus Octopus vulgaris have been determined. A phylogenic tree of multicellular animals has been constructed from 73 5S rRNA sequences available at present including those from the above four sequences. The tree suggests that the mesozoan is the most ancient multicellular animal identified so far, its emergence time being almost the same as that of flagellated or ciliated protozoans. The branching points of planarians and nematodes are a little later than that of the mesozoan but are clearly earlier than other metazoan groups including sponges and jellyfishes. Many metazoan groups seem to have diverged within a relatively short period. PMID:6539911

  10. Labeled nucleotide phosphate (NP) probes

    DOEpatents

    Korlach, Jonas; Webb, Watt W.; Levene, Michael; Turner, Stephen; Craighead, Harold G.; Foquet, Mathieu

    2009-02-03

    The present invention is directed to a method of sequencing a target nucleic acid molecule having a plurality of bases. In its principle, the temporal order of base additions during the polymerization reaction is measured on a molecule of nucleic acid, i.e. the activity of a nucleic acid polymerizing enzyme on the template nucleic acid molecule to be sequenced is followed in real time. The sequence is deduced by identifying which base is being incorporated into the growing complementary strand of the target nucleic acid by the catalytic activity of the nucleic acid polymerizing enzyme at each step in the sequence of base additions. A polymerase on the target nucleic acid molecule complex is provided in a position suitable to move along the target nucleic acid molecule and extend the oligonucleotide primer at an active site. A plurality of labelled types of nucleotide analogs are provided proximate to the active site, with each distinguishable type of nucleotide analog being complementary to a different nucleotide in the target nucleic acid sequence. The growing nucleic acid strand is extended by using the polymerase to add a nucleotide analog to the nucleic acid strand at the active site, where the nucleotide analog being added is complementary to the nucleotide of the target nucleic acid at the active site. The nucleotide analog added to the oligonucleotide primer as a result of the polymerizing step is identified. The steps of providing labelled nucleotide analogs, polymerizing the growing nucleic acid strand, and identifying the added nucleotide analog are repeated so that the nucleic acid strand is further extended and the sequence of the target nucleic acid is determined.

  11. The complete nucleotide sequence of a 16S ribosomal RNA gene from a blue-green alga, Anacystis nidulans.

    PubMed

    Tomioka, N; Sugiura, M

    1983-01-01

    The complete nucleotide sequence of a 16S ribosomal RNA gene from a blue-green alga, Anacystis nidulans, has been determined. Its coding region is estimated to be 1,487 base pairs long, which is nearly identical to those reported for chloroplast 16S rRNA genes and is about 4% shorter than that of the Escherichia coli gene. The 16S rRNA sequence of A. nidulans has 83% homology with that of tobacco chloroplast and 74% homology with that of E. coli. Possible stem and loop structures of A. nidulans 16S rRNA sequences resemble more closely those of chloroplast 16S rRNAs than those of E. coli 16S rRNA. These observations support the endosymbiotic theory of chloroplast origin. PMID:6412038

  12. The Identification of Discriminating Patterns from 16S rRNA Gene to Generate Signature for Bacillus Genus.

    PubMed

    More, Ravi P; Purohit, Hemant J

    2016-08-01

    The 16S ribosomal RNA (16S rRNA) gene has been widely used for the taxonomic classification of bacteria. A molecular signature is a set of nucleotide patterns, which constitute a regular expression that is specific to each particular taxon. Our main goal was to identify discriminating nucleotide patterns in 16S rRNA gene and then to generate signatures for taxonomic classification. To demonstrate our approach, we used the phylum Firmicutes as a model using representative taxa Bacilli (class), Bacillales (order), Bacillaceae (family), and Bacillus (genus), according to their dominance at each hierarchical taxonomic level. We applied combined composite vector and multiple sequence alignment approaches to generate gene-specific signatures. Further, we mapped all the patterns into the different hypervariable regions of 16S rRNA gene and confirmed the most appropriate distinguishing region as V3-V4 for targeted taxa. We also examined the evolution in discriminating patterns of signatures across taxonomic levels. We assessed the comparative classification accuracy of signatures with other methods (i.e., RDP Classifier, KNN, and SINA). Results revealed that the signatures for taxa Bacilli, Bacillales, Bacillaceae, and Bacillus could correctly classify isolate sequences with sensitivity of 0.99, 0.97, 0.94, and 0.89, respectively, and specificity close to 0.99. We developed signature-based software DNA Barcode Identification (DNA BarID) for taxonomic classification that is available at website http://www.neeri.res.in/DNA_BarID.htm . This pattern-based study provides a deeper understanding of taxon-specific discriminating patterns in 16S rRNA gene with respect to taxonomic classification. PMID:27104769

  13. Recent Trends in Nucleotide Synthesis.

    PubMed

    Roy, Béatrice; Depaix, Anaïs; Périgaud, Christian; Peyrottes, Suzanne

    2016-07-27

    Focusing on the recent literature (since 2000), this review outlines the main synthetic approaches for the preparation of 5'-mono-, 5'-di-, and 5'-triphosphorylated nucleosides, also known as nucleotides, as well as several derivatives, namely, cyclic nucleotides and dinucleotides, dinucleoside 5',5'-polyphosphates, sugar nucleotides, and nucleolipids. Endogenous nucleotides and their analogues can be obtained enzymatically, which is often restricted to natural substrates, or chemically. In chemical synthesis, protected or unprotected nucleosides can be used as the starting material, depending on the nature of the reagents selected from P(III) or P(V) species. Both solution-phase and solid-support syntheses have been developed and are reported here. Although a considerable amount of research has been conducted in this field, further work is required because chemists are still faced with the challenge of developing a universal methodology that is compatible with a large variety of nucleoside analogues. PMID:27319940

  14. Discrimination of Bacillus anthracis from closely related microorganisms by analysis of 16S and 23S rRNA with oligonucleotide microchips

    DOEpatents

    Bavykin, Sergei G.; Mirzabekova, legal representative, Natalia V.; Mirzabekov, deceased, Andrei D.

    2007-12-04

    The present invention relates to methods and compositions for using nucleotide sequence variations of 16S and 23S rRNA within the B. cereus group to discriminate a highly infectious bacterium B. anthracis from closely related microorganisms. Sequence variations in the 16S and 23S rRNA of the B. cereus subgroup including B. anthracis are utilized to construct an array that can detect these sequence variations through selective hybridizations and discriminate B. cereus group that includes B. anthracis. Discrimination of single base differences in rRNA was achieved with a microchip during analysis of B. cereus group isolates from both single and in mixed samples, as well as identification of polymorphic sites. Successful use of a microchip to determine the appropriate subgroup classification using eight reference microorganisms from the B. cereus group as a study set, was demonstrated.

  15. Phylogenetic tree derived from bacterial, cytosol and organelle 5S rRNA sequences.

    PubMed Central

    Küntzel, H; Heidrich, M; Piechulla, B

    1981-01-01

    A phylogenetic tree was constructed by computer analysis of 47 completely determined 5S rRNA sequences. The wheat mitochondrial sequence is significantly more related to prokaryotic than to eukaryotic sequences, and its affinity to that of the thermophilic Gram-negative bacterium Thermus aquaticus is comparable to the affinity between Anacystis nidulans and chloroplastic sequences. This strongly supports the idea of an endosymbiotic origin of plant mitochondria. A comparison of the plant cytosol and chloroplast sub-trees suggests a similar rate of nucleotide substitution in nuclear genes and chloroplastic genes. Other features of the tree are a common precursor of protozoa and metazoa, which appears to be more related to the fungal than to the plant protosequence, and an early divergence of the archebacterial sequence (Halobacterium cutirubrum) from the prokaryotic branch. PMID:6785727

  16. 43 CFR 420.3 - Adjacent lands.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 43 Public Lands: Interior 1 2014-10-01 2014-10-01 false Adjacent lands. 420.3 Section 420.3 Public Lands: Interior Regulations Relating to Public Lands BUREAU OF RECLAMATION, DEPARTMENT OF THE INTERIOR...-managing agencies on adjacent lands (both public and private)....

  17. 43 CFR 420.3 - Adjacent lands.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 43 Public Lands: Interior 1 2013-10-01 2013-10-01 false Adjacent lands. 420.3 Section 420.3 Public Lands: Interior Regulations Relating to Public Lands BUREAU OF RECLAMATION, DEPARTMENT OF THE INTERIOR...-managing agencies on adjacent lands (both public and private)....

  18. 43 CFR 420.3 - Adjacent lands.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 43 Public Lands: Interior 1 2011-10-01 2011-10-01 false Adjacent lands. 420.3 Section 420.3 Public Lands: Interior Regulations Relating to Public Lands BUREAU OF RECLAMATION, DEPARTMENT OF THE INTERIOR OFF-ROAD VEHICLE USE § 420.3 Adjacent lands. When administratively feasible, the regulation of off-road vehicle use on Reclamation lands will...

  19. 43 CFR 420.3 - Adjacent lands.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... 43 Public Lands: Interior 1 2012-10-01 2011-10-01 true Adjacent lands. 420.3 Section 420.3 Public Lands: Interior Regulations Relating to Public Lands BUREAU OF RECLAMATION, DEPARTMENT OF THE INTERIOR...-managing agencies on adjacent lands (both public and private)....

  20. 43 CFR 420.3 - Adjacent lands.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 43 Public Lands: Interior 1 2010-10-01 2010-10-01 false Adjacent lands. 420.3 Section 420.3 Public Lands: Interior Regulations Relating to Public Lands BUREAU OF RECLAMATION, DEPARTMENT OF THE INTERIOR OFF-ROAD VEHICLE USE § 420.3 Adjacent lands. When administratively feasible, the regulation of...

  1. Multi-site-specific 16S rRNA Methyltransferase RsmF from Thermus thermophilus

    SciTech Connect

    Demirci, H.; Larsen, L; Hansen, T; Rasmussen, A; Cadambi, A; Gregory, S; Kirpekar, F; Jogl, G

    2010-01-01

    Cells devote a significant effort toward the production of multiple modified nucleotides in rRNAs, which fine tune the ribosome function. Here, we report that two methyltransferases, RsmB and RsmF, are responsible for all four 5-methylcytidine (m{sup 5}C) modifications in 16S rRNA of Thermus thermophilus. Like Escherichia coli RsmB, T. thermophilus RsmB produces m{sup 5}C967. In contrast to E. coli RsmF, which introduces a single m{sup 5}C1407 modification, T. thermophilus RsmF modifies three positions, generating m{sup 5}C1400 and m{sup 5}C1404 in addition to m{sup 5}C1407. These three residues are clustered near the decoding site of the ribosome, but are situated in distinct structural contexts, suggesting a requirement for flexibility in the RsmF active site that is absent from the E. coli enzyme. Two of these residues, C1400 and C1404, are sufficiently buried in the mature ribosome structure so as to require extensive unfolding of the rRNA to be accessible to RsmF. In vitro, T. thermophilus RsmF methylates C1400, C1404, and C1407 in a 30S subunit substrate, but only C1400 and C1404 when naked 16S rRNA is the substrate. The multispecificity of T. thermophilus RsmF is potentially explained by three crystal structures of the enzyme in a complex with cofactor S-adenosyl-methionine at up to 1.3 {angstrom} resolution. In addition to confirming the overall structural similarity to E. coli RsmF, these structures also reveal that key segments in the active site are likely to be dynamic in solution, thereby expanding substrate recognition by T. thermophilus RsmF.

  2. Functional Specialization of Domains Tandemly Duplicated Witin 16S rRNA Methyltransferase RsmC

    SciTech Connect

    Sunita,S.; Purta, E.; Durawa, M.; Tkaczuk, K.; Swaathi, J.; Bujnicki, J.; Sivaraman, J.

    2007-01-01

    RNA methyltransferases (MTases) are important players in the biogenesis and regulation of the ribosome, the cellular machine for protein synthesis. RsmC is a MTase that catalyzes the transfer of a methyl group from S-adenosyl-L-methionine (SAM) to G1207 of 16S rRNA. Mutations of G1207 have dominant lethal phenotypes in Escherichia coli, underscoring the significance of this modified nucleotide for ribosome function. Here we report the crystal structure of E. coli RsmC refined to 2.1 Angstroms resolution, which reveals two homologous domains tandemly duplicated within a single polypeptide. We characterized the function of the individual domains and identified key residues involved in binding of rRNA and SAM, and in catalysis. We also discovered that one of the domains is important for the folding of the other. Domain duplication and subfunctionalization by complementary degeneration of redundant functions (in particular substrate binding versus catalysis) has been reported for many enzymes, including those involved in RNA metabolism. Thus, RsmC can be regarded as a model system for functional streamlining of domains accompanied by the development of dependencies concerning folding and stability.

  3. Detailed analysis of RNA-protein interactions within the bacterial ribosomal protein L5/5S rRNA complex.

    PubMed Central

    Perederina, Anna; Nevskaya, Natalia; Nikonov, Oleg; Nikulin, Alexei; Dumas, Philippe; Yao, Min; Tanaka, Isao; Garber, Maria; Gongadze, George; Nikonov, Stanislav

    2002-01-01

    The crystal structure of ribosomal protein L5 from Thermus thermophilus complexed with a 34-nt fragment comprising helix III and loop C of Escherichia coli 5S rRNA has been determined at 2.5 A resolution. The protein specifically interacts with the bulged nucleotides at the top of loop C of 5S rRNA. The rRNA and protein contact surfaces are strongly stabilized by intramolecular interactions. Charged and polar atoms forming the network of conserved intermolecular hydrogen bonds are located in two narrow planar parallel layers belonging to the protein and rRNA, respectively. The regions, including these atoms conserved in Bacteria and Archaea, can be considered an RNA-protein recognition module. Comparison of the T. thermophilus L5 structure in the RNA-bound form with the isolated Bacillus stearothermophilus L5 structure shows that the RNA-recognition module on the protein surface does not undergo significant changes upon RNA binding. In the crystal of the complex, the protein interacts with another RNA molecule in the asymmetric unit through the beta-sheet concave surface. This protein/RNA interface simulates the interaction of L5 with 23S rRNA observed in the Haloarcula marismortui 50S ribosomal subunit. PMID:12515387

  4. Compilation of 5S rRNA and 5S rRNA gene sequences

    PubMed Central

    Specht, Thomas; Wolters, Jörn; Erdmann, Volker A.

    1990-01-01

    The BERLIN RNA DATABANK as of Dezember 31, 1989, contains a total of 667 sequences of 5S rRNAs or their genes, which is an increase of 114 new sequence entries over the last compilation (1). It covers sequences from 44 archaebacteria, 267 eubacteria, 20 plastids, 6 mitochondria, 319 eukaryotes and 11 eukaryotic pseudogenes. The hardcopy shows only the list (Table 1) of those organisms whose sequences have been determined. The BERLIN RNA DATABANK uses the format of the EMBL Nucleotide Sequence Data Library complemented by a Sequence Alignment (SA) field including secondary structure information. PMID:1692116

  5. Structure of ERA in complex with the 3′ end of 16S rRNA: Implications for ribosome biogenesis

    SciTech Connect

    Tu, Chao; Zhou, Xiaomei; Tropea, Joseph E.; Austin, Brian P.; Waugh, David S.; Court, Donald L.; Ji, Xinhua

    2009-10-09

    ERA, composed of an N-terminal GTPase domain followed by an RNA-binding KH domain, is essential for bacterial cell viability. It binds to 16S rRNA and the 30S ribosomal subunit. However, its RNA-binding site, the functional relationship between the two domains, and its role in ribosome biogenesis remain unclear. We have determined two crystal structures of ERA, a binary complex with GDP and a ternary complex with a GTP-analog and the {sub 1531}AUCACCUCCUUA{sub 1542} sequence at the 3' end of 16S rRNA. In the ternary complex, the first nine of the 12 nucleotides are recognized by the protein. We show that GTP binding is a prerequisite for RNA recognition by ERA and that RNA recognition stimulates its GTP-hydrolyzing activity. Based on these and other data, we propose a functional cycle of ERA, suggesting that the protein serves as a chaperone for processing and maturation of 16S rRNA and a checkpoint for assembly of the 30S ribosomal subunit. The AUCA sequence is highly conserved among bacteria, archaea, and eukaryotes, whereas the CCUCC, known as the anti-Shine-Dalgarno sequence, is conserved in noneukaryotes only. Therefore, these data suggest a common mechanism for a highly conserved ERA function in all three kingdoms of life by recognizing the AUCA, with a 'twist' for noneukaryotic ERA proteins by also recognizing the CCUCC.

  6. Sulfur-oxidizing bacterial endosymbionts: analysis of phylogeny and specificity by 16S rRNA sequences.

    PubMed

    Distel, D L; Lane, D J; Olsen, G J; Giovannoni, S J; Pace, B; Pace, N R; Stahl, D A; Felbeck, H

    1988-06-01

    The 16S rRNAs from the bacterial endosymbionts of six marine invertebrates from diverse environments were isolated and partially sequenced. These symbionts included the trophosome symbiont of Riftia pachyptila, the gill symbionts of Calyptogena magnifica and Bathymodiolus thermophilus (from deep-sea hydrothermal vents), and the gill symbionts of Lucinoma annulata, Lucinoma aequizonata, and Codakia orbicularis (from relatively shallow coastal environments). Only one type of bacterial 16S rRNA was detected in each symbiosis. Using nucleotide sequence comparisons, we showed that each of the bacterial symbionts is distinct from the others and that all fall within a limited domain of the gamma subdivision of the purple bacteria (one of the major eubacterial divisions previously defined by 16S rRNA analysis [C. R. Woese, Microbiol. Rev. 51: 221-271, 1987]). Two host specimens were analyzed in five of the symbioses; in each case, identical bacterial rRNA sequences were obtained from conspecific host specimens. These data indicate that the symbioses examined are species specific and that the symbiont species are unique to and invariant within their respective host species. PMID:3286609

  7. The Cfr rRNA Methyltransferase Confers Resistance to Phenicols, Lincosamides, Oxazolidinones, Pleuromutilins, and Streptogramin A Antibiotics

    PubMed Central

    Long, Katherine S.; Poehlsgaard, Jacob; Kehrenberg, Corinna; Schwarz, Stefan; Vester, Birte

    2006-01-01

    A novel multidrug resistance phenotype mediated by the Cfr rRNA methyltransferase is observed in Staphylococcus aureus and Escherichia coli. The cfr gene has previously been identified as a phenicol and lincosamide resistance gene on plasmids isolated from Staphylococcus spp. of animal origin and recently shown to encode a methyltransferase that modifies 23S rRNA at A2503. Antimicrobial susceptibility testing shows that S. aureus and E. coli strains expressing the cfr gene exhibit elevated MICs to a number of chemically unrelated drugs. The phenotype is named PhLOPSA for resistance to the following drug classes: Phenicols, Lincosamides, Oxazolidinones, Pleuromutilins, and Streptogramin A antibiotics. Each of these five drug classes contains important antimicrobial agents that are currently used in human and/or veterinary medicine. We find that binding of the PhLOPSA drugs, which bind to overlapping sites at the peptidyl transferase center that abut nucleotide A2503, is perturbed upon Cfr-mediated methylation. Decreased drug binding to Cfr-methylated ribosomes has been confirmed by footprinting analysis. No other rRNA methyltransferase is known to confer resistance to five chemically distinct classes of antimicrobials. In addition, the findings described in this study represent the first report of a gene conferring transferable resistance to pleuromutilins and oxazolidinones. PMID:16801432

  8. The EMBL Nucleotide Sequence Database.

    PubMed

    Stoesser, G; Tuli, M A; Lopez, R; Sterk, P

    1999-01-01

    The EMBL Nucleotide Sequence Database (http://www.ebi.ac.uk/embl.html) constitutes Europe's primary nucleotide sequence resource. Main sources for DNA and RNA sequences are direct submissions from individual researchers, genome sequencing projects and patent applications. While automatic procedures allow incorporation of sequence data from large-scale genome sequencing centres and from the European Patent Office (EPO), the preferred submission tool for individual submitters is Webin (WWW). Through all stages, dataflow is monitored by EBI biologists communicating with the sequencing groups. In collaboration with DDBJ and GenBank the database is produced, maintained and distributed at the European Bioinformatics Institute (EBI). Database releases are produced quarterly and are distributed on CD-ROM. Network services allow access to the most up-to-date data collection via Internet and World Wide Web interface. EBI's Sequence Retrieval System (SRS) is a Network Browser for Databanks in Molecular Biology, integrating and linking the main nucleotide and protein databases, plus many specialised databases. For sequence similarity searching a variety of tools (e.g. Blitz, Fasta, Blast etc) are available for external users to compare their own sequences against the most currently available data in the EMBL Nucleotide Sequence Database and SWISS-PROT. PMID:9847133

  9. Automated Identification of Medically Important Bacteria by 16S rRNA Gene Sequencing Using a Novel Comprehensive Database, 16SpathDB▿

    PubMed Central

    Woo, Patrick C. Y.; Teng, Jade L. L.; Yeung, Juilian M. Y.; Tse, Herman; Lau, Susanna K. P.; Yuen, Kwok-Yung

    2011-01-01

    Despite the increasing use of 16S rRNA gene sequencing, interpretation of 16S rRNA gene sequence results is one of the most difficult problems faced by clinical microbiologists and technicians. To overcome the problems we encountered in the existing databases during 16S rRNA gene sequence interpretation, we built a comprehensive database, 16SpathDB (http://147.8.74.24/16SpathDB) based on the 16S rRNA gene sequences of all medically important bacteria listed in the Manual of Clinical Microbiology and evaluated its use for automated identification of these bacteria. Among 91 nonduplicated bacterial isolates collected in our clinical microbiology laboratory, 71 (78%) were reported by 16SpathDB as a single bacterial species having >98.0% nucleotide identity with the query sequence, 19 (20.9%) were reported as more than one bacterial species having >98.0% nucleotide identity with the query sequence, and 1 (1.1%) was reported as no match. For the 71 bacterial isolates reported as a single bacterial species, all results were identical to their true identities as determined by a polyphasic approach. For the 19 bacterial isolates reported as more than one bacterial species, all results contained their true identities as determined by a polyphasic approach and all of them had their true identities as the “best match in 16SpathDB.” For the isolate (Gordonibacter pamelaeae) reported as no match, the bacterium has never been reported to be associated with human disease and was not included in the Manual of Clinical Microbiology. 16SpathDB is an automated, user-friendly, efficient, accurate, and regularly updated database for 16S rRNA gene sequence interpretation in clinical microbiology laboratories. PMID:21389154

  10. On the time-course of adjacent and non-adjacent transposed-letter priming

    PubMed Central

    Ktori, Maria; Kingma, Brechtsje; Hannagan, Thomas; Holcomb, Phillip J.; Grainger, Jonathan

    2014-01-01

    We compared effects of adjacent (e.g., atricle-ARTICLE) and non-adjacent (e.g., actirle-ARTICLE) transposed-letter (TL) primes in an ERP study using the sandwich priming technique. TL priming was measured relative to the standard double-substitution condition. We found significantly stronger priming effects for adjacent transpositions than non-adjacent transpositions (with 2 intervening letters) in behavioral responses (lexical decision latencies), and the adjacent priming effects emerged earlier in the ERP signal, at around 200 ms post-target onset. Non-adjacent priming effects emerged about 50 ms later and were short-lived, being significant only in the 250-300 ms time-window. Adjacent transpositions on the other hand continued to produce priming in the N400 time-window (300-500 ms post-target onset). This qualitatively different pattern of priming effects for adjacent and non-adjacent transpositions is discussed in the light of different accounts of letter transposition effects, and the utility of drawing a distinction between positional flexibility and positional noise. PMID:25364497

  11. Quantitative Northern Blot Analysis of Mammalian rRNA Processing.

    PubMed

    Wang, Minshi; Pestov, Dimitri G

    2016-01-01

    Assembly of eukaryotic ribosomes is an elaborate biosynthetic process that begins in the nucleolus and requires hundreds of cellular factors. Analysis of rRNA processing has been instrumental for studying the mechanisms of ribosome biogenesis and effects of stress conditions on the molecular milieu of the nucleolus. Here, we describe the quantitative analysis of the steady-state levels of rRNA precursors, applicable to studies in mammalian cells and other organisms. We include protocols for gel electrophoresis and northern blotting of rRNA precursors using procedures optimized for the large size of these RNAs. We also describe the ratio analysis of multiple precursors, a technique that facilitates the accurate assessment of changes in the efficiency of individual pre-rRNA processing steps. PMID:27576717

  12. A nested array of rRNA targeted probes for the detection and identification of enterococci by reverse hybridization.

    PubMed

    Behr, T; Koob, C; Schedl, M; Mehlen, A; Meier, H; Knopp, D; Frahm, E; Obst, U; Schleifer, K; Niessner, R; Ludwig, W

    2000-12-01

    Complete 23S and almost complete 16S rRNA gene sequences were determined for the type strains of the validly described Enterococcus species, Melissococcus pluton and Tetragenococcus halophilus. A comprehensive set of rRNA targeted specific oligonucleotide hybridization probes was designed according to the multiple probe concept. In silico probe design and evaluation was performed using the respective tools of the ARB program package in combination with the ARB databases comprising the currently available 16S as well as 23S rRNA primary structures. The probes were optimized with respect to their application for reverse hybridization in microplate format. The target comprising 16S and 23S rDNA was amplified and labeled by PCR (polymerase chain reaction) using general primers targeting a wide spectrum of bacteria. Alternatively, amplification of two adjacent rDNA fragments of enterococci was performed by using specific primers. In vitro evaluation of the probe set was done including all Enterococcus type strains, and a selection of other representatives of the gram-positive bacteria with a low genomic DNA G+C content. The optimized probe set was used to analyze enriched drinking water samples as well as original samples from waste water treatment plants. PMID:11249027

  13. Archaea box C/D enzymes methylate two distinct substrate rRNA sequences with different efficiency.

    PubMed

    Graziadei, Andrea; Masiewicz, Pawel; Lapinaite, Audrone; Carlomagno, Teresa

    2016-05-01

    RNA modifications confer complexity to the 4-nucleotide polymer; nevertheless, their exact function is mostly unknown. rRNA 2'-O-ribose methylation concentrates to ribosome functional sites and is important for ribosome biogenesis. The methyl group is transferred to rRNA by the box C/D RNPs: The rRNA sequence to be methylated is recognized by a complementary sequence on the guide RNA, which is part of the enzyme. In contrast to their eukaryotic homologs, archaeal box C/D enzymes can be assembled in vitro and are used to study the mechanism of 2'-O-ribose methylation. In Archaea, each guide RNA directs methylation to two distinct rRNA sequences, posing the question whether this dual architecture of the enzyme has a regulatory role. Here we use methylation assays and low-resolution structural analysis with small-angle X-ray scattering to study the methylation reaction guided by the sR26 guide RNA fromPyrococcus furiosus We find that the methylation efficacy at sites D and D' differ substantially, with substrate D' turning over more efficiently than substrate D. This observation correlates well with structural data: The scattering profile of the box C/D RNP half-loaded with substrate D' is similar to that of the holo complex, which has the highest activity. Unexpectedly, the guide RNA secondary structure is not responsible for the functional difference at the D and D' sites. Instead, this difference is recapitulated by the nature of the first base pair of the guide-substrate duplex. We suggest that substrate turnover may occur through a zip mechanism that initiates at the 5'-end of the product. PMID:26925607

  14. Typification of virulent and low virulence Babesia bigemina clones by 18S rRNA and rap-1c.

    PubMed

    Thompson, C; Baravalle, M E; Valentini, B; Mangold, A; Torioni de Echaide, S; Ruybal, P; Farber, M; Echaide, I

    2014-06-01

    The population structure of original Babesia bigemina isolates and reference strains with a defined phenotypic profile was assessed using 18S rRNA and rap-1c genes. Two reference strains, BbiS2P-c (virulent) and BbiS1A-c (low virulence), were biologically cloned in vitro. The virulence profile of the strains and clones was assessed in vivo. One fully virulent and one low-virulence clone were mixed in identical proportions to evaluate their growth efficiency in vitro. Each clone was differentiated by two microsatellites and the gene gp45. The 18S rRNA and rap-1c genes sequences from B. bigemina biological clones and their parental strains, multiplied exclusively in vivo or in vitro, were compared with strain JG-29. The virulence of clones derived from the BbiS2P-c strain was variable. Virulent clone Bbi9P1 grew more efficiently in vitro than did the low-virulence clone Bbi2A1. The haplotypes generated by the nucleotide polymorphism, localized in the V4 region of the 18S rRNA, allowed the identification of three genotypes. The rap-1c haplotypes allowed defining four genotypes. Parental and original strains were defined by multiple haplotypes identified in both genes. The rap-1c gene, analyzed by high-resolution melting (HRM), allowed discrimination between two genotypes according to their phenotype, and both were different from JG-29. B. bigemina biological clones made it possible to define the population structure of isolates and strains. The polymorphic regions of the 18S rRNA and rap-1c genes allowed the identification of different subpopulations within original B. bigemina isolates by the definition of several haplotypes and the differentiation of fully virulent from low virulence clones. PMID:24681200

  15. Imprint of Ancient Evolution on rRNA Folding.

    PubMed

    Lanier, Kathryn A; Athavale, Shreyas S; Petrov, Anton S; Wartell, Roger; Williams, Loren Dean

    2016-08-23

    In a model describing the origin and evolution of the translation system, ribosomal RNA (rRNA) grew in size by accretion [Petrov, A. S., et al. (2015) History of the Ribosome and the Origin of Translation. Proc. Natl. Acad. Sci. U.S.A. 112, 15396-15401]. Large rRNAs were built up by iterative incorporation and encasement of small folded RNAs, in analogy with addition of new LEGOs onto the surface of a preexisting LEGO assembly. In this model, rRNA robustness in folding arises from inherited autonomy of local folding. We propose that rRNAs can be decomposed at various granularities, retaining folding mechanism and folding competence. To test these predictions, we disassembled Domain III of the large ribosomal subunit (LSU). We determined whether local rRNA structure, stability, and folding pathways are autonomous. Thermal melting, chemical footprinting, and circular dichroism were used to infer rules that govern folding of rRNA. We deconstructed Domain III of the LSU rRNA by mapping out its complex multistep melting pathway. We studied Domain III and two equal-size "sub-Domains" of Domain III. The combined results are consistent with a model in which melting transitions of Domain III are conserved upon cleavage into sub-Domains. Each of the eight melting transitions of Domain III corresponds in Tm and ΔH with a transition observed in one of the two isolated sub-Domains. The results support a model in which structure, stability, and folding mechanisms are dominated by local interactions and are unaffected by separation of the sub-Domains. Domain III rRNA is distinct from RNAs that form long-range cooperative interaction networks at early stages of folding or that do not fold reversibly. PMID:27428664

  16. ANALYSIS OF THE INTERACTION OF THE EG5 LOOP5 WITH THE NUCLEOTIDE SITE

    PubMed Central

    Harrington, Timothy D.; Naber, Nariman; Larson, Adam G.; Cooke, Roger; Rice, Sarah; Pate, Edward

    2011-01-01

    Loop 5 (L5) is a conserved loop that projects from the α2-helix adjacent to the nucleotide site of all kinesin-family motors. L5 is critical to the function of the mitotic kinesin-5 family motors and is the binding site for several kinesin-5 inhibitors that are currently in clinical trials. Its conformational dynamics and its role in motor function are not fully understood. Our previous work using EPR spectroscopy suggested that L5 alters the nucleotide pocket conformation of the kinesin-5 motor Eg5 [1]. EPR spectra of a spin-labeled nucleotide analog bound at the nucleotide site of Eg5 display a highly immobilized component that is absent if L5 is shortened or if the inhibitor STLC is added [1], which X-ray structures suggest stabilizes a L5 conformation pointing away from the nucleotide site. These data, coupled with the proximity of L5 to the nucleotide site suggest L5 could interact with a bound nucleotide, modulating function. Here we use molecular dynamics (MD) simulations of Eg5 to explore the interaction of L5 with the nucleotide site in greater detail. We performed MD simulations in which the L5-domain of the Eg5•ADP X-ray structure was manually deformed via backbone bond rotations. The L5-domain of Eg5 was sufficiently lengthy that portions of L5 could be located in proximity to bound ADP. The MD simulations evolved to thermodynamically stable structures at 300K showing that L5 can interact directly with bound nucleotide with significant impingement on the ribose hydroxyls, consistent with the EPR spectroscopy results. Taken together, these data provide support for the hypothesis that L5 modulates Eg5 function via interaction with the nucleotide-binding site. PMID:21872609

  17. Analysis of the interaction of the Eg5 Loop5 with the nucleotide site.

    PubMed

    Harrington, Timothy D; Naber, Nariman; Larson, Adam G; Cooke, Roger; Rice, Sarah E; Pate, Edward

    2011-11-21

    Loop 5 (L5) is a conserved loop that projects from the α2-helix adjacent to the nucleotide site of all kinesin-family motors. L5 is critical to the function of the mitotic kinesin-5 family motors and is the binding site for several kinesin-5 inhibitors that are currently in clinical trials. Its conformational dynamics and its role in motor function are not fully understood. Our previous work using EPR spectroscopy suggested that L5 alters the nucleotide pocket conformation of the kinesin-5 motor Eg5 (Larson et al., 2010). EPR spectra of a spin-labeled nucleotide analog bound at the nucleotide site of Eg5 display a highly immobilized component that is absent if L5 is shortened or if the inhibitor STLC is added (Larson et al., 2010), which X-ray structures suggest stabilizes an L5 conformation pointing away from the nucleotide site. These data, coupled with the proximity of L5 to the nucleotide site suggest L5 could interact with a bound nucleotide, modulating function. Here we use molecular dynamics (MD) simulations of Eg5 to explore the interaction of L5 with the nucleotide site in greater detail. We performed MD simulations in which the L5-domain of the Eg5·ADP X-ray structure was manually deformed via backbone bond rotations. The L5-domain of Eg5 was sufficiently lengthy that portions of L5 could be located in proximity to bound ADP. The MD simulations evolved to thermodynamically stable structures at 300 K showing that L5 can interact directly with bound nucleotide with significant impingement on the ribose hydroxyls, consistent with the EPR spectroscopy results. Taken together, these data provide support for the hypothesis that L5 modulates Eg5 function via interaction with the nucleotide-binding site. PMID:21872609

  18. Analysis of the Interaction of the Eg5 Loop5 with the Nucleotide Site

    SciTech Connect

    Harrington, Timothy D.; Naber, Nariman; Larson, Adam G.; Cooke, Roger; Rice, Sarah E.; Pate, Edward F.

    2011-11-21

    Loop 5 (L5) is a conserved loop that projects from the α2-helix adjacent to the nucleotide site of all kinesin-family motors. L5 is critical to the function of the mito tickinesin-5 family motors and is the binding site for several kinesin-5 inhibitors that are currently in clinical trials. Its conformational dynamics and its role in motor function are not fully understood. Our previous work using EPR spectroscopy suggested that L5 alters the nucleotide pocket conformation of the kinesin-5 motor Eg5 (Larsonetal.,2010). EPR spectra of a spin-labeled nucleotide analog bound at the nucleotide site of Eg5 display a highly immobilized component that is absent if L5 is shortened or if the inhibitor STLC is added (Larson etal.,2010), which X-ray structures suggest stabilizes an L5 conformation pointing away from the nucleotide site. These data, coupled with the proximity of L5 to the nucleotide site suggest L5 could interact with a bound nucleotide, modulating function. Here we use molecular dynamics (MD) simulations of Eg5 to explore the interaction of L5 with the nucleotide site in greater detail. We performed MD simulations in which the L5-domain of the Eg5•ADP X-ray structure was manually deformed via backbone bond rotations. The L5-domain of Eg5 was sufficiently lengthy that portions of L5 could belocated in proximity to bound ADP. The MD simulations evolved to thermodynamically stable structures at 300K showing that L5 can interact directly with bound nucleotide with significant impingement on the ribosehydroxyls, consistent with the EPR spectroscopy results. Taken together, these data provide support for the hypothes is that L5 modulates Eg5 function via interaction with the nucleotide-binding site.

  19. Applications of adenine nucleotide measurements in oceanography

    NASA Technical Reports Server (NTRS)

    Holm-Hansen, O.; Hodson, R.; Azam, F.

    1975-01-01

    The methodology involved in nucleotide measurements is outlined, along with data to support the premise that ATP concentrations in microbial cells can be extrapolated to biomass parameters. ATP concentrations in microorganisms and nucleotide analyses are studied.

  20. The Single Nucleotide Polymorphism Consortium

    NASA Technical Reports Server (NTRS)

    Morgan, Michael

    2003-01-01

    I want to discuss both the Single Nucleotide Polymorphism (SNP) Consortium and the Human Genome Project. I am afraid most of my presentation will be thin on law and possibly too high on rhetoric. Having been engaged in a personal and direct way with these issues as a trained scientist, I find it quite difficult to be always as objective as I ought to be.

  1. Nucleotide sequences 1986/1987

    SciTech Connect

    Not Available

    1987-01-01

    These eight volumes are the third annual published compendium of nucleic acid sequences included in the European Molecular Biology Laboratory Nucleotide Sequence Data Library and the GenBank Genetic Sequences Data Bank. Each volume surveys one or more subdivisions of the database. The volume subtitles are: Primates; Rodents; Other Vertebrates and Invertebrates, Plants and Organelles, Bacteria and Bacteriophage, Viruses, Structural RNA, Synthetic and Unannotated Sequences, and Database Directory and Master Indices.

  2. Nucleotides in neuroregeneration and neuroprotection.

    PubMed

    Miras-Portugal, M Teresa; Gomez-Villafuertes, Rosa; Gualix, Javier; Diaz-Hernandez, Juan Ignacio; Artalejo, Antonio R; Ortega, Felipe; Delicado, Esmerilda G; Perez-Sen, Raquel

    2016-05-01

    Brain injury generates the release of a multitude of factors including extracellular nucleotides, which exhibit bi-functional properties and contribute to both detrimental actions in the acute phase and also protective and reparative actions in the later recovery phase to allow neuroregeneration. A promising strategy toward restoration of neuronal function is based on activation of endogenous adult neural stem/progenitor cells. The implication of purinergic signaling in stem cell biology, including regulation of proliferation, differentiation, and cell death has become evident in the last decade. In this regard, current strategies of acute transplantation of ependymal stem/progenitor cells after spinal cord injury restore altered expression of P2X4 and P2X7 receptors and improve functional locomotor recovery. The expression of both receptors is transcriptionally regulated by Sp1 factor, which plays a key role in the startup of the transcription machinery to induce regeneration-associated genes expression. Finally, general signaling pathways triggered by nucleotide receptors in neuronal populations converge on several intracellular kinases, such as PI3K/Akt, GSK3 and ERK1,2, as well as the Nrf-2/heme oxigenase-1 axis, which specifically link them to neuroprotection. In this regard, regulation of dual specificity protein phosphatases can become novel mechanism of actions for nucleotide receptors that associate them to cell homeostasis regulation. This article is part of the Special Issue entitled 'Purines in Neurodegeneration and Neuroregeneration'. PMID:26359530

  3. Necessary relations for nucleotide frequencies.

    PubMed

    Sinclair, Robert

    2015-06-01

    Genome composition analysis of di-, tri- and tetra-nucleotide frequencies is known to be evolutionarily informative, and useful in metagenomic studies, where binning of raw sequence data is often an important first step. Patterns appearing in genome composition analysis may be due to evolutionary processes or purely mathematical relations. For example, the total number of dinucleotides in a sequence is equal to the sum of the individual totals of the sixteen types of dinucleotide, and this is entirely independent of any assumptions made regarding mutation or selection, or indeed any physical or chemical process. Before any statistical analysis can be attempted, a knowledge of all necessary mathematical relations is required. I show that 25% of di-, tri- and tetra-nucleotide frequencies can be written as simple sums and differences of the remainder. The vast majority of organisms have circular genomes, for which these relations are exact and necessary. In the case of linear molecules, the absolute error is very nearly zero, and does not grow with contiguous sequence length. As a result of the new, necessary relations presented here, the foundations of the statistical analysis of di-, tri- and tetra-nucleotide frequencies, and k-mer analysis in general, need to be revisited. PMID:25843217

  4. Guanine nucleotide depletion inhibits pre-ribosomal RNA synthesis and causes nucleolar disruption.

    PubMed

    Huang, Min; Ji, Yanshan; Itahana, Koji; Zhang, Yanping; Mitchell, Beverly

    2008-01-01

    Inosine monophosphate dehydrogenase (IMPDH) is a pivotal enzyme in the de novo pathway of guanine nucleotide biosynthesis. Inhibitors of this enzyme decrease intracellular guanine nucleotide levels by 50-80% and have potential as anti-neoplastic agents. Both mycophenolic acid (MPA) and AVN-944 are highly specific inhibitors of IMPDH that cause cell cycle arrest or apoptosis in lymphocytes and leukemic cell lines. We have examined the mechanisms by which these two agents cause cytotoxicity. Both MPA and AVN-944 inhibit the growth of K562 cells, and induce apoptosis in Raji B and CCRF-CEM T cells. Both compounds strikingly inhibit RNA synthesis within 2 h of exposure. Depletion of guanine nucleotides by MPA and AVN-944 also causes an early and near-complete reduction in levels of the 45S precursor rRNA synthesis and the concomitant translocation of nucleolar proteins including nucleolin, nucleophosmin, and nucleostemin from the nucleolus to the nucleoplasm. This efflux correlates temporally with the sustained induction of p53 in cell lines with wild-type p53. We conclude that inhibition of IMPDH causes a primary reduction in rRNA synthesis and secondary nucleolar disruption and efflux of nucleolar proteins that most likely mediate cell cycle arrest or apoptosis. The ability of AVN-944 to induce apoptosis in a number of leukemic cell lines supports its potential utility in the treatment of hematologic malignancies. PMID:17462731

  5. Nucleotide sequences of 5S rRNAs from sponge Halichondria japonica and tunicate Halocynthia roretzi and their phylogenetic positions

    PubMed Central

    Komiya, Hiroyuki; Hasegawa, Masami; Takemura, Shosuke

    1983-01-01

    The nucleotide sequences of 5S rRNAs from sponge Halichondria japonica and tunicate Halocynthia roretzi were determined by chemical and enzymatic gel methods. Their phylogenetic positions among metazoans were derived from the 5S rRNA sequences by a computer analysis based on the maximum parsimony principle. It was suggested that the sponge is closely related to several invertebrates and the tunicate has affinity to vertebrates rather than invertebrates. PMID:6835845

  6. Border separation for adjacent orthogonal fields

    SciTech Connect

    Werner, B.L.; Khan, F.M.; Sharma, S.C.; Lee, C.K.; Kim, T.H. )

    1991-06-01

    Field border separations for adjacent orthogonal fields can be calculated geometrically, given the validity of some important assumptions such as beam alignment and field uniformity. Thermoluminescent dosimetry (TLD) measurements were used to investigate dose uniformity across field junctions as a function of field separation and, in particular, to review the CCSG recommendation for the treatment of medulloblastoma with separate head and spine fields.

  7. SOLIDS TRANSPORT BETWEEN ADJACENT CAFB FLUIDIZED BEDS

    EPA Science Inventory

    The report gives results of an experimental investigation of a pulsed, dense-phase pneumatic transport system for controlled circulation between adjacent fluidized beds. A model was developed to predict performance. The program provides technical support for EPA's program to demo...

  8. Interactions of the TnaC nascent peptide with rRNA in the exit tunnel enable the ribosome to respond to free tryptophan.

    PubMed

    Martínez, Allyson K; Gordon, Emily; Sengupta, Arnab; Shirole, Nitin; Klepacki, Dorota; Martinez-Garriga, Blanca; Brown, Lewis M; Benedik, Michael J; Yanofsky, Charles; Mankin, Alexander S; Vazquez-Laslop, Nora; Sachs, Matthew S; Cruz-Vera, Luis R

    2014-01-01

    A transcriptional attenuation mechanism regulates expression of the bacterial tnaCAB operon. This mechanism requires ribosomal arrest induced by the regulatory nascent TnaC peptide in response to free L-tryptophan (L-Trp). In this study we demonstrate, using genetic and biochemical analyses, that in Escherichia coli, TnaC residue I19 and 23S rRNA nucleotide A2058 are essential for the ribosome's ability to sense free L-Trp. We show that the mutational change A2058U in 23S rRNA reduces the concentration dependence of L-Trp-mediated tna operon induction, whereas the TnaC I19L change suppresses this phenotype, restoring the sensitivity of the translating A2058U mutant ribosome to free L-Trp. These findings suggest that interactions between TnaC residue I19 and 23S rRNA nucleotide A2058 contribute to the creation of a regulatory L-Trp binding site within the translating ribosome. PMID:24137004

  9. Nucleotide composition of CO1 sequences in Chelicerata (Arthropoda): detecting new mitogenomic rearrangements.

    PubMed

    Arabi, Juliette; Judson, Mark L I; Deharveng, Louis; Lourenço, Wilson R; Cruaud, Corinne; Hassanin, Alexandre

    2012-02-01

    Here we study the evolution of nucleotide composition in third codon-positions of CO1 sequences of Chelicerata, using a phylogenetic framework, based on 180 taxa and three markers (CO1, 18S, and 28S rRNA; 5,218 nt). The analyses of nucleotide composition were also extended to all CO1 sequences of Chelicerata found in GenBank (1,701 taxa). The results show that most species of Chelicerata have a positive strand bias in CO1, i.e., in favor of C nucleotides, including all Amblypygi, Palpigradi, Ricinulei, Solifugae, Uropygi, and Xiphosura. However, several taxa show a negative strand bias, i.e., in favor of G nucleotides: all Scorpiones, Opisthothelae spiders and several taxa within Acari, Opiliones, Pseudoscorpiones, and Pycnogonida. Several reversals of strand-specific bias can be attributed to either a rearrangement of the control region or an inversion of a fragment containing the CO1 gene. Key taxa for which sequencing of complete mitochondrial genomes will be necessary to determine the origin and nature of mtDNA rearrangements involved in the reversals are identified. Acari, Opiliones, Pseudoscorpiones, and Pycnogonida were found to show a strong variability in nucleotide composition. In addition, both mitochondrial and nuclear genomes have been affected by higher substitution rates in Acari and Pseudoscorpiones. The results therefore indicate that these two orders are more liable to fix mutations of all types, including base substitutions, indels, and genomic rearrangements. PMID:22362465

  10. Analysis of the primary sequence and secondary structure of the unusually long SSU rRNA of the soil bug, Armadillidium vulgare.

    PubMed

    Choe, C P; Hancock, J M; Hwang, U W; Kim, W

    1999-12-01

    The complete nucleotide sequence of the SSU rRNA gene from the soil bug, Armadillidium vulgare (Crustacea, Isopoda), was determined. It is 3214 bp long, with a GC content of 56.3%. It is not only the longest SSU rRNA gene among Crustacea but also longer than any other SSU rRNA gene except that of the strepsipteran insect, Xenos vesparum (3316 bp). The unusually long sequence of this species is explained by the long sequences of variable regions V4 and V7, which make up more than half of the total length. RT-PCR analysis of these two regions showed that the long sequences also exist in the mature rRNA and sequence simplicity analysis revealed the presence of slippage motifs in these two regions. The putative secondary structure of the rRNA is typical for eukaryotes except for the length and shape variations of the V2, V4, V7, and V9 regions. Each of the V2, V4, and V7 regions was elongated, while the V9 region was shortened. In V2, two bulges, located between helix 8 and helix 9 and between helix 9 and helix 10, were elongated. In V4, stem E23-3 was dramatically expanded, with several small branched stems. In V7, stem 43 was branched and expanded. Comparisons with the unusually long SSU rRNAs of other organisms imply that the increase in total length of SSU rRNA is due mainly to expansion in the V4 and V7 regions. PMID:10594181

  11. Mitochondrial m.1584A 12S m62A rRNA methylation in families with m.1555A>G associated hearing loss.

    PubMed

    O'Sullivan, Mary; Rutland, Paul; Lucas, Deirdre; Ashton, Emma; Hendricks, Sebastian; Rahman, Shamima; Bitner-Glindzicz, Maria

    2015-02-15

    The mitochondrial DNA mutation m.1555A>G predisposes to hearing loss following aminoglycoside antibiotic exposure in an idiosyncratic dose-independent manner. However, it may also cause maternally inherited hearing loss in the absence of aminoglycoside exposure or any other clinical features (non-syndromic hearing loss). Although m.1555A>G was identified as a cause of deafness more than twenty years ago, the pathogenic mechanism of this mutation of ribosomal RNA remains controversial. Different mechanistic concepts have been proposed. Most recently, evidence from cell lines and animal models suggested that patients with m.1555A>G may have more 12S rRNA N6, N6-dimethyladenosine (m(6) 2A) methylation than controls, so-called 'hypermethylation'. This has been implicated as a pathogenic mechanism of mitochondrial dysfunction but has yet to be validated in patients. 12S m(6) 2A rRNA methylation, by the mitochondrial transcription factor 1 (TFB1M) enzyme, occurs at two successive nucleotides (m.1584A and m.1583A) in close proximity to m.1555A>G. We examined m(6) 2A methylation in 14 patients with m.1555A>G, and controls, and found all detectable 12S rRNA transcripts to be methylated in both groups. Moreover, different RNA samples derived from the same patient (lymphocyte, fibroblast and lymphoblast) revealed that only transformed cells contained some unmethylated 12S rRNA transcripts, with all detectable 12S rRNA transcripts derived from primary samples m(6) 2A-methylated. Our data indicate that TFB1M 12S m(6) 2A rRNA hypermethylation is unlikely to be a pathogenic mechanism and may be an artefact of previous experimental models studied. We propose that RNA methylation studies in experimental models should be validated in primary clinical samples to ensure that they are applicable to the human situation. PMID:25305075

  12. Assessing hog lagoon waste contamination in the Cape Fear Watershed using Bacteroidetes 16S rRNA gene pyrosequencing.

    PubMed

    Arfken, Ann M; Song, Bongkeun; Mallin, Michael A

    2015-09-01

    Hog lagoons can be major sources of waste and nutrient contamination to watersheds adjacent to pig farms. Fecal source tracking methods targeting Bacteroidetes 16S rRNA genes in pig fecal matter may underestimate or fail to detect hog lagoon contamination in riverine environments. In order to detect hog lagoon wastewater contamination in the Cape Fear Watershed, where a large number of hog farms are present, we conducted pyrosequencing analyses of Bacteroidetes 16S rRNA genes in hog lagoon waste and identified new hog lagoon-specific marker sequences. Additional pyrosequencing analyses of Bacteroidetes 16S rRNA genes were conducted with surface water samples collected at 4 sites during 5 months in the Cape Fear Watershed. Using an operational taxonomic unit (OTU) identity cutoff value of 97 %, these newly identified hog lagoon markers were found in 3 of the river samples, while only 1 sample contained the pig fecal marker. In the sample containing the pig fecal marker, there was a relatively high percentage (14.1 %) of the hog lagoon markers and a low pig fecal marker relative abundance of 0.4 % in the Bacteroidetes 16S rRNA gene sequences. This suggests that hog lagoon contamination must be somewhat significant in order for pig fecal markers to be detected, and low levels of hog lagoon contamination cannot be detected targeting only pig-specific fecal markers. Thus, new hog lagoon markers have a better detection capacity for lagoon waste contamination, and in conjunction with a pig fecal marker, provide a more comprehensive and accurate detection of hog lagoon waste contamination in susceptible watersheds. PMID:26189016

  13. Regulation of Arabidopsis thaliana 5S rRNA Genes.

    PubMed

    Vaillant, Isabelle; Tutois, Sylvie; Cuvillier, Claudine; Schubert, Ingo; Tourmente, Sylvette

    2007-05-01

    The Arabidopsis thaliana genome comprises around 1,000 copies of 5S rRNA genes encoding both major and minor 5S rRNAs. In mature wild-type leaves, the minor 5S rRNA genes are silent. Using different mutants of DNA methyltransferases (met1, cmt3 and met1 cmt3), components of the RNAi pathway (ago4) or post-translational histone modifier (hda6/sil1), we show that the corresponding proteins are needed to maintain proper methylation patterns at heterochromatic 5S rDNA repeats. Using reverse transcription-PCR and cytological analyses, we report that a decrease of 5S rDNA methylation at CG or CNG sites in these mutants leads to the release of 5S rRNA gene silencing which occurred without detectable changes of the 5S rDNA chromatin structure. In spite of severely reduced DNA methylation, the met1 cmt3 double mutant revealed no increase in minor 5S rRNA transcripts. Furthermore, the release of silencing of minor 5S rDNAs can be achieved without increased formation of euchromatic loops by 5S rDNA, and is independent from the global heterochromatin content. Additionally, fluorescence in situ hybridization with centromeric 180 bp repeats confirmed that these highly repetitive sequences, in spite of their elevated transcriptional activity in the DNA methyltransferase mutants (met1, cmt3 and met1 cmt3), remain within chromocenters of the mutant nuclei. PMID:17412735

  14. Phylogeny and evolutionary genetics of Frankia strains based on 16S rRNA and nifD-K gene sequences.

    PubMed

    Mishra, Arun Kumar; Singh, Pawan Kumar; Singh, Prashant; Singh, Anumeha; Singh, Satya Shila; Srivastava, Amrita; Srivastava, Alok Kumar; Sarma, Hridip Kumar

    2015-08-01

    16S rRNA and nifD-nifK sequences were used to study the molecular phylogeny and evolutionary genetics of Frankia strains isolated from Hippöphae salicifolia D. Don growing at different altitudes (ecologically classified as riverside and hillside isolates) of the Eastern Himalayan region of North Sikkim, India. Genetic information for the small subunit rRNA (16S rRNA) revealed that the riverside Frankia isolates markedly differed from the hillside isolates suggesting that the riverside isolates are genetically compact. Further, for enhanced resolutions, the partial sequence of nifD (3' end), nifK (5' end) and nifD-K IGS region have been investigated. The sequences obtained, failed to separate riverside isolates and hillside isolates, thus suggesting a possible role of genetic transfer events either from hillside to riverside or vice versa. The evolutionary genetic analyses using evogenomic extrapolations of gene sequence data obtained from 16S rRNA and nifD-K provided differing equations with the pace of evolution being more appropriately, intermediate. Values of recombination frequency (R), nucleotide diversity per site (Pi), and DNA divergence estimates supported the existence of an intermixed zone where spatial isolations occurred in sync with the temporal estimates. J. Basic Microbiol. 2015, 54, 1-9. PMID:25871924

  15. Natural Microbial Community Compositions Compared by a Back-Propagating Neural Network and Cluster Analysis of 5S rRNA

    PubMed Central

    Noble, P. A.; Bidle, K. D.; Fletcher, M.

    1997-01-01

    The community compositions of free-living and particle-associated bacteria in the Chesapeake Bay estuary were analyzed by comparing banding patterns of stable low-molecular-weight RNA (SLMW RNA) which include 5S rRNA and tRNA molecules. By analyzing images of autoradiographs of SLMW RNAs on polyacrylamide gels, band intensities of 5S rRNA were converted to binary format for transmission to a back-propagating neural network (NN). The NN was trained to relate binary input to sample stations, collection times, positions in the water column, and sample types (e.g., particle-associated versus free-living communities). Dendrograms produced by using Euclidean distance and average and Ward's linkage methods on data of three independently trained NNs yielded the following results. (i) Community compositions of Chesapeake Bay water samples varied both seasonally and spatially. (ii) Although there was no difference in the compositions of free-living and particle-associated bacteria in the summer, these community types differed significantly in the winter. (iii) In the summer, most bay samples had a common 121-nucleotide 5S rRNA molecule. Although this band occurred in the top water of midbay samples, it did not occur in particle-associated communities of bottom-water samples. (iv) Regardless of the season, midbay samples had the greatest variety of 5S rRNA sizes. The utility of NNs for interpreting complex banding patterns in electrophoresis gels was demonstrated. PMID:16535593

  16. Adjacent Segment Pathology after Lumbar Spinal Fusion.

    PubMed

    Lee, Jae Chul; Choi, Sung-Woo

    2015-10-01

    One of the major clinical issues encountered after lumbar spinal fusion is the development of adjacent segment pathology (ASP) caused by increased mechanical stress at adjacent segments, and resulting in various radiographic changes and clinical symptoms. This condition may require surgical intervention. The incidence of ASP varies with both the definition and methodology adopted in individual studies; various risk factors for this condition have been identified, although a significant controversy still exists regarding their significance. Motion-preserving devices have been developed, and some studies have shown their efficacy of preventing ASP. Surgeons should be aware of the risk factors of ASP when planning a surgery, and accordingly counsel their patients preoperatively. PMID:26435804

  17. Adjacent Segment Pathology after Anterior Cervical Fusion

    PubMed Central

    Chung, Jae Yoon; Park, Jong-Beom; Seo, Hyoung-Yeon

    2016-01-01

    Anterior cervical fusion has become a standard of care for numerous pathologic conditions of the cervical spine. However, subsequent development of clinically significant disc disease at levels adjacent to fused discs is a serious long-term complication of this procedure. As more patients live longer after surgery, it is foreseeable that adjacent segment pathology (ASP) will develop in increasing numbers of patients. Also, ASP has been studied more intensively with the recent popularity of motion preservation technologies like total disc arthroplasty. The true nature and scope of ASP remains poorly understood. The etiology of ASP is most likely multifactorial. Various factors including altered biomechanical stresses, surgical disruption of soft tissue and the natural history of cervical disc disease contribute to the development of ASP. General factors associated with disc degeneration including gender, age, smoking and sports may play a role in the development of ASP. Postoperative sagittal alignment and type of surgery are also considered potential causes of ASP. Therefore, a spine surgeon must be particularly careful to avoid unnecessary disruption of the musculoligamentous structures, reduced risk of direct injury to the disc during dissection and maintain a safe margin between the plate edge and adjacent vertebrae during anterior cervical fusion. PMID:27340541

  18. Adjacent Segment Pathology after Anterior Cervical Fusion.

    PubMed

    Chung, Jae Yoon; Park, Jong-Beom; Seo, Hyoung-Yeon; Kim, Sung Kyu

    2016-06-01

    Anterior cervical fusion has become a standard of care for numerous pathologic conditions of the cervical spine. However, subsequent development of clinically significant disc disease at levels adjacent to fused discs is a serious long-term complication of this procedure. As more patients live longer after surgery, it is foreseeable that adjacent segment pathology (ASP) will develop in increasing numbers of patients. Also, ASP has been studied more intensively with the recent popularity of motion preservation technologies like total disc arthroplasty. The true nature and scope of ASP remains poorly understood. The etiology of ASP is most likely multifactorial. Various factors including altered biomechanical stresses, surgical disruption of soft tissue and the natural history of cervical disc disease contribute to the development of ASP. General factors associated with disc degeneration including gender, age, smoking and sports may play a role in the development of ASP. Postoperative sagittal alignment and type of surgery are also considered potential causes of ASP. Therefore, a spine surgeon must be particularly careful to avoid unnecessary disruption of the musculoligamentous structures, reduced risk of direct injury to the disc during dissection and maintain a safe margin between the plate edge and adjacent vertebrae during anterior cervical fusion. PMID:27340541

  19. Coordinated tissue-specific regulation of adjacent alternative 3′ splice sites in C. elegans

    PubMed Central

    Ragle, James Matthew; Katzman, Sol; Akers, Taylor F.; Barberan-Soler, Sergio; Zahler, Alan M.

    2015-01-01

    Adjacent alternative 3′ splice sites, those separated by ≤18 nucleotides, provide a unique problem in the study of alternative splicing regulation; there is overlap of the cis-elements that define the adjacent sites. Identification of the intron's 3′ end depends upon sequence elements that define the branchpoint, polypyrimidine tract, and terminal AG dinucleotide. Starting with RNA-seq data from germline-enriched and somatic cell-enriched Caenorhabditis elegans samples, we identify hundreds of introns with adjacent alternative 3′ splice sites. We identify 203 events that undergo tissue-specific alternative splicing. For these, the regulation is monodirectional, with somatic cells preferring to splice at the distal 3′ splice site (furthest from the 5′ end of the intron) and germline cells showing a distinct shift toward usage of the adjacent proximal 3′ splice site (closer to the 5′ end of the intron). Splicing patterns in somatic cells follow C. elegans consensus rules of 3′ splice site definition; a short stretch of pyrimidines preceding an AG dinucleotide. Splicing in germline cells occurs at proximal 3′ splice sites that lack a preceding polypyrimidine tract, and in three instances the germline-specific site lacks the AG dinucleotide. We provide evidence that use of germline-specific proximal 3′ splice sites is conserved across Caenorhabditis species. We propose that there are differences between germline and somatic cells in the way that the basal splicing machinery functions to determine the intron terminus. PMID:25922281

  20. Structural Rearrangements in the Active Site of the Thermus thermophilus 16S rRNA Methyltransferase KsgA in a Binary Complex with 5'-Methylthioadenosine

    SciTech Connect

    Demirci, H.; Belardinelli, R; Seri, E; Gregory, S; Gualerzi, C; Dahlberg, A; Jogl, G

    2009-01-01

    Posttranscriptional modification of ribosomal RNA (rRNA) occurs in all kingdoms of life. The S-adenosyl-l-methionine-dependent methyltransferase KsgA introduces the most highly conserved rRNA modification, the dimethylation of A1518 and A1519 of 16S rRNA. Loss of this dimethylation confers resistance to the antibiotic kasugamycin. Here, we report biochemical studies and high-resolution crystal structures of KsgA from Thermus thermophilus. Methylation of 30S ribosomal subunits by T. thermophilus KsgA is more efficient at low concentrations of magnesium ions, suggesting that partially unfolded RNA is the preferred substrate. The overall structure is similar to that of other methyltransferases but contains an additional ?-helix in a novel N-terminal extension. Comparison of the apoenzyme with complex structures with 5?-methylthioadenosine or adenosine bound in the cofactor-binding site reveals novel features when compared with related enzymes. Several mobile loop regions that restrict access to the cofactor-binding site are observed. In addition, the orientation of residues in the substrate-binding site indicates that conformational changes are required for binding two adjacent residues of the substrate rRNA.

  1. Frequency and spectrum of mitochondrial 12S rRNA variants in 440 Han Chinese hearing impaired pediatric subjects from two otology clinics

    PubMed Central

    2011-01-01

    Background Aminoglycoside ototoxicity is one of the common health problems. Mitochondrial 12S rRNA mutations are one of the important causes of aminoglycoside ototoxicity. However, the incidences of 12S rRNA mutations associated with aminoglycoside ototoxicity are less known. Methods A total of 440 Chinese pediatric hearing-impaired subjects were recruited from two otology clinics in the Ningbo and Wenzhou cities of Zhejiang Province, China. These subjects underwent clinical, genetic evaluation and molecular analysis of mitochondrial 12S rRNA. Resultant mtDNA variants were evaluated by structural and phylogenetic analysis. Results The study samples consisted of 227 males and 213 females. The age of all participants ranged from 1 years old to 18 years, with the median age of 9 years. Ninety-eight subjects (58 males and 40 females) had a history of exposure to aminoglycosides, accounting for 22.3% cases of hearing loss in this cohort. Molecular analysis of 12S rRNA gene identified 41 (39 known and 2 novel) variants. The incidences of the known deafness-associated 1555A > G, 1494C > T and 1095T > C mutations were 7.5%, 0.45% and 0.91% in this entire hearing-impaired subjects, respectively, and 21.4%, 2% and 2% among 98 subjects with aminoglycoside ototoxicity, respectively. The structural and phylogenetic evaluations showed that a novel 747A > G variant and known 839A > G, 1027A > G, 1310C > T and 1413T > C variants conferred increased sensitivity to aminoglycosides or nonsyndromic deafness as they were absent in 449 Chinese controls and localized at highly conserved nucleotides of this rRNA. However, other variants were polymorphisms. Of 44 subjects carrying one of definite or putative deafness-related 12S rRNA variants, only one subject carrying the 1413T > C variant harbored the 235DelC/299DelAT mutations in the GJB2 gene, while none of mutations in GJB2 gene was detected in other 43 subjects. Conclusions Mutations in mitochondrial 12S rRNA accounted for ~30% cases

  2. ATCG nucleotide fluctuation of Deinococcus radiodurans radiation genes

    NASA Astrophysics Data System (ADS)

    Holden, Todd; Subramaniam, R.; Sullivan, R.; Cheung, E.; Schneider, C.; Tremberger, G., Jr.; Flamholz, A.; Lieberman, D. H.; Cheung, T. D.

    2007-09-01

    The radiation resistance-repair genes in Deinococcus radiodurans (DR) and E-coli were analyzed in terms of the A, T, C, G nucleotide fluctuations. The studied genes were Rec-A, Rec-Q, and the unique DR PprA gene. In an ATCG sequence, each base was assigned a number equal to its atomic number. The resulting numerical sequence was the basis of the statistical analysis. Fractal analysis using the Higuchi method gave a fractal dimension increase of the Deinococcus radiodurans genes as compared to E-coli, which is comparable to the enhancement observed in the human HAR1 region (HAR1F gene) over that of the chimpanzee. Near neighbor fluctuation was also studied via the Black-Scholes model where the increment sequence was treated as a random walk series. The Deinococcus radiodurans radiation gene standard deviations were consistently higher than that of the E-coli deviations, and agree with the fractal analysis results. The sequence stacking interaction was studied using the published nucleotide-pair melting free energy values and Deinococcus radiodurans radiation genes were shown to possess larger negative free energies. The high sensitivity of the fractal dimension as a biomarker was tested with correlation analysis of the gamma ray dose versus fractal dimension, and the R square values were found to be above 0.9 (N=5). When compared with other nucleotide sequences such as the rRNA sequences, HAR1 and its chimpanzee counterpart, the higher fluctuation (correlated randomness) and larger negative free energy of a DR radiation gene suggested that a radiation resistance-repair sequence exhibited higher complexity. As the HAR1 nucleotide sequence complexity and its transcription activity of co-expressing cortex protein reelin supported a positive selection event in humans, a similar inference of positive selection of coding genes could be drawn for Deinococcus radiodurans when compared to E-coli. The origin of such a positive selection would be consistent with that of a

  3. The effect of imidazole, cyanamide, and polyornithine on the condensation of nucleotides in aqueous systems.

    NASA Technical Reports Server (NTRS)

    Ibanez, J.; Kimball, A. P.; Oro, J.

    1971-01-01

    Development of two models for the condensation of nucleotides under possibly prebiotic conditions. In the first of these models this type of reaction is promoted by the presence of imidazole and substituted imidazole compounds. The second model involves the condensation of mononucleotides with cyanamide in the presence and absence of a prototemplate such as polyornithine. A tentative mechanism for the role of imidazole catalysis in phosphodiester bond formation between adjacent TMP molecules is suggested.

  4. Alternate rRNA secondary structures as regulators of translation.

    PubMed

    Feng, Shu; Li, Heng; Zhao, Jing; Pervushin, Konstantin; Lowenhaupt, Ky; Schwartz, Thomas U; Dröge, Peter

    2011-02-01

    Structural dynamics of large molecular assemblies are intricately linked to function. For ribosomes, macromolecular changes occur especially during mRNA translation and involve participation of ribosomal RNA. Without suitable probes specific to RNA secondary structure, however, elucidation of more subtle dynamic ribosome structure-function relationships, especially in vivo, remains challenging. Here we report that the Z-DNA- and Z-RNA-binding domain Zα, derived from the human RNA editing enzyme ADAR1-L, binds with high stability to specific rRNA segments of Escherichia coli and human ribosomes. Zα impaired in Z-RNA recognition does not associate with ribosomes. Notably, Zα(ADAR1)-ribosome interaction blocks translation in vitro and in vivo, with substantial physiological consequences. Our study shows that ribosomes can be targeted by a protein that specifically recognizes an alternate rRNA secondary structure, and suggests a new mechanism of translational regulation on the ribosome. PMID:21217697

  5. Analysis of a 5S rRNA gene cloned from Euplotes eurstomus

    SciTech Connect

    Roberson, A.E.; Wolffe, A.; Olins, D.E.

    1987-05-01

    The macronucleus of the hypotrichous ciliated protozoan Euplotes eurystomus lends itself to the study of eukaryotic gene and chromatin structure because native macronuclear DNA exists as linear, gene-sized fragments between 400 and 20,000 bp in length. The macronuclear chromatin, while arranged in a typical nucleosomal structure, is freely soluble in low ionic strength buffers without treatment by nucleases. Thus, specific genes may be enriched as native, intact chromatin molecules. The 5S rRNA gene from Euplotes has been cloned to facilitate investigation of 5S gene-chromatin following characterization of the gene at the DNA level. It has been demonstrated that the gene, while in circular or linear form, can be transcribed in vitro by a Xenopus oocyte nuclear extract. The transcript generated in vitro is 120 nucleotides in length and is synthesized by RNA polymerase III. Anti-Xenopus TFIIIA antibodies recognize a Euplotes macronuclear chromatin-associated protein which is approx. 80 KD in size. It has been established that the sequence of the telomere flanking the 5S gene in Euplotes eurystomus is the same telomeric sequence published for Euplotes aediculatus.

  6. Two distinct promoter elements in the human rRNA gene identified by linker scanning mutagenesis.

    PubMed Central

    Haltiner, M M; Smale, S T; Tjian, R

    1986-01-01

    A cell-free RNA polymerase I transcription system was used to evaluate the transcription efficiency of 21 linker scanning mutations that span the human rRNA gene promoter. Our analysis revealed the presence of two major control elements, designated the core and upstream elements, that affect the level of transcription initiation. The core element extends from -45 to +18 relative to the RNA start site, and transcription is severely affected (up to 100-fold) by linker scanning mutations in this region. Linker scanning and deletion mutations in the upstream element, located between nucleotides -156 and -107, cause a three- to fivefold reduction in transcription. Under certain reaction conditions, such as the presence of a high ratio of protein to template or supplementation of the reaction with partially purified protein fractions, sequences upstream of the core element can have an even greater effect (20- to 50-fold) on RNA polymerase I transcription. Primer extension analysis showed that RNA synthesized from all of these mutant templates is initiated at the correct in vivo start site. To examine the functional relationship between the core and the upstream region, mutant promoters were constructed that alter the orientation, distance, or multiplicity of these control elements relative to each other. The upstream control element appears to function in only one orientation, and its position relative to the core is constrained within a fairly narrow region. Moreover, multiple core elements in close proximity to each other have an inhibitory effect on transcription. Images PMID:3785147

  7. Robust Computational Analysis of rRNA Hypervariable Tag Datasets

    PubMed Central

    Sipos, Maksim; Jeraldo, Patricio; Chia, Nicholas; Qu, Ani; Dhillon, A. Singh; Konkel, Michael E.; Nelson, Karen E.; White, Bryan A.; Goldenfeld, Nigel

    2010-01-01

    Next-generation DNA sequencing is increasingly being utilized to probe microbial communities, such as gastrointestinal microbiomes, where it is important to be able to quantify measures of abundance and diversity. The fragmented nature of the 16S rRNA datasets obtained, coupled with their unprecedented size, has led to the recognition that the results of such analyses are potentially contaminated by a variety of artifacts, both experimental and computational. Here we quantify how multiple alignment and clustering errors contribute to overestimates of abundance and diversity, reflected by incorrect OTU assignment, corrupted phylogenies, inaccurate species diversity estimators, and rank abundance distribution functions. We show that straightforward procedural optimizations, combining preexisting tools, are effective in handling large () 16S rRNA datasets, and we describe metrics to measure the effectiveness and quality of the estimators obtained. We introduce two metrics to ascertain the quality of clustering of pyrosequenced rRNA data, and show that complete linkage clustering greatly outperforms other widely used methods. PMID:21217830

  8. A phylogenetic comparison of the 16S rRNA sequence of the fish pathogen, Renibacterium salmoninarum, to gram-positive bacteria.

    PubMed

    Gutenberger, S K; Giovannoni, S J; Field, K G; Fryer, J L; Rohovec, J S

    1991-01-15

    The 16S rRNA of Renibacterium salmoninarum, the causative agent of bacterial kidney disease in salmonids, was sequenced by reverse transcriptase to produce a nearly complete sequence (97%) of 1475 nucleotides. Phylogenetic comparisons to seventeen genera and signature sequence analysis indicated that R. salmoninarum was a member of the high G + C Gram-positive eubacterial subdivision although the reported G + C value is only 53%. A phylogenetic tree details the relationship of R. salmoninarum to ten actinomycetes from diverse environments. PMID:1709893

  9. Complete sequence and gene organization of the Nosema spodopterae rRNA gene.

    PubMed

    Tsai, Shu-Jen; Huang, Wei-Fone; Wang, Chung-Hsiung

    2005-01-01

    By sequencing the entire ribosomal RNA (rRNA) gene of Nosema spodopterae, we show here that its gene organization follows a pattern similar to the Nosema type species, Nosema bombycis, i.e. 5'-large subunit rRNA (2,497 bp)-internal transcribed spacer (185 bp)-small subunit rRNA (1,232 bp)-intergenic spacer (277 bp)-5S rRNA (114 bp)-3'. Gene sequences and the secondary structures of large subunit rRNA, small subunit rRNA, and 5S rRNA are compared with the known corresponding sequences and structures of closely related microsporidia. The results suggest that the Nosema genus may be heterogeneous and that the rRNA gene organization may be a useful characteristic for determining which species are closely related to the type species. PMID:15702980

  10. Reconstructing genome mixtures from partial adjacencies.

    PubMed

    Mahmoody, Ahmad; Kahn, Crystal L; Raphael, Benjamin J

    2012-01-01

    Many cancer genome sequencing efforts are underway with the goal of identifying the somatic mutations that drive cancer progression. A major difficulty in these studies is that tumors are typically heterogeneous, with individual cells in a tumor having different complements of somatic mutations. However, nearly all DNA sequencing technologies sequence DNA from multiple cells, thus resulting in measurement of mutations from a mixture of genomes. Genome rearrangements are a major class of somatic mutations in many tumors, and the novel adjacencies (i.e. breakpoints) resulting from these rearrangements are readily detected from DNA sequencing reads. However, the assignment of each rearrangement, or adjacency, to an individual cancer genome in the mixture is not known. Moreover, the quantity of DNA sequence reads may be insufficient to measure all rearrangements in all genomes in the tumor. Motivated by this application, we formulate the k-minimum completion problem (k-MCP). In this problem, we aim to reconstruct k genomes derived from a single reference genome, given partial information about the adjacencies present in the mixture of these genomes. We show that the 1-MCP is solvable in linear time in the cases where: (i) the measured, incomplete genome has a single circular or linear chromosome; (ii) there are no restrictions on the chromosomal content of the measured, incomplete genome. We also show that the k-MCP problem, for k ≥ 3 in general, and the 2-MCP problem with the double-cut-and-join (DCJ) distance are NP-complete, when there are no restriction on the chromosomal structure of the measured, incomplete genome. These results lay the foundation for future algorithmic studies of the k-MCP and the application of these algorithms to real cancer sequencing data. PMID:23282028

  11. Nucleotide sequences encoding a thermostable alkaline protease

    DOEpatents

    Wilson, David B.; Lao, Guifang

    1998-01-01

    Nucleotide sequences, derived from a thermophilic actinomycete microorganism, which encode a thermostable alkaline protease are disclosed. Also disclosed are variants of the nucleotide sequences which encode a polypeptide having thermostable alkaline proteolytic activity. Recombinant thermostable alkaline protease or recombinant polypeptide may be obtained by culturing in a medium a host cell genetically engineered to contain and express a nucleotide sequence according to the present invention, and recovering the recombinant thermostable alkaline protease or recombinant polypeptide from the culture medium.

  12. Nucleotide sequences encoding a thermostable alkaline protease

    DOEpatents

    Wilson, D.B.; Lao, G.

    1998-01-06

    Nucleotide sequences, derived from a thermophilic actinomycete microorganism, which encode a thermostable alkaline protease are disclosed. Also disclosed are variants of the nucleotide sequences which encode a polypeptide having thermostable alkaline proteolytic activity. Recombinant thermostable alkaline protease or recombinant polypeptide may be obtained by culturing in a medium a host cell genetically engineered to contain and express a nucleotide sequence according to the present invention, and recovering the recombinant thermostable alkaline protease or recombinant polypeptide from the culture medium. 3 figs.

  13. Mosaic organization of DNA nucleotides

    NASA Technical Reports Server (NTRS)

    Peng, C. K.; Buldyrev, S. V.; Havlin, S.; Simons, M.; Stanley, H. E.; Goldberger, A. L.

    1994-01-01

    Long-range power-law correlations have been reported recently for DNA sequences containing noncoding regions. We address the question of whether such correlations may be a trivial consequence of the known mosaic structure ("patchiness") of DNA. We analyze two classes of controls consisting of patchy nucleotide sequences generated by different algorithms--one without and one with long-range power-law correlations. Although both types of sequences are highly heterogenous, they are quantitatively distinguishable by an alternative fluctuation analysis method that differentiates local patchiness from long-range correlations. Application of this analysis to selected DNA sequences demonstrates that patchiness is not sufficient to account for long-range correlation properties.

  14. Nucleotide excision repair in humans.

    PubMed

    Spivak, Graciela

    2015-12-01

    The demonstration of DNA damage excision and repair replication by Setlow, Howard-Flanders, Hanawalt and their colleagues in the early 1960s, constituted the discovery of the ubiquitous pathway of nucleotide excision repair (NER). The serial steps in NER are similar in organisms from unicellular bacteria to complex mammals and plants, and involve recognition of lesions, adducts or structures that disrupt the DNA double helix, removal of a short oligonucleotide containing the offending lesion, synthesis of a repair patch copying the opposite undamaged strand, and ligation, to restore the DNA to its original form. The transcription-coupled repair (TCR) subpathway of NER, discovered nearly two decades later, is dedicated to the removal of lesions from the template DNA strands of actively transcribed genes. In this review I will outline the essential factors and complexes involved in NER in humans, and will comment on additional factors and metabolic processes that affect the efficiency of this important process. PMID:26388429

  15. Resistance to ketolide antibiotics by coordinated expression of rRNA methyltransferases in a bacterial producer of natural ketolides

    PubMed Central

    Almutairi, Mashal M.; Park, Sung Ryeol; Rose, Simon; Hansen, Douglas A.; Vázquez-Laslop, Nora; Douthwaite, Stephen; Sherman, David H.; Mankin, Alexander S.

    2015-01-01

    Ketolides are promising new antimicrobials effective against a broad range of Gram-positive pathogens, in part because of the low propensity of these drugs to trigger the expression of resistance genes. A natural ketolide pikromycin and a related compound methymycin are produced by Streptomyces venezuelae strain ATCC 15439. The producer avoids the inhibitory effects of its own antibiotics by expressing two paralogous rRNA methylase genes pikR1 and pikR2 with seemingly redundant functions. We show here that the PikR1 and PikR2 enzymes mono- and dimethylate, respectively, the N6 amino group in 23S rRNA nucleotide A2058. PikR1 monomethylase is constitutively expressed; it confers low resistance at low fitness cost and is required for ketolide-induced activation of pikR2 to attain high-level resistance. The regulatory mechanism controlling pikR2 expression has been evolutionary optimized for preferential activation by ketolide antibiotics. The resistance genes and the induction mechanism remain fully functional when transferred to heterologous bacterial hosts. The anticipated wide use of ketolide antibiotics could promote horizontal transfer of these highly efficient resistance genes to pathogens. Taken together, these findings emphasized the need for surveillance of pikR1/pikR2-based bacterial resistance and the preemptive development of drugs that can remain effective against the ketolide-specific resistance mechanism. PMID:26438831

  16. Mutations in 23S rRNA gene associated with decreased susceptibility to tiamulin and valnemulin in Mycoplasma gallisepticum.

    PubMed

    Li, Bei-Bei; Shen, Jian-Zhong; Cao, Xing-Yuan; Wang, Yang; Dai, Lei; Huang, Si-Yang; Wu, Cong-Ming

    2010-07-01

    Mycoplasma gallisepticum is a major etiological agent of chronic respiratory disease (CRD) in chickens and sinusitis in turkeys. The pleuromutilin antibiotics tiamulin and valnemulin are currently used in the treatment of M. gallisepticum infection. We studied the in vitro development of pleuromutilin resistance in M. gallisepticum and investigated the molecular mechanisms involved in this process. Pleuromutilin-resistant mutants were selected by serial passages of M. gallisepticum strains PG31 and S6 in broth medium containing subinhibitory concentrations of tiamulin or valnemulin. A portion of the gene encoding 23S rRNA gene (domain V) and the gene encoding ribosome protein L3 were amplified and sequenced. No mutation could be detected in ribosome protein L3. Mutations were found at nucleotide positions 2058, 2059, 2061, 2447 and 2503 of 23S rRNA gene (Escherichia coli numbering). Although a single mutation could cause elevation of tiamulin and valnemulin MICs, combinations of two or three mutations were necessary to produce high-level resistance. All the mutants were cross-resistant to lincomycin, chloramphenicol and florfenicol. Mutants with the A2058G or the A2059G mutation exhibited cross-resistance to macrolide antibiotics erythromycin, tilmicosin and tylosin. PMID:20487023

  17. Mitochondrial 12S rRNA variants in 1642 Han Chinese pediatric subjects with aminoglycoside-induced and nonsyndromic hearing loss

    PubMed Central

    Lu, Jianxin; Li, Zhiyuan; Zhu, Yi; Yang, Aifen; Li, Ronghua; Zheng, Jing; Cai, Qin; Peng, Guanghua; Zheng, Wuwei; Tang, Xiaowen; Chen, Bobei; Chen, Jianfu; Liao, Zhisu; Yang, Li; Li, Yongyan; You, Junyan; Ding, Yu; Yu, Hong; Wang, Jindan; Sun, Dongmei; Zhao, Jianyue; Xue, Ling; Wang, Jieying; Guan, Min-Xin

    2010-01-01

    In this report, we investigated the frequency and spectrum of mitochondrial 12S rRNA variants in a large cohort of 1642 Han Chinese pediatric subjects with aminoglycoside-induced and nonsyndromic hearing loss. Mutational analysis of 12S rRNA gene in these subjects identified 68 (54 known and 14 novel) variants. The frequencies of known 1555A>G and 1494C>T mutations were 3.96% and 0.18%, respectively, in this cohort with nonsyndromic and aminoglycoside-induced hearing loss. Prevalence of other putative deafness-associated mutation at positions 1095 and 961 were 0.61% and 1.7% in this cohort, respectively. Furthermore, the 745A>G, 792C>T, 801A>G, 839A>G, 856A>G, 1027A>G, 1192C>T, 1192C>A, 1310C>T, 1331A>G, 1374A>G and 1452T>C variants conferred increased sensitivity to ototoxic drugs or nonsyndromic deafness as they were absent in 449 Chinese controls and localized at highly conserved nucleotides of this rRNA. However, other variants appeared to be polymorphisms. Moreover, 65 Chinese subjects carrying the 1555A>G mutation exhibited bilateral and sensorineural hearing loss. A wide range of severity, age-of-onset and audiometric configuration was observed among these subjects. In particular, the sloping and flat shaped patterns were the common audiograms in individuals carrying the 1555A>G mutation. The phenotypic variability in subjects carrying these 12S rRNA mutations indicated the involvement of nuclear modifier genes, mitochondrial haplotypes, epigenetic and environmental factors in the phenotypic manifestation of these mutations. Therefore, our data demonstrated that mitochondrial 12S rRNA is the hot spot for mutations associated with aminoglycoside ototoxicity. PMID:20100600

  18. Extremely Acidophilic Protists from Acid Mine Drainage Host Rickettsiales-Lineage Endosymbionts That Have Intervening Sequences in Their 16S rRNA Genes

    PubMed Central

    Baker, Brett J.; Hugenholtz, Philip; Dawson, Scott C.; Banfield, Jillian F.

    2003-01-01

    During a molecular phylogenetic survey of extremely acidic (pH < 1), metal-rich acid mine drainage habitats in the Richmond Mine at Iron Mountain, Calif., we detected 16S rRNA gene sequences of a novel bacterial group belonging to the order Rickettsiales in the Alphaproteobacteria. The closest known relatives of this group (92% 16S rRNA gene sequence identity) are endosymbionts of the protist Acanthamoeba. Oligonucleotide 16S rRNA probes were designed and used to observe members of this group within acidophilic protists. To improve visualization of eukaryotic populations in the acid mine drainage samples, broad-specificity probes for eukaryotes were redesigned and combined to highlight this component of the acid mine drainage community. Approximately 4% of protists in the acid mine drainage samples contained endosymbionts. Measurements of internal pH of the protists showed that their cytosol is close to neutral, indicating that the endosymbionts may be neutrophilic. The endosymbionts had a conserved 273-nucleotide intervening sequence (IVS) in variable region V1 of their 16S rRNA genes. The IVS does not match any sequence in current databases, but the predicted secondary structure forms well-defined stem loops. IVSs are uncommon in rRNA genes and appear to be confined to bacteria living in close association with eukaryotes. Based on the phylogenetic novelty of the endosymbiont sequences and initial culture-independent characterization, we propose the name “Candidatus Captivus acidiprotistae.” To our knowledge, this is the first report of an endosymbiotic relationship in an extremely acidic habitat. PMID:12957940

  19. Leuconostoc pseudomesenteroides WCFur3 partial 16S rRNA gene

    Technology Transfer Automated Retrieval System (TEKTRAN)

    This study used a partial 535 base pair 16S rRNA gene sequence to identify a bacterial isolate. Fatty acid profiles are consistent with the 16S rRNA gene sequence identification of this bacterium. The isolate was obtained from a compost bin in Fort Collins, Colorado, USA. The 16S rRNA gene sequen...

  20. The Era GTPase recognizes the GAUCACCUCC sequence and binds helix 45 near the 3; end of 16S rRNA

    SciTech Connect

    Tu, Chao; Zhou, Xiaomei; Tarasov, Sergey G.; Tropea, Joseph E.; Austin, Brian P.; Waugh, David S.; Court, Donald L.; Ji, Xinhua

    2012-03-26

    Era, composed of a GTPase domain and a K homology domain, is essential for bacterial cell viability. It is required for the maturation of 16S rRNA and assembly of the 30S ribosomal subunit. We showed previously that the protein recognizes nine nucleotides (1531{sup AUCACCUCC}1539) near the 3{prime} end of 16S rRNA, and that this recognition stimulates GTP-hydrolyzing activity of Era. In all three kingdoms of life, the 1530{sup GAUCA}1534 sequence and helix 45 (h45) (nucleotides 1506-1529) are highly conserved. It has been shown that the 1530{sup GA}1531 to 1530{sup AG}1531 double mutation severely affects the viability of bacteria. However, whether Era interacts with G1530 and/or h45 and whether such interactions (if any) contribute to the stimulation of Era's GTPase activity were not known. Here, we report two RNA structures that contain nucleotides 1506-1542 (RNA301), one in complex with Era and GDPNP (GNP), a nonhydrolysable GTP-analogue, and the other in complex with Era, GNP, and the KsgA methyltransferase. The structures show that Era recognizes 10 nucleotides, including G1530, and that Era also binds h45. Moreover, GTPase assay experiments show that G1530 does not stimulate Era's GTPase activity. Rather, A1531 and A1534 are most important for stimulation and h45 further contributes to the stimulation. Although G1530 does not contribute to the intrinsic GTPase activity of Era, its interaction with Era is important for binding and is essential for the protein to function, leading to the discovery of a new cold-sensitive phenotype of Era.

  1. Single nucleotide markers of D-loop for identification of Indian wild pig (Sus scrofa cristatus)

    PubMed Central

    Srivastava, Gaurav Kumar; Rajput, Nidhi; Jadav, Kajal Kumar; Shrivastav, Avadh Bihari; Joshi, Himanshu R.

    2015-01-01

    Aim: Partial fragment of D-loop region extending from 35 to 770 were compared with corresponding sequences of 16 wild pigs and 9 domestic pig breeds from different parts of the world for detection of single nucleotide polymorphism (SNP) markers in the region. The paper also reappraises SNP markers from two fragments of cytochrome b gene and a fragment 12S rRNA gene distinguishing the Indian wild pig from other pig species of the world. Materials and Methods: Deoxyribonucleic acid (DNA) was isolated from 14 and 12 tissue samples of Indian wild and domestic pigs, respectively, collected from Central India for characterization of the D-loop DNA sequences using universal primers. The sequences obtained were aligned along with the retrieved sequences to analyze species-specific SNP marker. Results: A total of 58 mitochondrial D-loop gene sequences of pig races were aligned to identify 1349 polymorphic sites in the fragment from nucleotide positions 35-770 bp and four SNP markers were identified to differentiate Indian wild pig from all the sequences investigated in this study. With the inclusion of cytochrome b gene and 12S rRNA gene fragments, the present study contributes to the total 15 SNP markers, which have been identified in the mitochondrial fragment of 1936 bp for identification of Indian wild pig. Conclusion: SNP markers have advantages over other marker types and do not require subsequent standardization to compare data across studies or laboratories. PMID:27047129

  2. Phylogenetic analysis of oryx species using partial sequences of mitochondrial rRNA genes.

    PubMed

    Khan, H A; Arif, I A; Al Farhan, A H; Al Homaidan, A A

    2008-01-01

    We conducted a comparative evaluation of 12S rRNA and 16S rRNA genes of the mitochondrial genome for molecular differentiation among three oryx species (Oryx leucoryx, Oryx dammah and Oryx gazella) with respect to two closely related outgroups, addax and roan. Our findings showed the failure of 12S rRNA gene to differentiate between the genus Oryx and addax, whereas a 342-bp partial sequence of 16S rRNA accurately grouped all five taxa studied, suggesting the utility of 16S rRNA segment for molecular phylogeny of oryx at the genus and possibly species levels. PMID:19048493

  3. Specific multiplex analysis of pathogens using a direct 16S rRNA hybridization in microarray system.

    PubMed

    Hwang, Byeong Hee; Shin, Hwa Hui; Seo, Jeong Hyun; Cha, Hyung Joon

    2012-06-01

    For the rapid multiplex analysis of pathogens, 16S rRNAs from cell lysates were directly applied onto a DNA microarray at room temperature (RT) for RNA-DNA hybridization. To eliminate the labeling step, seven fluorescent-labeled detector probes were cohybridized with 16S rRNA targets and adjacent specific capture probes. We found that eight pathogens were successfully discriminated by the 16S rRNA-based direct method, which showed greater specificity than the polymerase chain reaction (PCR)-labeled method due to chaperone and distance effects. A new specificity criterion for a perfect match between RNA and DNA was suggested to be 21-41% dissimilarity using correlation analysis between the mismatch and the sequence according to the guanine-cytosine (GC) percentage or the distribution of mismatches. Six categories of food matrix (egg, meat, milk, rice, vegetable, and mixed) were also tested, and the target pathogen was successfully discriminated within statistically significant levels. Finally, we found that the intrinsic abundance of 16S rRNA molecules successfully substituted PCR-based amplification with a low limit of detection of 10-10(3) cells mL(-1) and a high quantitative linear correlation. Collectively, our suggested 16S rRNA-based direct method enables the highly sensitive, specific, and quantitative analysis of selected pathogens at RT within 2 h, even in food samples. PMID:22551354

  4. Exchange coupling between laterally adjacent nanomagnets.

    PubMed

    Dey, H; Csaba, G; Bernstein, G H; Porod, W

    2016-09-30

    We experimentally demonstrate exchange-coupling between laterally adjacent nanomagnets. Our results show that two neighboring nanomagnets that are each antiferromagnetically exchange-coupled to a common ferromagnetic bottom layer can be brought into strong ferromagnetic interaction. Simulations show that interlayer exchange coupling effectively promotes ferromagnetic alignment between the two nanomagnets, as opposed to antiferromagnetic alignment due to dipole-coupling. In order to experimentally demonstrate the proposed scheme, we fabricated arrays of pairs of elongated, single-domain nanomagnets. Magnetic force microscopy measurements show that most of the pairs are ferromagnetically ordered. The results are in agreement with micromagnetic simulations. The presented scheme can achieve coupling strengths that are significantly stronger than dipole coupling, potentially enabling far-reaching applications in Nanomagnet Logic, spin-wave devices and three-dimensional storage and computing. PMID:27535227

  5. Boundary Layers of Air Adjacent to Cylinders

    PubMed Central

    Nobel, Park S.

    1974-01-01

    Using existing heat transfer data, a relatively simple expression was developed for estimating the effective thickness of the boundary layer of air surrounding cylinders. For wind velocities from 10 to 1000 cm/second, the calculated boundary-layer thickness agreed with that determined for water vapor diffusion from a moistened cylindrical surface 2 cm in diameter. It correctly predicted the resistance for water vapor movement across the boundary layers adjacent to the (cylindrical) inflorescence stems of Xanthorrhoea australis R. Br. and Scirpus validus Vahl and the leaves of Allium cepa L. The boundary-layer thickness decreased as the turbulence intensity increased. For a turbulence intensity representative of field conditions (0.5) and for νwindd between 200 and 30,000 cm2/second (where νwind is the mean wind velocity and d is the cylinder diameter), the effective boundary-layer thickness in centimeters was equal to [Formula: see text]. PMID:16658855

  6. Seismicity in Azerbaijan and Adjacent Caspian Sea

    SciTech Connect

    Panahi, Behrouz M.

    2006-03-23

    So far no general view on the geodynamic evolution of the Black Sea to the Caspian Sea region is elaborated. This is associated with the geological and structural complexities of the region revealed by geophysical, geochemical, petrologic, structural, and other studies. A clash of opinions on geodynamic conditions of the Caucasus region, sometimes mutually exclusive, can be explained by a simplified interpretation of the seismic data. In this paper I analyze available data on earthquake occurrences in Azerbaijan and the adjacent Caspian Sea region. The results of the analysis of macroseismic and instrumental data, seismic regime, and earthquake reoccurrence indicate that a level of seismicity in the region is moderate, and seismic event are concentrated in the shallow part of the lithosphere. Seismicity is mostly intra-plate, and spatial distribution of earthquake epicenters does not correlate with the plate boundaries.

  7. Determining Fungi rRNA Copy Number by PCR

    PubMed Central

    Black, Jonathan; Dean, Timothy; Byfield, Grace; Foarde, Karin; Menetrez, Marc

    2013-01-01

    The goal of this project is to improve the quantification of indoor fungal pollutants via the specific application of quantitative PCR (qPCR). Improvement will be made in the controls used in current qPCR applications. This work focuses on the use of two separate controls within a standard qPCR reaction. The first control developed was the internal standard control gene, benA. This gene encodes for β-tubulin and was selected based on its single-copy nature. The second control developed was the standard control plasmid, which contained a fragment of the ribosomal RNA (rRNA) gene and produced a specific PCR product. The results confirm the multicopy nature of the rRNA region in several filamentous fungi and show that we can quantify fungi of unknown genome size over a range of spore extractions by inclusion of these two standard controls. Advances in qPCR have led to extremely sensitive and quantitative methods for single-copy genes; however, it has not been well established that the rRNA can be used to quantitate fungal contamination. We report on the use of qPCR, combined with two controls, to identify and quantify indoor fungal contaminants with a greater degree of confidence than has been achieved previously. Advances in indoor environmental health have demonstrated that contamination of the built environment by the filamentous fungi has adverse impacts on the health of building occupants. This study meets the need for more accurate and reliable methods for fungal identification and quantitation in the indoor environment. PMID:23543828

  8. Long-range correlations in nucleotide sequences

    NASA Astrophysics Data System (ADS)

    Peng, C.-K.; Buldyrev, S. V.; Goldberger, A. L.; Havlin, S.; Sciortino, F.; Simons, M.; Stanley, H. E.

    1992-03-01

    DNA SEQUENCES have been analysed using models, such as an it-step Markov chain, that incorporate the possibility of short-range nucleotide correlations1. We propose here a method for studying the stochastic properties of nucleotide sequences by constructing a 1:1 map of the nucleotide sequence onto a walk, which we term a 'DNA walk'. We then use the mapping to provide a quantitative measure of the correlation between nucleotides over long distances along the DNA chain. Thus we uncover in the nucleotide sequence a remarkably long-range power law correlation that implies a new scale-invariant property of DNA. We find such long-range correlations in intron-containing genes and in nontranscribed regulatory DNA sequences, but not in complementary DNA sequences or intron-less genes.

  9. Interactions of aminoglycoside antibiotics with rRNA.

    PubMed

    Trylska, Joanna; Kulik, Marta

    2016-08-15

    Aminoglycoside antibiotics are protein synthesis inhibitors applied to treat infections caused mainly by aerobic Gram-negative bacteria. Due to their adverse side effects they are last resort antibiotics typically used to combat pathogens resistant to other drugs. Aminoglycosides target ribosomes. We describe the interactions of aminoglycoside antibiotics containing a 2-deoxystreptamine (2-DOS) ring with 16S rRNA. We review the computational studies, with a focus on molecular dynamics (MD) simulations performed on RNA models mimicking the 2-DOS aminoglycoside binding site in the small ribosomal subunit. We also briefly discuss thermodynamics of interactions of these aminoglycosides with their 16S RNA target. PMID:27528743

  10. Growth rate regulation of rRNA content of a marine Synechococcus (cyanobacterium) strain

    SciTech Connect

    Binder, B.J.; Liu, Y.C.

    1998-09-01

    The relationship between growth rate and rRNA content in a marine Synechococcus strain was examined. A combination of flow cytometry and whole-cell hybridization with fluorescently labeled 16S rRNA-targeted oligonucleotide probes was used to measure the rRNA content of Synechococcus strain WH8101 cells grown at a range of light-limited growth rates. The sensitivity of this approach was sufficient for the analysis of rRNA even in very slowly growing Synechococcus cells. The relationship between growth rate and cellular rRNA content comprised three phases: (1) at low growth rates, rRNA cell{sup {minus}1} remained approximately constant; (2) at intermediate rates, rRNA cell{sup {minus}1} increased proportionally with growth rate; and (3) at the highest, light-saturated rates, rRNA cell{sup {minus}1} dropped abruptly. Total cellular RNA was well correlated with the probe-based measure of rRNA and varied in a similar manner with growth rate. Mean cell volume and rRNA concentration were related to growth rate in a manner similar to rRNA cell{sup {minus}1}, although the overall magnitude linear increase in ribosome efficiency with increasing growth rate, which is consistent with the prevailing prokaryotic model at low growth rates. Taken together, these results support the notion that measurements of cellular rRNA content might be useful for estimating in situ growth rates in natural Synechococcus populations.

  11. 33 CFR 80.1395 - Puget Sound and adjacent waters.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 33 Navigation and Navigable Waters 1 2010-07-01 2010-07-01 false Puget Sound and adjacent waters... INTERNATIONAL NAVIGATION RULES COLREGS DEMARCATION LINES Thirteenth District § 80.1395 Puget Sound and adjacent waters. The 72 COLREGS shall apply on all waters of Puget Sound and adjacent waters, including Lake...

  12. 33 CFR 80.1395 - Puget Sound and adjacent waters.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 33 Navigation and Navigable Waters 1 2013-07-01 2013-07-01 false Puget Sound and adjacent waters... INTERNATIONAL NAVIGATION RULES COLREGS DEMARCATION LINES Thirteenth District § 80.1395 Puget Sound and adjacent waters. The 72 COLREGS shall apply on all waters of Puget Sound and adjacent waters, including Lake...

  13. 33 CFR 80.1395 - Puget Sound and adjacent waters.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 33 Navigation and Navigable Waters 1 2011-07-01 2011-07-01 false Puget Sound and adjacent waters... INTERNATIONAL NAVIGATION RULES COLREGS DEMARCATION LINES Thirteenth District § 80.1395 Puget Sound and adjacent waters. The 72 COLREGS shall apply on all waters of Puget Sound and adjacent waters, including Lake...

  14. 33 CFR 80.1395 - Puget Sound and adjacent waters.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 33 Navigation and Navigable Waters 1 2014-07-01 2014-07-01 false Puget Sound and adjacent waters... INTERNATIONAL NAVIGATION RULES COLREGS DEMARCATION LINES Thirteenth District § 80.1395 Puget Sound and adjacent waters. The 72 COLREGS shall apply on all waters of Puget Sound and adjacent waters, including Lake...

  15. 33 CFR 80.1395 - Puget Sound and adjacent waters.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 33 Navigation and Navigable Waters 1 2012-07-01 2012-07-01 false Puget Sound and adjacent waters... INTERNATIONAL NAVIGATION RULES COLREGS DEMARCATION LINES Thirteenth District § 80.1395 Puget Sound and adjacent waters. The 72 COLREGS shall apply on all waters of Puget Sound and adjacent waters, including Lake...

  16. 30 CFR 56.9103 - Clearance on adjacent tracks.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 30 Mineral Resources 1 2010-07-01 2010-07-01 false Clearance on adjacent tracks. 56.9103 Section..., Hauling, and Dumping Traffic Safety § 56.9103 Clearance on adjacent tracks. Railcars shall not be left on side tracks unless clearance is provided for traffic on adjacent tracks....

  17. 30 CFR 57.9103 - Clearance on adjacent tracks.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 30 Mineral Resources 1 2011-07-01 2011-07-01 false Clearance on adjacent tracks. 57.9103 Section..., Hauling, and Dumping Traffic Safety § 57.9103 Clearance on adjacent tracks. Railcars shall not be left on side tracks unless clearance is provided for traffic on adjacent tracks....

  18. 30 CFR 56.9103 - Clearance on adjacent tracks.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 30 Mineral Resources 1 2011-07-01 2011-07-01 false Clearance on adjacent tracks. 56.9103 Section..., Hauling, and Dumping Traffic Safety § 56.9103 Clearance on adjacent tracks. Railcars shall not be left on side tracks unless clearance is provided for traffic on adjacent tracks....

  19. 30 CFR 57.9103 - Clearance on adjacent tracks.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 30 Mineral Resources 1 2010-07-01 2010-07-01 false Clearance on adjacent tracks. 57.9103 Section..., Hauling, and Dumping Traffic Safety § 57.9103 Clearance on adjacent tracks. Railcars shall not be left on side tracks unless clearance is provided for traffic on adjacent tracks....

  20. ATP half-sites in RadA and RAD51 recombinases bind nucleotides.

    PubMed

    Marsh, May E; Scott, Duncan E; Ehebauer, Matthias T; Abell, Chris; Blundell, Tom L; Hyvönen, Marko

    2016-05-01

    Homologous recombination is essential for repair of DNA double-strand breaks. Central to this process is a family of recombinases, including archeal RadA and human RAD51, which form nucleoprotein filaments on damaged single-stranded DNA ends and facilitate their ATP-dependent repair. ATP binding and hydrolysis are dependent on the formation of a nucleoprotein filament comprising RadA/RAD51 and single-stranded DNA, with ATP bound between adjacent protomers. We demonstrate that truncated, monomeric Pyrococcus furiosus RadA and monomerised human RAD51 retain the ability to bind ATP and other nucleotides with high affinity. We present crystal structures of both apo and nucleotide-bound forms of monomeric RadA. These structures reveal that while phosphate groups are tightly bound, RadA presents a shallow, poorly defined binding surface for the nitrogenous bases of nucleotides. We suggest that RadA monomers would be constitutively bound to nucleotides in the cell and that the bound nucleotide might play a structural role in filament assembly. PMID:27419043

  1. Examination of ClpB Quaternary Structure and Linkage to Nucleotide Binding.

    PubMed

    Lin, JiaBei; Lucius, Aaron L

    2016-03-29

    Escherichia coli caseinolytic peptidase B (ClpB) is a molecular chaperone with the unique ability to catalyze protein disaggregation in collaboration with the KJE system of chaperones. Like many AAA+ molecular motors, ClpB assembles into hexameric rings, and this reaction is thermodynamically linked to nucleotide binding. Here we show that ClpB exists in a dynamic equilibrium of monomers, dimers, tetramers, and hexamers in the presence of both limiting and excess ATPγS. We find that ClpB monomer is only able to bind one nucleotide, whereas all 12 sites in the hexameric ring are bound by nucleotide at saturating concentrations. Interestingly, dimers and tetramers exhibit stoichiometries of ∼3 and 7, respectively, which is one fewer than the maximum number of binding sites in the formed oligomer. This observation suggests an open conformation for the intermediates based on the need for an adjacent monomer to fully form the binding pocket. We also report the protein-protein interaction constants for dimers, tetramers, and hexamers and their dependencies on nucleotide. These interaction constants make it possible to predict the concentration of hexamers present and able to bind to cochaperones and polypeptide substrates. Such information is essential for the interpretation of many in vitro studies. Finally, the strategies presented here are broadly applicable to a large number of AAA+ molecular motors that assemble upon nucleotide binding and interact with partner proteins. PMID:26891079

  2. Characterising the Canine Oral Microbiome by Direct Sequencing of Reverse-Transcribed rRNA Molecules

    PubMed Central

    McDonald, James E.; Larsen, Niels; Pennington, Andrea; Connolly, John; Wallis, Corrin; Rooks, David J.; Hall, Neil; McCarthy, Alan J.; Allison, Heather E.

    2016-01-01

    PCR amplification and sequencing of phylogenetic markers, primarily Small Sub-Unit ribosomal RNA (SSU rRNA) genes, has been the paradigm for defining the taxonomic composition of microbiomes. However, ‘universal’ SSU rRNA gene PCR primer sets are likely to miss much of the diversity therein. We sequenced a library comprising purified and reverse-transcribed SSU rRNA (RT-SSU rRNA) molecules from the canine oral microbiome and compared it to a general bacterial 16S rRNA gene PCR amplicon library generated from the same biological sample. In addition, we have developed BIONmeta, a novel, open-source, computer package for the processing and taxonomic classification of the randomly fragmented RT-SSU rRNA reads produced. Direct RT-SSU rRNA sequencing revealed that 16S rRNA molecules belonging to the bacterial phyla Actinobacteria, Bacteroidetes, Firmicutes, Proteobacteria and Spirochaetes, were most abundant in the canine oral microbiome (92.5% of total bacterial SSU rRNA). The direct rRNA sequencing approach detected greater taxonomic diversity (1 additional phylum, 2 classes, 1 order, 10 families and 61 genera) when compared with general bacterial 16S rRNA amplicons from the same sample, simultaneously provided SSU rRNA gene inventories of Bacteria, Archaea and Eukarya, and detected significant numbers of sequences not recognised by ‘universal’ primer sets. Proteobacteria and Spirochaetes were found to be under-represented by PCR-based analysis of the microbiome, and this was due to primer mismatches and taxon-specific variations in amplification efficiency, validated by qPCR analysis of 16S rRNA amplicons from a mock community. This demonstrated the veracity of direct RT-SSU rRNA sequencing for molecular microbial ecology. PMID:27276347

  3. Characterising the Canine Oral Microbiome by Direct Sequencing of Reverse-Transcribed rRNA Molecules.

    PubMed

    McDonald, James E; Larsen, Niels; Pennington, Andrea; Connolly, John; Wallis, Corrin; Rooks, David J; Hall, Neil; McCarthy, Alan J; Allison, Heather E

    2016-01-01

    PCR amplification and sequencing of phylogenetic markers, primarily Small Sub-Unit ribosomal RNA (SSU rRNA) genes, has been the paradigm for defining the taxonomic composition of microbiomes. However, 'universal' SSU rRNA gene PCR primer sets are likely to miss much of the diversity therein. We sequenced a library comprising purified and reverse-transcribed SSU rRNA (RT-SSU rRNA) molecules from the canine oral microbiome and compared it to a general bacterial 16S rRNA gene PCR amplicon library generated from the same biological sample. In addition, we have developed BIONmeta, a novel, open-source, computer package for the processing and taxonomic classification of the randomly fragmented RT-SSU rRNA reads produced. Direct RT-SSU rRNA sequencing revealed that 16S rRNA molecules belonging to the bacterial phyla Actinobacteria, Bacteroidetes, Firmicutes, Proteobacteria and Spirochaetes, were most abundant in the canine oral microbiome (92.5% of total bacterial SSU rRNA). The direct rRNA sequencing approach detected greater taxonomic diversity (1 additional phylum, 2 classes, 1 order, 10 families and 61 genera) when compared with general bacterial 16S rRNA amplicons from the same sample, simultaneously provided SSU rRNA gene inventories of Bacteria, Archaea and Eukarya, and detected significant numbers of sequences not recognised by 'universal' primer sets. Proteobacteria and Spirochaetes were found to be under-represented by PCR-based analysis of the microbiome, and this was due to primer mismatches and taxon-specific variations in amplification efficiency, validated by qPCR analysis of 16S rRNA amplicons from a mock community. This demonstrated the veracity of direct RT-SSU rRNA sequencing for molecular microbial ecology. PMID:27276347

  4. HCV IRES interacts with the 18S rRNA to activate the 40S ribosome for subsequent steps of translation initiation

    PubMed Central

    Malygin, Alexey A.; Kossinova, Olga A.; Shatsky, Ivan N.; Karpova, Galina G.

    2013-01-01

    Previous analyses of complexes of 40S ribosomal subunits with the hepatitis C virus (HCV) internal ribosome entry site (IRES) have revealed contacts made by the IRES with ribosomal proteins. Here, using chemical probing, we show that the HCV IRES also contacts the backbone and bases of the CCC triplet in the 18S ribosomal RNA (rRNA) expansion segment 7. These contacts presumably provide interplay between IRES domain II and the AUG codon close to ribosomal protein S5, which causes a rearrangement of 18S rRNA structure in the vicinity of the universally conserved nucleotide G1639. As a result, G1639 becomes exposed and the corresponding site of the 40S subunit implicated in transfer RNA discrimination can select . These data are the first demonstration at nucleotide resolution of direct IRES–rRNA interactions and how they induce conformational transition in the 40S subunit allowing the HCV IRES to function without AUG recognition initiation factors. PMID:23873958

  5. The rRNA evolution and procaryotic phylogeny

    NASA Technical Reports Server (NTRS)

    Fox, G. E.

    1986-01-01

    Studies of ribosomal RNA primary structure allow reconstruction of phylogenetic trees for prokaryotic organisms. Such studies reveal major dichotomy among the bacteria that separates them into eubacteria and archaebacteria. Both groupings are further segmented into several major divisions. The results obtained from 5S rRNA sequences are essentially the same as those obtained with the 16S rRNA data. In the case of Gram negative bacteria the ribosomal RNA sequencing results can also be directly compared with hybridization studies and cytochrome c sequencing studies. There is again excellent agreement among the several methods. It seems likely then that the overall picture of microbial phylogeny that is emerging from the RNA sequence studies is a good approximation of the true history of these organisms. The RNA data allow examination of the evolutionary process in a semi-quantitative way. The secondary structures of these RNAs are largely established. As a result it is possible to recognize examples of local structural evolution. Evolutionary pathways accounting for these events can be proposed and their probability can be assessed.

  6. Higher-order structure of rRNA

    NASA Technical Reports Server (NTRS)

    Gutell, R. R.; Woese, C. R.

    1986-01-01

    A comparative search for phylogenetically covarying basepair replacements within potential helices has been the only reliable method to determine the correct secondary structure of the 3 rRNAs, 5S, 16S, and 23S. The analysis of 16S from a wide phylogenetic spectrum, that includes various branches of the eubacteria, archaebacteria, eucaryotes, in addition to the mitochondria and chloroplast, is beginning to reveal the constraints on the secondary structures of these rRNAs. Based on the success of this analysis, and the assumption that higher order structure will also be phylogenetically conserved, a comparative search was initiated for positions that show co-variation not involved in secondary structure helices. From a list of potential higher order interactions within 16S rRNA, two higher-order interactions are presented. The first of these interactions involves positions 570 and 866. Based on the extent of phylogenetic covariation between these positions while maintaining Watson-Crick pairing, this higher-order interaction is considered proven. The other interaction involves a minimum of six positions between the 1400 and 1500 regions of the 16S rRNA. Although these patterns of covariation are not as striking as the 570/866 interaction, the fact that they all exist in an anti-parallel fashion and that experimental methods previously implicated these two regions of the molecule in tRNA function suggests that these interactions be given serious consideration.

  7. Chromosomal Organization of Rrna Operons in Bacillus Subtilis

    PubMed Central

    Jarvis, E. D.; Widom, R. L.; LaFauci, G.; Setoguchi, Y.; Richter, I. R.; Rudner, R.

    1988-01-01

    Integrative mapping with vectors containing ribosomal DNA sequences were used to complete the mapping of the 10 rRNA gene sets in the endospore forming bacterium Bacillus subtilis. Southern hybridizations allowed the assignment of nine operons to distinct BclI restriction fragments and their genetic locus identified by transductional crosses. Nine of the ten rRNA gene sets are located between 0 and 70° on the genomic map. In the region surrounding cysA14, two sets of closely spaced tandem clusters are present. The first (rrnJ and rrnW) is located between purA16 and cysA14 closely linked to the latter; the second (rrnI, rrnH and rrnG) previously mapped within this area is located between attSPO2 and glpT6. The operons at or near the origin of replication (rrnO,rrnA and rrnJ,rrnW) represent ``hot spots'' of plasmid insertion. PMID:2465199

  8. Sulfur-oxidizing bacterial endosymbionts: analysis of phylogeny and specificity by 16S rRNA sequences. [Calyptogena magnifica; Bathymodiolus thermophilus; Lucinoma annulata; Lucinoma aequizonata; Codakia orbicularis

    SciTech Connect

    Distel, D.L.; Lane, D.J.; Olsen, G.J.; Giovannoni, S.J.; Pace, B.; Pace, N.R.; Stahl, D.A.; Felbeck, H.

    1988-06-01

    The 16S rRNAs from the bacterial endosymbionts of six marine invertebrates from diverse environments were isolated and partially sequenced. These symbionts included the trophosome symbiont of Riftia pachyptila, the gill symbionts of Calyptogena magnifica and Bathymodiolus thermophilus (from deep-sea hydrothermal vents), and the gill symbionts of Lucinoma annulata, Lucinoma aequizonata, and Codakia orbicularis (from relatively shallow coastal environments). Only one type of bacterial 16S rRNA was detected in each symbiosis. Using nucleotide sequence comparisons, we showed that each of the bacterial symbionts is distinct from the others and that all fall within a limited domain of the gamma subdivision of the purple bacteria (one of the major eubacterial divisions previously defined by 16S rRNA analysis. Two host specimens were analyzed in five of the symbioses; in each case, identical bacterial rRNA sequences were obtained from conspecific host specimens. These data indicate that the symbioses examined are species specific and that the symbiont species are unique to and invariant within their respective host species.

  9. UV light-induced DNA lesions cause dissociation of yeast RNA polymerases-I and establishment of a specialized chromatin structure at rRNA genes

    PubMed Central

    Tremblay, Maxime; Charton, Romain; Wittner, Manuel; Levasseur, Geneviève; Griesenbeck, Joachim; Conconi, Antonio

    2014-01-01

    The cytotoxicity of UV light-induced DNA lesions results from their interference with transcription and replication. DNA lesions arrest elongating RNA polymerases, an event that triggers transcription-coupled nucleotide excision repair. Since arrested RNA polymerases reduce the accessibility of repair factors to DNA lesions, they might be displaced. The fate of arrested RNA polymerases-II at DNA lesions has been extensively studied, yielding partially contradictory results. Considerably less is known about RNA polymerases-I that transcribe nucleosomes-depleted rRNA genes at very high rate. To investigate the fate of arrested RNA polymerases-I at DNA lesions, chromatin-immunoprecipitation, electron microscopy, transcription run-on, psoralen-cross-linking and chromatin-endogenous cleavage were employed. We found that RNA polymerases-I density increased at the 5′-end of the gene, likely due to continued transcription initiation followed by elongation and pausing/release at the first DNA lesion. Most RNA polymerases-I dissociated downstream of the first DNA lesion, concomitant with chromatin closing that resulted from deposition of nucleosomes. Although nucleosomes were deposited, the high mobility group-box Hmo1 (component of actively transcribed rRNA genes) remained associated. After repair of DNA lesions, Hmo1 containing chromatin might help to restore transcription elongation and reopening of rRNA genes chromatin. PMID:24097442

  10. Nucleotide Salvage Deficiencies, DNA Damage and Neurodegeneration

    PubMed Central

    Fasullo, Michael; Endres, Lauren

    2015-01-01

    Nucleotide balance is critically important not only in replicating cells but also in quiescent cells. This is especially true in the nervous system, where there is a high demand for adenosine triphosphate (ATP) produced from mitochondria. Mitochondria are particularly prone to oxidative stress-associated DNA damage because nucleotide imbalance can lead to mitochondrial depletion due to low replication fidelity. Failure to maintain nucleotide balance due to genetic defects can result in infantile death; however there is great variability in clinical presentation for particular diseases. This review compares genetic diseases that result from defects in specific nucleotide salvage enzymes and a signaling kinase that activates nucleotide salvage after DNA damage exposure. These diseases include Lesch-Nyhan syndrome, mitochondrial depletion syndromes, and ataxia telangiectasia. Although treatment options are available to palliate symptoms of these diseases, there is no cure. The conclusions drawn from this review include the critical role of guanine nucleotides in preventing neurodegeneration, the limitations of animals as disease models, and the need to further understand nucleotide imbalances in treatment regimens. Such knowledge will hopefully guide future studies into clinical therapies for genetic diseases. PMID:25923076

  11. [Study of the heterogeneity of 16s rRNA gene and groESL operone in the dna samples of Anaplasma phagocytophilum, Ehrlichia muris, and "Candidatus Neoehrlichia mikurensis" determined in the Ixodes persulcatus ticks in the area of Urals, Siberia, and far east of Russia].

    PubMed

    Rar, V A; Epikhina, T I; Livanova, N N; Panov, V V; Doroshenko, E K; Pukhovskaia, N M; Vysochina, N P; Ivanov, L I

    2011-01-01

    A total of 3552 Ixodes persulcatus from Sverdlovsk, Chelyabinsk, Novosibirsk, Irkutsk regions and Khabarovsk Territory were examined on the Ehrlichia and Anaplasma presence by nested PCR based on the 16S rRNA gene. Both Anaplasma phagocytophilum and Ehrlichia muris DNA were found in I. persulcatus in all studied regions. A. Phagocytophilum was detected in 1.3-6.3% of ticks and E. muris - in 2.0-14.1% of ticks. Moreover, "Candidatus Neoehrlichia mikurensis" DNA was found in 8 ticks collected in Novosibirsk, Irkutsk Regions and Khabarovsk Territory. Partial nucleotide sequences of 16S rRNA gene and groESL operone (1240-1300 bp) were determined for 65 samples of A. Phagocytophilum, 17 samples of E. muris and 4 samples of "Candidatus Neoehrlichia mikurensis". Nucleotide sequences of 16S rRNA gene and groESL operone of E. muris and "Candidatus Neoehrlichia mikurensis" were shown to be highly conservative, and nucleotide sequences of groESL operone of both E. muris and "Candidatus Neoehrlichia mikurensis" differed from the sequences found previously in other species of Ixodid tick. On the basis of analysis of the 16S rRNA gene and groESL operone sequences it was concluded that all revealed samples A. Phagocytophilum could be divided into 2 groups. GroESL operone sequences of A. Phagocytophilum from the first group were identical to each other but significantly differed from the known groESL operone sequences (less than 98.2% of similarity), whereas their 16S rRNA gene sequences were identical to the sequence of widely distributed and pathogenic for human A. Phagocytophilum genetic variant (CAHU-HGEl, GenBank AF093788) or differed from it by a single nucleotide substitution. The nucleotide sequences of groESL operone of A. Phagocytophilum from the second group differed from each other by 1-4 nucleotides and were closely related (99.2-99.4% of similarity) to the sequences of groESL operone ofA. phagocytophilum isolates found in Europe in Ixodes ricinus and roe deer. The

  12. Interaction between adjacent lightning discharges in clouds

    NASA Astrophysics Data System (ADS)

    Wang, Yanhui; Zhang, Guangshu; Zhang, Tong; Li, Yajun; Wu, Bin; Zhang, Tinglong

    2013-07-01

    Using a 3D lightning radiation source locating system (LLS), three pairs of associated lightning discharges (two or more adjacent lightning discharges following an arbitrary rule that their space-gap was less than 10 km and their time-gap was less than 800 ms) were observed, and the interaction between associated lightning discharges was analyzed. All these three pairs of associated lightning discharges were found to involve three or more charge regions (the ground was considered as a special charge region). Moreover, at least one charge region involved two lightning discharges per pair of associated lightning discharges. Identified from electric field changes, the subsequent lightning discharges were suppressed by the prior lightning discharges. However, it is possible that the prior lightning discharge provided a remaining discharge channel to facilitate the subsequent lightning discharge. The third case provided evidence of this possibility. Together, the results suggested that, if the charges in the main negative charge region can be consumed using artificial lightning above the main negative charge regions, lightning accidents on the ground could be greatly reduced, on the condition that the height of the main negative charge region and the charge intensity of the lower positive charge region are suitable.

  13. The Regulation of rRNA Gene Transcription during Directed Differentiation of Human Embryonic Stem Cells

    PubMed Central

    Liu, Zhong; Zhao, Rui; Giles, Keith E.

    2016-01-01

    It has become increasingly clear that proper cellular control of pluripotency and differentiation is related to the regulation of rRNA synthesis. To further our understanding of the role that the regulation of rRNA synthesis has in pluripotency we monitored rRNA synthesis during the directed differentiation of human embryonic stem cells (hESCs). We discovered that the rRNA synthesis rate is reduced ~50% within 6 hours of ACTIVIN A treatment. This precedes reductions in expression of specific stem cell markers and increases in expression of specific germ layer markers. The reduction in rRNA synthesis is concomitant with dissociation of the Pol I transcription factor, UBTF, from the rRNA gene promoter and precedes any increase to heterochromatin throughout the rRNA gene. To directly investigate the role of rRNA synthesis in pluripotency, hESCs were treated with the Pol I inhibitor, CX-5461. The direct reduction of rRNA synthesis by CX-5461 induces the expression of markers for all three germ layers, reduces the expression of pluripotency markers, and is overall similar to the ACTIVIN A induced changes. This work indicates that the dissociation of UBTF from the rRNA gene, and corresponding reduction in transcription, represent early regulatory events during the directed differentiation of pluripotent stem cells. PMID:27299313

  14. Single Nucleotide Polymorphisms and Osteoarthritis

    PubMed Central

    Wang, Ting; Liang, Yuting; Li, Hong; Li, Haibo; He, Quanze; Xue, Ying; Shen, Cong; Zhang, Chunhua; Xiang, Jingjing; Ding, Jie; Qiao, Longwei; Zheng, Qiping

    2016-01-01

    Abstract Osteoarthritis (OA) is a complex disorder characterized by degenerative articular cartilage and is largely attributed to genetic risk factors. Single nucleotide polymorphisms (SNPs) are common DNA variants that have shown promising and efficiency, compared with positional cloning, to map candidate genes of complex diseases, including OA. In this study, we aim to provide an overview of multiple SNPs from a number of genes that have recently been linked to OA susceptibility. We also performed a comprehensive meta-analysis to evaluate the association of SNP rs7639618 of double von Willebrand factor A domains (DVWA) gene with OA susceptibility. A systematic search of studies on the association of SNPs with susceptibility to OA was conducted in PubMed and Google scholar. Studies subjected to meta-analysis include human and case-control studies that met the Hardy–Weinberg equilibrium model and provide sufficient data to calculate an odds ratio (OR). A total of 9500 OA cases and 9365 controls in 7 case-control studies relating to SNP rs7639618 were included in this study and the ORs with 95% confidence intervals (CIs) were calculated. Over 50 SNPs from different genes have been shown to be associated with either hip (23), or knee (20), or both (13) OA. The ORs of these SNPs for OA and the subtypes are not consistent. As to SNP rs7639618 of DVWA, increased knee OA risk was observed in all genetic models analyzed. Specifically, people from Asian with G-allele showed significantly increased risk of knee OA (A versus G: OR = 1.28, 95% CI 1.13–1.46; AA versus GG: OR = 1.60, 95% CI 1.25–2.05; GA versus GG: OR = 1.31, 95% CI 1.18–1.44; AA versus GA+GG: OR = 1.34, 95% CI 1.12–1.61; AA+GA versus GG: OR = 1.40, 95% CI 1.19–1.64), but not in Caucasians or with hip OA. Our results suggest that multiple SNPs play different roles in the pathogenesis of OA and its subtypes; SNP rs7639618 of DVWA gene is associated with a significantly increased

  15. Advances in targeting cyclic nucleotide phosphodiesterases

    PubMed Central

    Maurice, Donald H.; Ke, Hengming; Ahmad, Faiyaz; Wang, Yousheng; Chung, Jay; Manganiello, Vincent C.

    2014-01-01

    Cyclic nucleotide phosphodiesterases (PDEs) catalyse the hydrolysis of cyclic AMP and cyclic GMP, thereby regulating the intracellular concentrations of these cyclic nucleotides, their signalling pathways and, consequently, myriad biological responses in health and disease. Currently, a small number of PDE inhibitors are used clinically for treating the pathophysiological dysregulation of cyclic nucleotide signalling in several disorders, including erectile dysfunction, pulmonary hypertension, acute refractory cardiac failure, intermittent claudication and chronic obstructive pulmonary disease. However, pharmaceutical interest in PDEs has been reignited by the increasing understanding of the roles of individual PDEs in regulating the subcellular compartmentalization of specific cyclic nucleotide signalling pathways, by the structure-based design of novel specific inhibitors and by the development of more sophisticated strategies to target individual PDE variants. PMID:24687066

  16. Advances in targeting cyclic nucleotide phosphodiesterases.

    PubMed

    Maurice, Donald H; Ke, Hengming; Ahmad, Faiyaz; Wang, Yousheng; Chung, Jay; Manganiello, Vincent C

    2014-04-01

    Cyclic nucleotide phosphodiesterases (PDEs) catalyse the hydrolysis of cyclic AMP and cyclic GMP, thereby regulating the intracellular concentrations of these cyclic nucleotides, their signalling pathways and, consequently, myriad biological responses in health and disease. Currently, a small number of PDE inhibitors are used clinically for treating the pathophysiological dysregulation of cyclic nucleotide signalling in several disorders, including erectile dysfunction, pulmonary hypertension, acute refractory cardiac failure, intermittent claudication and chronic obstructive pulmonary disease. However, pharmaceutical interest in PDEs has been reignited by the increasing understanding of the roles of individual PDEs in regulating the subcellular compartmentalization of specific cyclic nucleotide signalling pathways, by the structure-based design of novel specific inhibitors and by the development of more sophisticated strategies to target individual PDE variants. PMID:24687066

  17. Gut microbiome compositional and functional differences between tumor and non-tumor adjacent tissues from cohorts from the US and Spain

    PubMed Central

    Allali, Imane; Delgado, Susana; Marron, Pablo Isidro; Astudillo, Aurora; Yeh, Jen Jen; Ghazal, Hassan; Amzazi, Saaïd; Keku, Temitope; Azcarate-Peril, M Andrea

    2015-01-01

    Colorectal cancer (CRC) is the third most common cancer in the world and the second leading cause of cancer deaths in the US and Spain. The molecular mechanisms involved in the etiology of CRC are not yet elucidated due in part to the complexity of the human gut microbiota. In this study, we compared the microbiome composition of 90 tumor and matching adjacent tissue (adjacent) from cohorts from the US and Spain by 16S rRNA amplicon sequencing in order to determine the impact of the geographic origin on the CRC microbiome. Data showed a significantly (P < 0.05) higher Phylogenetic Diversity (PD) for the US (PD Adjacent = 26.3 ± 5.3, PD Tumor = 23.3 ± 6.2) compared to the Spanish cohort (PD Adjacent = 18.9 ± 5.9, PD Tumor = 18.7 ± 6.6) while no significant differences in bacterial diversity were observed between tumor and adjacent tissues for individuals from the same country. Adjacent tissues from the Spanish cohort were enriched in Firmicutes (SP = 43.9% and US = 22.2%, P = 0.0001) and Actinobacteria (SP = 1.6% and US = 0.5%, P = 0.0018) compared to US adjacent tissues, while adjacent tissues from the US had significantly higher abundances of Fusobacteria (US = 8.1% and SP = 1.5%, P = 0.0023) and Sinergistetes (US = 0.3% and SP = 0.1%, P = 0.0097). Comparisons between tumor and adjacent tissues in each cohort identified the genus Eikenella significantly over represented in US tumors (T = 0.024% and A = 0%, P = 0.03), and the genera Fusobacterium (T = 10.4% and A = 1.5%, P = <0.0001), Bulleida (T = 0.36% and A = 0.09%, P = 0.02), Gemella (T = 1.46% and A = 0.19%, P = 0.03), Parvimonas (T = 3.14% and A = 0.86%, P = 0.03), Campylobacter (T = 0.15% and A = 0.008%, P = 0.047), and Streptococcus (T = 2.84% and A = 2.19%, P = 0.05) significantly over represented in Spanish tumors. Predicted metagenome functional content from 16S rRNA surveys showed that bacterial motility proteins and proteins involved in flagellar assembly were over represented in adjacent tissues

  18. In vitro incorporation of LNA nucleotides.

    PubMed

    Veedu, Rakesh N; Vester, Birte; Wengel, Jesper

    2007-01-01

    An LNA modified nucleoside triphosphate 1 was synthesized in order to investigate its potential to act as substrate for DNA strand synthesis by polymerases. Primer extension assays for the incorporation experiments revealed that Phusion High Fidelity DNA polymerase is an efficient enzyme for incorporation of the LNA nucleotide and for extending strand to full length. It was also observed that pfu DNA polymerase could incorporate the LNA nucleotide but it failed to extend the strand to a full length product. PMID:18058567

  19. Petroleum basins of Sakhalin and adjacent shelf

    SciTech Connect

    Mavrinski, Y.; Koblov, E. )

    1993-09-01

    Sixty-seven oil and gas fields have been discovered on Sakhalin and the adjacent shelf but the distribution of fields is uneven in north Sakhalin, south Sakhalin, and the Tatar basins. The sedimentary cover is composed of sandy, clayey, and siliceous rocks, with volcanogenic and coal-bearing deposits of Upper Cretaceous, Paleogene, and Neogene 8-12 km thick. Marine clayey and siliceous oil source rocks are regionally developed in the section at different stratigraphic levels; the organic matter is of mixed type and the content varies from 0.5 to 1.5%. The upper Oligocene and middle-upper Miocene source rocks in the north Sakhalin basin are typical, and the organic carbon content ranges from 1 to 5%. The level of organic matter catagenesis and conversion into hydrocarbons is high because of the high differential geothermal gradient in the basins, 30-50[degrees]C per km. Porous sandstones in the Miocene form the reservoirs in all fields with the exception of Okruzhnoye, where the pay zone is a siliceous claystone. Growth-fault rollovers and anticlines form the main traps ranging in area from 5 to 300 km[sup 2], with amplitudes between 100 and 600 m. both stratigraphic and structural traps have been identified. Considerable volumes of reserves are associated with the Miocene deposits of north Sakhalin, which are characterized by an optimum combination of oil source rocks, focused migration paths, and thick sequences of reservoirs and cap rocks. Six large fields have been discovered in the past 15 yr. Oil and condensate reserves stand at over 300 million MT, and gas reserves are about 900 billion m[sup 3].

  20. The impact of rRNA secondary structure consideration in alignment and tree reconstruction: simulated data and a case study on the phylogeny of hexapods.

    PubMed

    Letsch, Harald O; Kück, Patrick; Stocsits, Roman R; Misof, Bernhard

    2010-11-01

    The use of secondary structures has been advocated to improve both the alignment and the tree reconstruction processes of ribosomal RNA (rRNA) data sets. We used simulated and empirical rRNA data to test the impact of secondary structure consideration in both steps of molecular phylogenetic analyses. A simulation approach was used to generate realistic rRNA data sets based on real 16S, 18S, and 28S sequences and structures in combination with different branch length and topologies. Alignment and tree reconstruction performance of four recent structural alignment methods was compared with exclusively sequence-based approaches. As empirical data, we used a hexapod rRNA data set to study the influence of nucleotide interdependencies in sequence alignment and tree reconstruction. Structural alignment methods delivered significantly better sequence alignments compared with pure sequence-based methods. Also, structural alignment methods delivered better trees judged by topological congruence to simulation base trees. However, the advantage of structural alignments was less pronounced and even vanished in several instances. For simulated data, application of mixed RNA/DNA models to stems and loops, respectively, led to significantly shorter branches. The application of mixed RNA/DNA models in the hexapod analyses delivered partly implausible relationships. This can be interpreted as a stronger sensitivity of mixed model setups to nonphylogenetic signal. Secondary structure consideration clearly influenced sequence alignment and tree reconstruction of ribosomal genes. Although sequence alignment quality can considerably be improved by the use of secondary structure information, the application of mixed models in tree reconstructions needs further studies to understand the observed effects. PMID:20530152

  1. Evaluation of the 16S and 12S rRNA genes as universal markers for the identification of commercial fish species in South Africa.

    PubMed

    Cawthorn, Donna-Mareè; Steinman, Harris Andrew; Witthuhn, R Corli

    2012-01-01

    The development of DNA-based methods for the identification of fish species is important for fisheries research and control, as well as for the detection of unintentional or fraudulent species substitutions in the marketplace. The aim of this study was to generate a comprehensive reference database of DNA sequences from the mitochondrial 16S and 12S ribosomal RNA (rRNA) genes for 53 commercial fish species in South Africa and to evaluate the applicability of these genetic markers for the identification of fish at the species level. The DNA extracted from all target species was readily amplified using universal primers targeting both rRNA gene regions. Sequences from the 16S and 12S rRNA genes were submitted to GenBank for the first time for 34% and 53% of the fish species, respectively. Cumulative analysis of the 16S rRNA gene sequences revealed mean conspecific, congeneric and confamilial Kimura two parameter (K2P) distances of 0.03%, 0.70% and 5.10% and the corresponding values at the 12S level were 0.03%, 1.00% and 5.57%. K2P neighbour-joining trees based on both sequence datasets generally clustered species in accordance with their taxonomic classifications. The nucleotide variation in both the 16S and 12S sequences was suitable for identifying the large majority of the examined fish specimens to at least the level of genus, but was found to be less useful for the explicit differentiation of certain congeneric fish species. It is recommended that one or more faster-evolving DNA regions be analysed to confirm the identities of closely-related fish species in South Africa. PMID:21963445

  2. Regulation of mammalian nucleotide metabolism and biosynthesis

    PubMed Central

    Lane, Andrew N.; Fan, Teresa W.-M.

    2015-01-01

    Nucleotides are required for a wide variety of biological processes and are constantly synthesized de novo in all cells. When cells proliferate, increased nucleotide synthesis is necessary for DNA replication and for RNA production to support protein synthesis at different stages of the cell cycle, during which these events are regulated at multiple levels. Therefore the synthesis of the precursor nucleotides is also strongly regulated at multiple levels. Nucleotide synthesis is an energy intensive process that uses multiple metabolic pathways across different cell compartments and several sources of carbon and nitrogen. The processes are regulated at the transcription level by a set of master transcription factors but also at the enzyme level by allosteric regulation and feedback inhibition. Here we review the cellular demands of nucleotide biosynthesis, their metabolic pathways and mechanisms of regulation during the cell cycle. The use of stable isotope tracers for delineating the biosynthetic routes of the multiple intersecting pathways and how these are quantitatively controlled under different conditions is also highlighted. Moreover, the importance of nucleotide synthesis for cell viability is discussed and how this may lead to potential new approaches to drug development in diseases such as cancer. PMID:25628363

  3. Exchange of Spacer Regions between Rrna Operons in Escherichia Coli

    PubMed Central

    Harvey, S.; Hill, C. W.

    1990-01-01

    The Escherichia coli rRNA operons each have one of two types of spacer separating the 16S and 23S coding regions. The spacers of four operons encode tRNA(Glu2) and the other three encode both tRNA(Ile) and tRNA(Ala 1 B). We have prepared a series of mutants in which the spacer region of a particular rrn operon has been replaced by the opposite type. Included among these were a mutant retaining only a single copy of the tRNA(Glu2) spacer (at rrnG) and another retaining only a single copy of the tRNA(Ile)-tRNA(Ala 1 B) spacer (at rrnA). While both mutants grew more slowly than controls, the mutant deficient in tRNA(Glu2) spacers was more severely affected. At a frequency of 6 X 10(-5), these mutants phenotypically reverted to faster growing types by increasing the copy number of the deficient spacer. In most of these phenotypic revertants, the deficient spacer type appeared in a rrn operon which previously contained the surplus type, bringing the ratio of spacer types closer to normal. In a few cases, these spacer changes were accompanied by an inversion of the chromosomal material between the donor and recipient rrn operons. Two examples of inversion of one-half of the E. coli chromosome between rrnG and rrnH were observed. The correlation of spacer change with inversion indicated that, in these particular cases, the change was due to an intrachromatid gene conversion event accompanied by a reciprocal crossover rather than reciprocal exchange between sister chromatids. PMID:2168847

  4. Evolution of small putative group I introns in the SSU rRNA gene locus of Phialophora species

    PubMed Central

    2011-01-01

    Background Group I introns (specifically subgroup IC1) are common in the nuclear ribosomal RNA genes of fungi. While most range in length from more than 200 to nearly 1800 nucleotides (nt) in length, several small putative (or degenerate) group I introns have been described that are between 56 and 81 nt. Although small, previously we demonstrated that the PaSSU intron in the rRNA small subunit gene of Phialophora americana isolate Wang 1046 is capable of in vitro splicing using a standard group I intron pathway, thus qualifying it as a functional ribozyme. Findings Here, we describe eight short putative group I introns, ranging in length from 63 to 75 nt, in the rRNA small subunit genes of Phialophora isolates, a fungal genus that ranges from saprobic to pathogenic on plants and animals. All contain putative pairing regions P1, P7, and P10, as well as a pairing region formed between the middle of the intron and part of the 3' exon. The other pairing regions common in the core of standard group I introns are absent. However, parts of the 3' exon may aid in the stabilization of these small introns. Although the eight putative group I introns were from at least three species of Phialophora, phylogenetic analysis indicated that the eight are monophyletic. They are also monophyletic with the small introns of two lichen-forming fungi, Porpidia crustulata and Arthonia lapidicola. Conclusions The small putative group I introns in Phialophora have common features that may represent group I introns at their minima. They appear to have a single origin as indicated by their monophyly in phylogenetic analyses. PMID:21781325

  5. Comparison of potential diatom 'barcode' genes (the 18S rRNA gene and ITS, COI, rbcL) and their effectiveness in discriminating and determining species taxonomy in the Bacillariophyta.

    PubMed

    Guo, Liliang; Sui, Zhenghong; Zhang, Shu; Ren, Yuanyuan; Liu, Yuan

    2015-04-01

    Diatoms form an enormous group of photoautotrophic micro-eukaryotes and play a crucial role in marine ecology. In this study, we evaluated typical genes to determine whether they were effective at different levels of diatom clustering analysis to assess the potential of these regions for barcoding taxa. Our test genes included nuclear rRNA genes (the nuclear small-subunit rRNA gene and the 5.8S rRNA gene+ITS-2), a mitochondrial gene (cytochrome c-oxidase subunit 1, COI), a chloroplast gene [ribulose-1,5-biphosphate carboxylase/oxygenase large subunit (rbcL)] and the universal plastid amplicon (UPA). Calculated genetic divergence was highest for the internal transcribed spacer (ITS; 5.8S+ITS-2) (p-distance of 1.569, 85.84% parsimony-informative sites) and COI (6.084, 82.14%), followed by the 18S rRNA gene (0.139, 57.69%), rbcL (0.120, 42.01%) and UPA (0.050, 14.97%), which indicated that ITS and COI were highly divergent compared with the other tested genes, and that their nucleotide compositions were variable within the whole group of diatoms. Bayesian inference (BI) analysis showed that the phylogenetic trees generated from each gene clustered diatoms at different phylogenetic levels. The 18S rRNA gene was better than the other genes in clustering higher diatom taxa, and both the 18S rRNA gene and rbcL performed well in clustering some lower taxa. The COI region was able to barcode species of some genera within the Bacillariophyceae. ITS was a potential marker for DNA based-taxonomy and DNA barcoding of Thalassiosirales, while species of Cyclotella, Skeletonema and Stephanodiscus gathered in separate clades, and were paraphyletic with those of Thalassiosira. Finally, UPA was too conserved to serve as a diatom barcode. PMID:25604341

  6. Structural basis for a six nucleotide genetic alphabet.

    PubMed

    Georgiadis, Millie M; Singh, Isha; Kellett, Whitney F; Hoshika, Shuichi; Benner, Steven A; Richards, Nigel G J

    2015-06-01

    Expanded genetic systems are most likely to work with natural enzymes if the added nucleotides pair with geometries that are similar to those displayed by standard duplex DNA. Here, we present crystal structures of 16-mer duplexes showing this to be the case with two nonstandard nucleobases (Z, 6-amino-5-nitro-2(1H)-pyridone and P, 2-amino-imidazo[1,2-a]-1,3,5-triazin-4(8H)one) that were designed to form a Z:P pair with a standard "edge on" Watson-Crick geometry, but joined by rearranged hydrogen bond donor and acceptor groups. One duplex, with four Z:P pairs, was crystallized with a reverse transcriptase host and adopts primarily a B-form. Another contained six consecutive Z:P pairs; it crystallized without a host in an A-form. In both structures, Z:P pairs fit canonical nucleobase hydrogen-bonding parameters and known DNA helical forms. Unique features include stacking of the nitro group on Z with the adjacent nucleobase ring in the A-form duplex. In both B- and A-duplexes, major groove widths for the Z:P pairs are approximately 1 Å wider than those of comparable G:C pairs, perhaps to accommodate the large nitro group on Z. Otherwise, ZP-rich DNA had many of the same properties as CG-rich DNA, a conclusion supported by circular dichroism studies in solution. The ability of standard duplexes to accommodate multiple and consecutive Z:P pairs is consistent with the ability of natural polymerases to biosynthesize those pairs. This, in turn, implies that the GACTZP synthetic genetic system can explore the entire expanded sequence space that additional nucleotides create, a major step forward in this area of synthetic biology. PMID:25961938

  7. Insights into the phylogenetic positions of photosynthetic bacteria obtained from 5S rRNA and 16S rRNA sequence data

    NASA Technical Reports Server (NTRS)

    Fox, G. E.

    1985-01-01

    Comparisons of complete 16S ribosomal ribonucleic acid (rRNA) sequences established that the secondary structure of these molecules is highly conserved. Earlier work with 5S rRNA secondary structure revealed that when structural conservation exists the alignment of sequences is straightforward. The constancy of structure implies minimal functional change. Under these conditions a uniform evolutionary rate can be expected so that conditions are favorable for phylogenetic tree construction.

  8. The complete nucleotide sequence of the mitochondrial genome of the Asian longhorn beetle, Anoplophora glabripennis (Coleoptera: Cerambycidae).

    PubMed

    Fang, Jie; Qian, Lu; Xu, Mei; Yang, Xiaojun; Wang, Baode; An, Yulin

    2016-09-01

    The complete mitochondrial genome of Anoplophora glabripennis has been investigated and analyzed. The genome is a circular molecule of 15,774 bp, containing 13 protein-coding genes (PCGs), 2 rRNA genes, 22 tRNA genes, and an A + T-rich region. The nucleotide composition of the A.glabripennis mitogenome is strongly biased toward A + T nucleotides (78.30%). Nine protein-coding genes and 14 tRNA genes are encoded on the H strand, and the other 4 protein-coding genes and 8 tRNA genes are encoded on the L strand. The arrangement of genes is identical to all know longhorn beetles mitochondrial genomes. PMID:25693709

  9. Ius Chasma Tributary Valleys and Adjacent Plains

    NASA Technical Reports Server (NTRS)

    2006-01-01

    This image covers valley tributaries of Ius Chasma, as well as the plains adjacent to the valleys. Ius Chasma is one of several canyons that make up the Valles Marineris canyon system. Valles Marineris likely formed by extension associated with the growth of the large volcanoes and topographic high of Tharsis to the northwest. As the ground was pulled apart, large and deep gaps resulted in the valleys seen in the top and bottom of this HiRISE image. Ice that was once in the ground could have also melted to create additional removal of material in the formation of the valleys. HiRISE is able to see the rocks along the walls of both these valleys and also impact craters in the image. Rock layers that appear lower down in elevation appear rougher and are shedding boulders. Near the top of the walls and also seen in patches along the smooth plains are brighter layers. These brighter layers are not shedding boulders so they must represent a different kind of rock formed in a different kind of environment than those further down the walls. Because they are highest in elevation, the bright layers are youngest in age. HiRISE is able to see dozens of the bright layers, which are perhaps only a meter in thickness. Darker sand dunes and ripples cover most of the plains and fill the floors of impact craters.

    Image PSP_001351_1715 was taken by the High Resolution Imaging Science Experiment (HiRISE) camera onboard the Mars Reconnaissance Orbiter spacecraft on November 9, 2006. The complete image is centered at -8.3 degrees latitude, 275.4 degrees East longitude. The range to the target site was 254.3 km (158.9 miles). At this distance the image scale ranges from 25.4 cm/pixel (with 1 x 1 binning) to 101.8 cm/pixel (with 4 x 4 binning). The image shown here has been map-projected to 25 cm/pixel and north is up. The image was taken at a local Mars time of 3:32 PM and the scene is illuminated from the west with a solar incidence angle of 59 degrees, thus the sun was about

  10. Proofreading of misincorporated nucleotides in DNA transcription.

    PubMed

    Voliotis, Margaritis; Cohen, Netta; Molina-París, Carmen; Liverpool, Tanniemola B

    2012-06-01

    The accuracy of DNA transcription is crucial for the proper functioning of the cell. Although RNA polymerases demonstrate selectivity for correct nucleotides, additional active mechanisms of transcriptional error correction are required to achieve observed levels of fidelity. Recent experimental findings have shed light on a particular mechanism of transcriptional error correction involving: (i) diffusive translocation of the RNA polymerase along the DNA (backtracking) and (ii) irreversible RNA cleavage. This mechanism achieves preferential cleavage of misincorporated nucleotides by biasing the local rates of translocation. Here, we study how misincorporated nucleotides affect backtracking dynamics and how this effect determines the level of transcriptional fidelity. We consider backtracking as a diffusive process in a periodic, one-dimensional energy landscape, which at a coarse-grained level gives rise to a hopping process between neighboring local minima. We propose a model for how misincorporated nucleotides deform this energy landscape and hence affect the hopping rates. In particular, we show that this model can be used to derive both the theoretical limit on the fidelity (i.e. the minimum fraction of misincorporated nucleotides) and the actual fidelity relative to this optimum, achieved for specific combinations of the cleavage and polymerization rates. Finally, we study how external factors influencing backtracking dynamics affect transcriptional fidelity. We show that biologically relevant loads, similar to those exerted by nucleosomes or other transcriptional barriers, increase error correction. PMID:22643861

  11. Proofreading of misincorporated nucleotides in DNA transcription.

    PubMed

    Voliotis, Margaritis; Cohen, Netta; Molina-París, Carmen; Liverpool, Tanniemola B

    2012-06-01

    The accuracy of DNA transcription is crucial for the proper functioning of the cell. Although RNA polymerases demonstrate selectivity for correct nucleotides, additional active mechanisms of transcriptional error correction are required to achieve observed levels of fidelity. Recent experimental findings have shed light on a particular mechanism of transcriptional error correction involving: (i) diffusive translocation of the RNA polymerase along the DNA (backtracking) and (ii) irreversible RNA cleavage. This mechanism achieves preferential cleavage of misincorporated nucleotides by biasing the local rates of translocation. Here, we study how misincorporated nucleotides affect backtracking dynamics and how this effect determines the level of transcriptional fidelity. We consider backtracking as a diffusive process in a periodic, one-dimensional energy landscape, which at a coarse-grained level gives rise to a hopping process between neighbouring local minima. We propose a model for how misincorporated nucleotides deform this energy landscape and hence affect the hopping rates. In particular, we show that this model can be used to derive both the theoretical limit on the fidelity (i.e. the minimum fraction of misincorporated nucleotides) and the actual fidelity relative to this optimum, achieved for specific combinations of the cleavage and polymerization rates. Finally, we study how external factors influencing backtracking dynamics affect transcriptional fidelity. We show that biologically relevant loads, similar to those exerted by nucleosomes or other transcriptional barriers, increase error correction. PMID:22551978

  12. Photoinitiator Nucleotide for Quantifying Nucleic Acid Hybridization

    PubMed Central

    Johnson, Leah M.; Hansen, Ryan R.; Urban, Milan; Kuchta, Robert D.; Bowman, Christopher N.

    2010-01-01

    This first report of a photoinitiator-nucleotide conjugate demonstrates a novel approach for sensitive, rapid and visual detection of DNA hybridization events. This approach holds potential for various DNA labeling schemes and for applications benefiting from selective DNA-based polymerization initiators. Here, we demonstrate covalent, enzymatic incorporation of an eosin-photoinitiator 2′-deoxyuridine-5′-triphosphate (EITC-dUTP) conjugate into surface-immobilized DNA hybrids. Subsequent radical chain photoinitiation from these sites using an acrylamide/bis-acrylamide formulation yields a dynamic detection range between 500pM and 50nM of DNA target. Increasing EITC-nucleotide surface densities leads to an increase in surface-based polymer film heights until achieving a film height plateau of 280nm ±20nm at 610 ±70 EITC-nucleotides/μm2. Film heights of 10–20 nm were obtained from eosin surface densities of approximately 20 EITC-nucleotides/μm2 while below the detection limit of ~10 EITC-nucleotides/μm2, no detectable films were formed. This unique threshold behavior is utilized for instrument-free, visual quantification of target DNA concentration ranges. PMID:20337438

  13. Secondary structure of mouse 28S rRNA and general model for the folding of the large rRNA in eukaryotes.

    PubMed Central

    Michot, B; Hassouna, N; Bachellerie, J P

    1984-01-01

    We present a secondary structure model for the entire sequence of mouse 28S rRNA (1) which is based on an extensive comparative analysis of the available eukaryotic sequences, i.e. yeast (2, 3), Physarum polycephalum (4), Xenopus laevis (5) and rat (6). It has been derived with close reference to the models previously proposed for yeast 26S rRNA (2) and for prokaryotic 23S rRNA (7-9). Examination of the recently published eukaryotic sequences confirms that all pro- and eukaryotic large rRNAs share a largely conserved secondary structure core, as already apparent from the previous analysis of yeast 26S rRNA (2). These new comparative data confirm most features of the yeast model (2). They also provide the basis for a few modifications and for new proposals which extend the boundaries of the common structural core (now representing about 85% of E. coli 23S rRNA length) and bring new insights for tracing the structural evolution, in higher eukaryotes, of the domains which have no prokaryotic equivalent and are inserted at specific locations within the common structural core of the large subunit rRNA. PMID:6374617

  14. A Nucleotide Phosphatase Activity in the Nucleotide Binding Domain of an Orphan Resistance Protein from Rice*

    PubMed Central

    Fenyk, Stepan; de San Eustaquio Campillo, Alba; Pohl, Ehmke; Hussey, Patrick J.; Cann, Martin J.

    2012-01-01

    Plant resistance proteins (R-proteins) are key components of the plant immune system activated in response to a plethora of different pathogens. R-proteins are P-loop NTPase superfamily members, and current models describe their main function as ATPases in defense signaling pathways. Here we show that a subset of R-proteins have evolved a new function to combat pathogen infection. This subset of R-proteins possesses a nucleotide phosphatase activity in the nucleotide-binding domain. Related R-proteins that fall in the same phylogenetic clade all show the same nucleotide phosphatase activity indicating a conserved function within at least a subset of R-proteins. R-protein nucleotide phosphatases catalyze the production of nucleoside from nucleotide with the nucleotide monophosphate as the preferred substrate. Mutation of conserved catalytic residues substantially reduced activity consistent with the biochemistry of P-loop NTPases. Kinetic analysis, analytical gel filtration, and chemical cross-linking demonstrated that the nucleotide-binding domain was active as a multimer. Nuclear magnetic resonance and nucleotide analogues identified the terminal phosphate bond as the target of a reaction that utilized a metal-mediated nucleophilic attack by water on the phosphoester. In conclusion, we have identified a group of R-proteins with a unique function. This biochemical activity appears to have co-evolved with plants in signaling pathways designed to resist pathogen attack. PMID:22157756

  15. Eukaryotic rRNA Modification by Yeast 5-Methylcytosine-Methyltransferases and Human Proliferation-Associated Antigen p120

    PubMed Central

    Gaspar, Imre; Aigueperse, Christelle; Schaefer, Matthias; Kellner, Stefanie; Helm, Mark; Motorin, Yuri

    2015-01-01

    Modified nucleotide 5-methylcytosine (m5C) is frequently present in various eukaryotic RNAs, including tRNAs, rRNAs and in other non-coding RNAs, as well as in mRNAs. RNA:m5C-methyltranferases (MTases) Nop2 from S. cerevisiae and human proliferation-associated nucleolar antigen p120 are both members of a protein family called Nop2/NSUN/NOL1. Protein p120 is well-known as a tumor marker which is over-expressed in various cancer tissues. Using a combination of RNA bisulfite sequencing and HPLC-MS/MS analysis, we demonstrated here that p120 displays an RNA:m5C- MTase activity, which restores m5C formation at position 2870 in domain V of 25S rRNA in a nop2Δ yeast strain. We also confirm that yeast proteins Nop2p and Rcm1p catalyze the formation of m5C in domains V and IV, respectively. In addition, we do not find any evidence of m5C residues in yeast 18S rRNA. We also performed functional complementation of Nop2-deficient yeasts by human p120 and studied the importance of different sequence and structural domains of Nop2 and p120 for yeast growth and m5C-MTase activity. Chimeric protein formed by Nop2 and p120 fragments revealed the importance of Nop2 N-terminal domain for correct protein localization and its cellular function. We also validated that the presence of Nop2, rather than the m5C modification in rRNA itself, is required for pre-rRNA processing. Our results corroborate that Nop2 belongs to the large family of pre-ribosomal proteins and possesses two related functions in pre-rRNA processing: as an essential factor for cleavages and m5C:RNA:modification. These results support the notion of quality control during ribosome synthesis by such modification enzymes. PMID:26196125

  16. Two-Dimensional Combinatorial Screening (2DCS) of a Bacterial rRNA A-site-like Motif Library: Defining Privileged Asymmetric Internal Loops that Bind Aminoglycosides

    PubMed Central

    Tran, Tuan; Disney, Matthew D.

    2010-01-01

    RNAs have diverse structures that are important for biological function. These structures include bulges and internal loops that can form tertiary contacts or serve as ligand binding sites. The most commonly exploited RNA drug target for small molecule intervention is the bacterial ribosome, more specifically the ribosomal RNA aminoacyl-tRNA site (rRNA A-site) which is a major target for the aminoglycoside class of antibiotics. The bacterial A-site is composed of a 1×1 nucleotide all-U internal loop and a 2×1 nucleotide all-A internal loop separated by a single GC base pair. Therefore, we probed the molecular recognition of a small library of four aminoglycosides for binding a 16384-member bacterial rRNA A-site-like internal loop library using Two-Dimensional Combinatorial Screening (2DCS). 2DCS is a microarray-based method that probes RNA and chemical spaces simultaneously. These studies sought to determine if aminoglycosides select their therapeutic target if given a choice of binding all possible internal loops derived from an A-site-like library. Results show that the bacterial rRNA A-site was not selected by any aminoglycoside. Analyses of selected sequences using the RNA Privileged Space Predictor (RNA-PSP) program show that each aminoglycoside preferentially binds different types of internal loops. For three of the aminoglycosides, 6″-azido-kanamycin A, 5-O-(2-azidoethyl) neamine, and 6″-azido-tobramycin, the selected internal loops bind with ~10-fold higher affinity than the bacterial rRNA A-site. The internal loops selected to bind 5″-azido-neomycin B bind with similar affinity as the therapeutic target. Selected internal loops that are unique for each aminoglycoside have dissociation constants ranging from 25 to 270 nM and are specific for the aminoglycoside they were selected to bind compared to the other arrayed aminoglycosides. These studies further establish a database of RNA motifs that are recognized by small molecules that could be used to

  17. Delayed Acquisition of Non-Adjacent Vocalic Distributional Regularities

    ERIC Educational Resources Information Center

    Gonzalez-Gomez, Nayeli; Nazzi, Thierry

    2016-01-01

    The ability to compute non-adjacent regularities is key in the acquisition of a new language. In the domain of phonology/phonotactics, sensitivity to non-adjacent regularities between consonants has been found to appear between 7 and 10 months. The present study focuses on the emergence of a posterior-anterior (PA) bias, a regularity involving two…

  18. View of north side from exterior stairs of adjacent building, ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    View of north side from exterior stairs of adjacent building, bottom cut off by fringed buildings, view facing south-southwest - U.S. Naval Base, Pearl Harbor, Industrial X-Ray Building, Off Sixth Street, adjacent to and south of Facility No. 11, Pearl City, Honolulu County, HI

  19. Learning Non-Adjacent Regularities at Age 0 ; 7

    ERIC Educational Resources Information Center

    Gervain, Judit; Werker, Janet F.

    2013-01-01

    One important mechanism suggested to underlie the acquisition of grammar is rule learning. Indeed, infants aged 0 ; 7 are able to learn rules based on simple identity relations (adjacent repetitions, ABB: "wo fe fe" and non-adjacent repetitions, ABA: "wo fe wo", respectively; Marcus et al., 1999). One unexplored issue is…

  20. The International Nucleotide Sequence Database Collaboration.

    PubMed

    Cochrane, Guy; Karsch-Mizrachi, Ilene; Takagi, Toshihisa

    2016-01-01

    The International Nucleotide Sequence Database Collaboration (INSDC; http://www.insdc.org) comprises three global partners committed to capturing, preserving and providing comprehensive public-domain nucleotide sequence information. The INSDC establishes standards, formats and protocols for data and metadata to make it easier for individuals and organisations to submit their nucleotide data reliably to public archives. This work enables the continuous, global exchange of information about living things. Here we present an update of the INSDC in 2015, including data growth and diversification, new standards and requirements by publishers for authors to submit their data to the public archives. The INSDC serves as a model for data sharing in the life sciences. PMID:26657633

  1. Nucleotide `maps' of digests of deoxyribonucleic acid

    PubMed Central

    Murray, K.

    1970-01-01

    Various digests of 32P-labelled DNA were examined by two-dimensional ionophoresis on cellulose acetate and DEAE-cellulose paper. The products from digestion with pancreatic deoxyribonuclease and Neurospora crassa endonuclease were qualitatively closely similar, but very complex, and were used to investigate the mapping behaviour of nucleotides in various ionophoretic systems. Ionophoresis on DEAE-cellulose paper in triethylamine carbonate, pH 9.7, followed by ionophoresis in the second dimension at pH1.9 gave high resolution of nucleotides in very complex mixtures and permitted the fractionation of larger quantities than is possible on cellulose acetate. High resolution of nucleotides in compact spots was obtained with two-dimensional ionophoresis on cellulose acetate and AE-cellulose paper, a system that is a useful supplement to those based on DEAE-cellulose paper. ImagesPLATE 7PLATE 1PLATE 2PLATE 3PLATE 4PLATE 5PLATE 6 PMID:5476726

  2. Ribosome heterogeneity in tumorigenesis: the rRNA point of view

    PubMed Central

    Marcel, Virginie; Catez, Frédéric; Diaz, Jean-Jacques

    2015-01-01

    The "specialized ribosome" concept proposes that ribosome variants are produced and differentially regulate translation. Examples supporting this notion demonstrated heterogeneity of ribosomal protein composition. However, ribosome translational activity is carried out by rRNA. We, and others, recently showed that rRNA heterogeneity regulates translation to generate distinct translatomes promoting tumorigenesis. PMID:27305893

  3. Tetrathiobacter kashmirensis Strain CA-1 16S rRNA gene complete sequence.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    This study used 1326 base pair 16S rRNA gene sequence methods to confirm the identification of a bacterium as Tetrathiobacter kashmirensis. Morphological, biochemical characteristics, and fatty acid profiles are consistent with the 16S rRNA gene sequence identification of the bacterium. The isolate...

  4. Exploiting the CRISPR/Cas9 PAM Constraint for Single-Nucleotide Resolution Interventions

    PubMed Central

    Li, Yi; Mendiratta, Saurabh; Ehrhardt, Kristina; Kashyap, Neha; White, Michael A.; Bleris, Leonidas

    2016-01-01

    CRISPR/Cas9 is an enabling RNA-guided technology for genome targeting and engineering. An acute DNA binding constraint of the Cas9 protein is the Protospacer Adjacent Motif (PAM). Here we demonstrate that the PAM requirement can be exploited to specifically target single-nucleotide heterozygous mutations while exerting no aberrant effects on the wild-type alleles. Specifically, we target the heterozygous G13A activating mutation of KRAS in colorectal cancer cells and we show reversal of drug resistance to a MEK small-molecule inhibitor. Our study introduces a new paradigm in genome editing and therapeutic targeting via the use of gRNA to guide Cas9 to a desired protospacer adjacent motif. PMID:26788852

  5. Characteristic archaebacterial 16S rRNA oligonucleotides

    NASA Technical Reports Server (NTRS)

    McGill, T. J.; Jurka, J.; Sobieski, J. M.; Pickett, M. H.; Woese, C. R.; Fox, G. E.

    1986-01-01

    A method of analyzing 16S rRNA catalog data has been developed in which groupings at various taxonomic levels can be characterized in terms of specific "signature" oligonucleotides. This approach provides an alternative means for evaluating higher order branching possibilities and can be used to assess the phylogenetic position of isolates that are poorly placed by the usual clustering procedures. This signature approach has been applied to forty archaebacterial catalogs and every oligonucleotide with significant signature value has been identified. Sets of specific oligonucleotides were identified for every major group on a dendrogram produced by cluster analysis procedures. Signatures that would establish between group relationships were also sought and found. In the case of the Methanobacteriaceae the clustering methods suggest a specific relationship to the Methanococcaceae. This inclusion is in fact supported by six strong signature oligonucleotides. However there are also significant numbers of signature oligonucleotides supporting a specific relationship of the Methanobacteriaceae to either the Halobacteriaceae or the Methanomicrobiaceae. Thus the placement of the Methanobacteriaceae is less certain than the usual dendrograms imply. The signature approach also was used to assess the phylogenetic position of Thermoplasma acidophilum which is found to be more closely related to the methanogen/halophile Division than to the sulfur dependent Division of the archaebacteria. This does not imply however that Thermoplasma acidophilum is properly regarded as being in the methanogen/halophile Division.

  6. Regulation of Ion Channels by Pyridine Nucleotides

    PubMed Central

    Kilfoil, Peter J.; Tipparaju, Srinivas M.; Barski, Oleg A.; Bhatnagar, Aruni

    2014-01-01

    Recent research suggests that in addition to their role as soluble electron carriers, pyridine nucleotides [NAD(P)(H)] also regulate ion transport mechanisms. This mode of regulation seems to have been conserved through evolution. Several bacterial ion–transporting proteins or their auxiliary subunits possess nucleotide-binding domains. In eukaryotes, the Kv1 and Kv4 channels interact with pyridine nucleotide–binding β-subunits that belong to the aldo-keto reductase superfamily. Binding of NADP+ to Kvβ removes N-type inactivation of Kv currents, whereas NADPH stabilizes channel inactivation. Pyridine nucleotides also regulate Slo channels by interacting with their cytosolic regulator of potassium conductance domains that show high sequence homology to the bacterial TrkA family of K+ transporters. These nucleotides also have been shown to modify the activity of the plasma membrane KATP channels, the cystic fibrosis transmembrane conductance regulator, the transient receptor potential M2 channel, and the intracellular ryanodine receptor calcium release channels. In addition, pyridine nucleotides also modulate the voltage-gated sodium channel by supporting the activity of its ancillary subunit—the glycerol-3-phosphate dehydrogenase-like protein. Moreover, the NADP+ metabolite, NAADP+, regulates intracellular calcium homeostasis via the 2-pore channel, ryanodine receptor, or transient receptor potential M2 channels. Regulation of ion channels by pyridine nucleotides may be required for integrating cell ion transport to energetics and for sensing oxygen levels or metabolite availability. This mechanism also may be an important component of hypoxic pulmonary vasoconstriction, memory, and circadian rhythms, and disruption of this regulatory axis may be linked to dysregulation of calcium homeostasis and cardiac arrhythmias. PMID:23410881

  7. Nucleotide chloramines and neutrophil-mediated cytotoxicity.

    PubMed

    Bernofsky, C

    1991-03-01

    Hypochlorite is a reactive oxidant formed as an end product of the respiratory burst in activated neutrophils. It is responsible for killing bacteria and has been implicated in neutrophil-mediated tissue injury associated with the inflammatory process. Although hypochlorite is a potent cytotoxic agent, the primary mechanism by which it exerts its effect is unclear. This review examines evidence that the primary event in hypochlorite cytotoxicity is the loss of adenine nucleotides from the target cell. This loss appears to be mediated by the formation of adenine nucleotide chloramines which are reactive intermediates with a free radical character and are capable of forming stable ligands with proteins and nucleic acids. PMID:1848195

  8. Evidence for the presence of 5S rRNA in mammalian mitochondria.

    PubMed

    Magalhães, P J; Andreu, A L; Schon, E A

    1998-09-01

    Mammalian mitochondrial ribosomes contain two prokaryotic-like rRNAs, 12S and 16S, both encoded by mitochondrial DNA. As opposed to cytosolic ribosomes, however, these ribosomes are not thought to contain 5S rRNA. For this reason, it has been unclear whether 5S rRNA, which can be detected in mitochondrial preparations, is an authentic organellar species imported from the cytosol or is merely a copurifying cytosol-derived contaminant. We now show that 5S rRNA is tightly associated with highly purified mitochondrial fractions of human and rat cells and that 5S rRNA transcripts derived from a synthetic gene transfected transiently into human cells are both expressed in vivo and present in highly purified mitochondria and mitoplasts. We conclude that 5S rRNA is imported into mammalian mitochondria, but its function there still remains to be clarified. PMID:9725900

  9. Structural and functional analysis of 5S rRNA in Saccharomyces cerevisiae

    PubMed Central

    Kiparisov, S.; Sergiev, P. V.; Dontsova, O. A.; Petrov, A.; Meskauskas, A.; Dinman, J. D.

    2005-01-01

    5S rRNA extends from the central protuberance of the large ribosomal subunit, through the A-site finger, and down to the GTPase-associated center. Here, we present a structure-function analysis of seven 5S rRNA alleles which are sufficient for viability in the yeast Saccharomyces cerevisiae when expressed in the absence of wild-type 5S rRNAs, and extend this analysis using a large bank of mutant alleles that show semidominant phenotypes in the presence of wild-type 5S rRNA. This analysis supports the hypothesis that 5S rRNA serves to link together several different functional centers of the ribosome. Data are also presented which suggest that in eukaryotic genomes selection has favored the maintenance of multiple alleles of 5S rRNA, and that these may provide cells with a mechanism to post-transcriptionally regulate gene expression. PMID:16047201

  10. Subnuclear partitioning of rRNA genes between the nucleolus and nucleoplasm reflects alternative epiallelic states

    PubMed Central

    Pontvianne, Frederic; Blevins, Todd; Chandrasekhara, Chinmayi; Mozgová, Iva; Hassel, Christiane; Pontes, Olga M.F.; Tucker, Sarah; Mokroš, Petr; Muchová, Veronika; Fajkus, Jiří; Pikaard, Craig S.

    2013-01-01

    Eukaryotes can have thousands of 45S ribosomal RNA (rRNA) genes, many of which are silenced during development. Using fluorescence-activated sorting techniques, we show that active rRNA genes in Arabidopsis thaliana are present within sorted nucleoli, whereas silenced rRNA genes are excluded. DNA methyltransferase (met1), histone deacetylase (hda6), or chromatin assembly (caf1) mutants that disrupt silencing abrogate this nucleoplasmic–nucleolar partitioning. Bisulfite sequencing data indicate that active nucleolar rRNA genes are nearly completely demethylated at promoter CGs, whereas silenced genes are nearly fully methylated. Collectively, the data reveal that rRNA genes occupy distinct but changeable nuclear territories according to their epigenetic state. PMID:23873938

  11. The influence of different land uses on the structure of archaeal communities in Amazonian anthrosols based on 16S rRNA and amoA genes.

    PubMed

    Taketani, Rodrigo Gouvêa; Tsai, Siu Mui

    2010-05-01

    Soil from the Amazonian region is usually regarded as unsuitable for agriculture because of its low organic matter content and low pH; however, this region also contains extremely rich soil, the Terra Preta Anthrosol. A diverse archaeal community usually inhabits acidic soils, such as those found in the Amazon. Therefore, we hypothesized that this community should be sensitive to changes in the environment. Here, the archaeal community composition of Terra Preta and adjacent soil was examined in four different sites in the Brazilian Amazon under different anthropic activities. The canonical correspondence analysis of terminal restriction fragment length polymorphisms has shown that the archaeal community structure was mostly influenced by soil attributes that differentiate the Terra Preta from the adjacent soil (i.e., pH, sulfur, and organic matter). Archaeal 16S rRNA gene clone libraries indicated that the two most abundant genera in both soils were Candidatus nitrosphaera and Canditatus nitrosocaldus. An ammonia monoxygenase gene (amoA) clone library analysis indicated that, within each site, there was no significant difference between the clone libraries of Terra Preta and adjacent soils. However, these clone libraries indicated there were significant differences between sites. Quantitative PCR has shown that Terra Preta soils subjected to agriculture displayed a higher number of amoA gene copy numbers than in adjacent soils. On the other hand, soils that were not subjected to agriculture did not display significant differences on amoA gene copy numbers between Terra Preta and adjacent soils. Taken together, our findings indicate that the overall archaeal community structure in these Amazonian soils is determined by the soil type and the current land use. PMID:20204349

  12. Nucleotide sequence of bacteriophage fd DNA.

    PubMed Central

    Beck, E; Sommer, R; Auerswald, E A; Kurz, C; Zink, B; Osterburg, G; Schaller, H; Sugimoto, K; Sugisaki, H; Okamoto, T; Takanami, M

    1978-01-01

    The sequence of the 6,408 nucleotides of bacteriophage fd DNA has been determined. This allows to deduce the exact organisation of the filamentous phage genome and provides easy access to DNA segments of known structure and function. PMID:745987

  13. Nucleotide-metabolizing enzymes in Chlamydomonas flagella.

    PubMed

    Watanabe, T; Flavin, M

    1976-01-10

    Nucleotides have at least two functions in eukaryotic cilia and flagella. ATP, originating in the cells, is utilized for motility by energy-transducing protein(s) called dynein, and the binding of guanine nucleotides to tubulin, and probably certain transformations of the bound nucleotides, are prerequisites for the assembly of microtubules. Besides dynein, which can be solubulized from Chlamydomonas flagella as a heterogeneous, Mg2+ or Ca2+-activated ATPase, we have purified and characterized five other flagellar enzymes involved in nucleotide transformations. A homogeneous, low molecular weight, Ca2+-specific adenosine triphosphatase was isolated, which was inhibited by Mg2+ and was not specific for ATP. This enzyme was not formed by treating purified dynein with proteases. It was absent from extracts of Tetrahymena cilia. Its function might be an auxiliary energy transducer, or in steering or tactic responses. Two species of adenylate kinase were isolated, one of which was much elevated in regenerating flagella; the latter was also present in cell bodies. A large part of flagellar nucleoside diphosphokinase activity could not be solubilized. Two soluble enzyme species were identified, one of which was also present in cell bodies. Since these enzymes are of interest because they might function in microtubule assembly, we studied the extent to which brain nucleoside diphosphokinase co-polymerizes with tubulin purified by repeated cycles of polymerization. Arginine kinase was not detected in Chlamydomonas flagellar extracts. PMID:397

  14. Thermoelastic response of thin metal films and their adjacent materials

    SciTech Connect

    Kang, S.; Yoon, Y.; Kim, J.; Kim, W.

    2013-01-14

    A pulsed laser beam applied to a thin metal film is capable of launching an acoustic wave due to thermal expansion. Heat transfer from the thin metal film to adjacent materials can also induce thermal expansion; thus, the properties of these adjacent materials (as well as the thin metal film) should be considered for a complete description of the thermoelastic response. Here, we show that adjacent materials with a small specific heat and large thermal expansion coefficient can generate an enhanced acoustic wave and we demonstrate a three-fold increase in the peak pressure of the generated acoustic wave on substitution of parylene for polydimethylsiloxane.

  15. Cyclic nucleotide phosphodiesterase 1 and vascular aging.

    PubMed

    Yan, Chen

    2015-12-01

    VSMCs (vascular smooth muscle cells) play critical roles in arterial remodelling with aging, hypertension and atherosclerosis. VSMCs exist in diverse phenotypes and exhibit phenotypic plasticity, e.g. changing from a quiescent/contractile phenotype to an active myofibroblast-like, often called 'synthetic', phenotype. Synthetic VSMCs are able to proliferate, migrate and secrete ECM (extracellular matrix) proteinases and ECM proteins. In addition, they produce pro-inflammatory molecules, providing an inflammatory microenvironment for leucocyte penetration, accumulation and activation. The aging VSMCs have also shown changes in cellular phenotype, responsiveness to contracting and relaxing mediators, replicating potential, matrix synthesis, inflammatory mediators and intracellular signalling. VSMC dysfunction plays a key role in age-associated vascular remodelling. Cyclic nucleotide PDEs (phosphodiesterases), by catalysing cyclic nucleotide hydrolysis, play a critical role in regulating the amplitude, duration and compartmentalization of cyclic nucleotide signalling. Abnormal alterations of PDEs and subsequent changes in cyclic nucleotide homoeostasis have been implicated in a number of different diseases. In the study published in the latest issue of Clinical Science, Bautista Niño and colleagues have shown that, in cultured senescent human VSMCs, PDE1A and PDE1C mRNA levels are significantly up-regulated and inhibition of PDE1 activity with vinpocetine reduced cellular senescent makers in senescent VSMCs. Moreover, in the premature aging mice with genomic instability (Ercc1(d/-)), impaired aortic ring relaxation in response to SNP (sodium nitroprusside), an NO (nitric oxide) donor, was also largely improved by vinpocetine. More interestingly, using data from human GWAS (genome-wide association studies), it has been found that PDE1A single nucleotide polymorphisms is significantly associated with diastolic blood pressure and carotid intima-media thickening, two

  16. Transferable Resistance to Aminoglycosides by Methylation of G1405 in 16S rRNA and to Hydrophilic Fluoroquinolones by QepA-Mediated Efflux in Escherichia coli▿

    PubMed Central

    Périchon, Bruno; Courvalin, Patrice; Galimand, Marc

    2007-01-01

    Plasmid pIP1206 was detected in Escherichia coli strain 1540 during the screening of clinical isolates of Enterobacteriaceae for high-level resistance to aminoglycosides. The sequence of this IncFI conjugative plasmid of ca. 100 kb was partially determined. pIP1206 carried the rmtB gene for a ribosome methyltransferase that was shown to modify the N7 position of nucleotide G1405, located in the A site of 16S rRNA. It also contained the qepA (quinolone efflux pump) gene that encodes a 14-transmembrane-segment putative efflux pump belonging to the major facilitator superfamily of proton-dependent transporters. Disruption of membrane proton potential by the efflux pump inhibitor carbonyl cyanide m-chlorophenylhydrazone in a transconjugant harboring the qepA gene resulted in elevation of norfloxacin accumulation. The transporter conferred resistance to the hydrophilic quinolones norfloxacin and ciprofloxacin. PMID:17470656

  17. Direct detection of 16S rRNA in soil extracts by using oligonucleotide microarrays.

    PubMed

    Small, J; Call, D R; Brockman, F J; Straub, T M; Chandler, D P

    2001-10-01

    We report on the development and validation of a simple microarray method for the direct detection of intact 16S rRNA from unpurified soil extracts. Total RNAs from Geobacter chapellei and Desulfovibrio desulfuricans were hybridized to an oligonucleotide array consisting of universal and species-specific 16S rRNA probes. PCR-amplified products from Geobacter and Desulfovibrio were easily and specifically detected under a range of hybridization times, temperatures, and buffers. However, reproducible, specific hybridization and detection of intact rRNA could be accomplished only by using a chaperone-detector probe strategy. With this knowledge, assay conditions were developed for rRNA detection using a 2-h hybridization time at room temperature. Hybridization specificity and signal intensity were enhanced using fragmented RNA. Formamide was required in the hybridization buffer in order to achieve species-specific detection of intact rRNA. With the chaperone detection strategy, we were able to specifically hybridize and detect G. chapellei 16S rRNA directly from a total-RNA soil extract, without further purification or removal of soluble soil constituents. The detection sensitivity for G. chapellei 16S rRNA in soil extracts was at least 0.5 microg of total RNA, representing approximately 7.5 x 10(6) Geobacter cell equivalents of RNA. These results suggest that it is now possible to apply microarray technology to the direct detection of microorganisms in environmental samples, without using PCR. PMID:11571176

  18. Trans-splicing and RNA editing of LSU rRNA in Diplonema mitochondria

    PubMed Central

    Valach, Matus; Moreira, Sandrine; Kiethega, Georgette N.; Burger, Gertraud

    2014-01-01

    Mitochondrial ribosomal RNAs (rRNAs) often display reduced size and deviant secondary structure, and sometimes are fragmented, as are their corresponding genes. Here we report a mitochondrial large subunit rRNA (mt-LSU rRNA) with unprecedented features. In the protist Diplonema, the rnl gene is split into two pieces (modules 1 and 2, 534- and 352-nt long) that are encoded by distinct mitochondrial chromosomes, yet the rRNA is continuous. To reconstruct the post-transcriptional maturation pathway of this rRNA, we have catalogued transcript intermediates by deep RNA sequencing and RT-PCR. Gene modules are transcribed separately. Subsequently, transcripts are end-processed, the module-1 transcript is polyuridylated and the module-2 transcript is polyadenylated. The two modules are joined via trans-splicing that retains at the junction ∼26 uridines, resulting in an extent of insertion RNA editing not observed before in any system. The A-tail of trans-spliced molecules is shorter than that of mono-module 2, and completely absent from mitoribosome-associated mt-LSU rRNA. We also characterize putative antisense transcripts. Antisense-mono-modules corroborate bi-directional transcription of chromosomes. Antisense-mt-LSU rRNA, if functional, has the potential of guiding concomitantly trans-splicing and editing of this rRNA. Together, these findings open a window on the investigation of complex regulatory networks that orchestrate multiple and biochemically diverse post-transcriptional events. PMID:24259427

  19. Depletion of pre-16S rRNA in starved Escherichia coli cells.

    PubMed

    Cangelosi, G A; Brabant, W H

    1997-07-01

    Specific hybridization assays for intermediates in rRNA synthesis (pre-rRNA) may become useful for monitoring the growth activity of individual microbial species in complex natural systems. This possibility depends upon the assumption that rRNA processing in microbial cells continues after growth and pre-rRNA synthesis cease, resulting in drainage of the pre-rRNA pool. This is not the case in many eukaryotic cells, but less is known about the situation in bacteria. Therefore, we used DNA probes to measure steady-state cellular pre-16S rRNA pools during growth state transitions in Escherichia coli. Pre-16S rRNA became undetectable when cells entered the stationary phase on rich medium and was replenished upon restoration of favorable growth conditions. These fluctuations were of much greater magnitude than concurrent fluctuations in the mature 16S rRNA pool. The extent of pre-16S rRNA depletion depended upon the circumstances limiting growth. It was significantly more pronounced in carbon-energy-starved cells than in nitrogen-starved cells or in cells treated with energy uncouplers. In the presence of the transcriptional inhibitor rifampin, rates of pre-16S rRNA depletion in carbon-energy-starved cells and nitrogen-starved cells were similar, suggesting that the difference between these conditions resides primarily at the level of pre-rRNA synthesis. Chloramphenicol, which inhibits the final steps in rRNA maturation, halted pre-16S rRNA depletion under all conditions. The data show that E. coli cells continue to process pre-rRNA after growth and rrn operon transcription cease, leading to drainage of the pre-rRNA pool. This supports the feasibility of using pre-rRNA-targeted probes to monitor bacterial growth in natural systems, with the caveat that patterns of pre-rRNA depletion vary with the conditions limiting growth. PMID:9226253

  20. Lock 4 View east of lock wall and adjacent ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Lock 4 - View east of lock wall and adjacent roadway built atop tow path. The gate pocket can be seen at center. - Savannah & Ogeechee Barge Canal, Between Ogeechee & Savannah Rivers, Savannah, Chatham County, GA

  1. 1. Ninth Street (west) facade. Adjacent on the north is ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    1. Ninth Street (west) facade. Adjacent on the north is the 9th Street facade of 816 E Street. Both buildings were originally one property. - Riley Building, Rendezvous Adult Magazines & Films, 437 Ninth Street, Northwest, Washington, District of Columbia, DC

  2. 2. THREEQUARTER VIEW FROM ADJACENT ACCESS ROAD SHOWING THREE SPANS ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    2. THREE-QUARTER VIEW FROM ADJACENT ACCESS ROAD SHOWING THREE SPANS AND NORTHWEST APPROACH SPANS, LOOKING SOUTHEAST - Red River Bridge, Spanning Red River at U.S. Highway 82, Garland, Miller County, AR

  3. 1. VIEW FROM ROOFTOP OF BUILDING (MOTEL) ADJACENT TO TECHWOOD ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    1. VIEW FROM ROOFTOP OF BUILDING (MOTEL) ADJACENT TO TECHWOOD HOMES, LOOKING SOUTH. GARAGE TO EXTREME LEFT, BUILDING 1 TO EXTREME RIGHT. - Techwood Homes (Public Housing), Bounded by North Avenue, Parker Street, William Street & Lovejoy Street, Atlanta, Fulton County, GA

  4. 3. View of north side of house facing from adjacent ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    3. View of north side of house facing from adjacent vacant property. Original wood lap siding and trim is covered by aluminum siding. Recessed side porch is in middle. - 645 South Eighteenth Street (House), Louisville, Jefferson County, KY

  5. 1. A BRICK AND CONCRETE FAN HOUSING ADJACENT TO ONE ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    1. A BRICK AND CONCRETE FAN HOUSING ADJACENT TO ONE OF THE ADIT OPENINGS (VIEW TO THE NORTH). - Foster Gulch Mine, Fan Housing, Bear Creek 1 mile Southwest of Town of Bear Creek, Red Lodge, Carbon County, MT

  6. 7. August, 1970 9 ORANGE STREET, ADJACENT TO UNITARIAN CHURCH ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    7. August, 1970 9 ORANGE STREET, ADJACENT TO UNITARIAN CHURCH (NOT IN STUDY AREA) - Orange & Union Streets Neighborhood Study, 8-31 Orange Street, 9-21 Union Street & Stone Alley, Nantucket, Nantucket County, MA

  7. OBLIQUE OF SOUTHWEST END AND SOUTHEAST SIDE, WITH ADJACENT FACILITY ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    OBLIQUE OF SOUTHWEST END AND SOUTHEAST SIDE, WITH ADJACENT FACILITY 391 IN THE FOREGROUND. - U.S. Naval Base, Pearl Harbor, Joint Intelligence Center, Makalapa Drive in Makalapa Administration Area, Pearl City, Honolulu County, HI

  8. Complications in exodontia--accidental dislodgment to adjacent anatomical areas.

    PubMed

    Grandini, S A; Barros, V M; Salata, L A; Rosa, A L; Soares, U N

    1993-01-01

    The authors report 4 cases of accidental dislodgement of teeth to adjacent anatomical areas during extraction. The causes and their prevention are discussed and solutions for the problem are suggested. PMID:8241759

  9. 6. Detail, vertical guides adjacent to east portal of Tunnel ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    6. Detail, vertical guides adjacent to east portal of Tunnel 28, view to southwest, 135mm lens with electronic flash fill. - Central Pacific Transcontinental Railroad, Tunnel No. 28, Milepost 134.75, Applegate, Placer County, CA

  10. Interior building details of Building A, dungeon cell adjacent to ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Interior building details of Building A, dungeon cell adjacent to northwest cell: granite and brick threshold, poured concrete floors, plastered finished walls, vaulted veiling; northwesterly view - San Quentin State Prison, Building 22, Point San Quentin, San Quentin, Marin County, CA

  11. 73. PASSAGE ADJACENT TO ROOM 232, EAST WING, SECOND FLOOR, ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    73. PASSAGE ADJACENT TO ROOM 232, EAST WING, SECOND FLOOR, LOOKING WEST BY NORTHWEST, SHOWING EASTERNMOST ARCH OF FORMER GREAT HALL NORTH ARCADE - Smithsonian Institution Building, 1000 Jefferson Drive, between Ninth & Twelfth Streets, Southwest, Washington, District of Columbia, DC

  12. View of viaduct, looking SE from roof of adjacent parking ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    View of viaduct, looking SE from roof of adjacent parking garage. - Mulberry Street Viaduct, Spanning Paxton Creek & Cameron Street (State Route 230) at Mulberry Street (State Route 3012), Harrisburg, Dauphin County, PA

  13. Cement Leakage into Adjacent Vertebral Body Following Percutaneous Vertebroplasty

    PubMed Central

    Park, Jae Hoo; Kim, Hyeun Sung

    2016-01-01

    Percutaneous vertebroplasty (PV) is a minimally invasive procedure for osteoporotic vertebral compression fractures that fail to respond to conventional conservative treatment. It significantly improves intolerable back pain within hours, and has a low complication rate. Although rare, PV is not free of complications, most of which are directly related to cement leakage. Because of its association with new adjacent fracture, the importance of cement leakage into the adjacent disc space is paramount. Here, we report an interesting case of cement leakage into the adjacent upper vertebral body as well as disc space following PV. To the best of our knowledge, there has been no report of cement leakage into the adjacent vertebral body following PV. This rare case is presented along with a review of the literature. PMID:27437018

  14. 1. HEBRONVILLE MILL COMPLEX ADJACENT TO NORTHEAST CORRIDOR. HEBRONVILLE, BRISTOL ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    1. HEBRONVILLE MILL COMPLEX ADJACENT TO NORTHEAST CORRIDOR. HEBRONVILLE, BRISTOL CO., MA. Sec. 4116, MP 193.75. - Northeast Railroad Corridor, Amtrak Route between RI/MA State Line & South Station, Boston, Suffolk County, MA

  15. 3. DODGEVILLE MILL COMPLEX ADJACENT TO NORTHEAST CORRIDOR DODGEVILLE, BRISTOL ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    3. DODGEVILLE MILL COMPLEX ADJACENT TO NORTHEAST CORRIDOR DODGEVILLE, BRISTOL CO., MA. Sec. 4116, MP 195.55. - Northeast Railroad Corridor, Amtrak Route between RI/MA State Line & South Station, Boston, Suffolk County, MA

  16. 33. HISTORIC PLAQUE MARKING WHERE JOHNSTON DIED, ADJACENT TO PATHWAY ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    33. HISTORIC PLAQUE MARKING WHERE JOHNSTON DIED, ADJACENT TO PATHWAY WITH CONCRETE CULVERT LEADING NORTH OUT OF RAVINE TOWARD JOHNSTON MEMORIAL SITE. VIEW NW. - Shiloh National Military Park Tour Roads, Shiloh, Hardin County, TN

  17. VIEW OF LAMP FIXTURE (EXTERIOR) ADJACENT TO ENTRANCE AT SOUTHWEST ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    VIEW OF LAMP FIXTURE (EXTERIOR) ADJACENT TO ENTRANCE AT SOUTHWEST CORNER OF BUILDING 23, FACING NORTH - Roosevelt Base, Auditorium-Gymnasium, West Virginia Street between Richardson & Reeves Avenues, Long Beach, Los Angeles County, CA

  18. 14. Charles Acey Cobb standing adjacent to the fish screen ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    14. Charles Acey Cobb standing adjacent to the fish screen he designed and installed in the Congdon Canal, facing southeast. Photo dates ca. late 1920's. - Congdon Canal, Fish Screen, Naches River, Yakima, Yakima County, WA

  19. 52. EASTSIDE PLANT: GENERAL VIEW OF GOVERNOR ADJACENT TO GENERATOR ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    52. EASTSIDE PLANT: GENERAL VIEW OF GOVERNOR ADJACENT TO GENERATOR - American Falls Water, Power & Light Company, Island Power Plant, Snake River, below American Falls Dam, American Falls, Power County, ID

  20. 24. INTERIOR VIEW, WILLIAM GRAY AT SIZING GUAGE ADJACENT TO ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    24. INTERIOR VIEW, WILLIAM GRAY AT SIZING GUAGE ADJACENT TO BRADLEY HAMMER; NOTE THIS IS THE SAME TOOL AS BEING FORGED ABOVE - Warwood Tool Company, Foot of Nineteenth Street, Wheeling, Ohio County, WV

  1. Detail exterior view looking north showing piping system adjacent to ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Detail exterior view looking north showing piping system adjacent to engine house. Gas cooling system is on far right. - Burnsville Natural Gas Pumping Station, Saratoga Avenue between Little Kanawha River & C&O Railroad line, Burnsville, Braxton County, WV

  2. VIEW OF CONSTRUCTION CAMP ROCK FEATURE WITH OVER, ADJACENT TO ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    VIEW OF CONSTRUCTION CAMP ROCK FEATURE WITH OVER, ADJACENT TO THE COLUMBIA SOUTHERN CANAL. LOOKING NORTHWEST - Tumalo Irrigation District, Tumalo Project, West of Deschutes River, Tumalo, Deschutes County, OR

  3. Pump house adjacent to the superintendent's house at the west ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Pump house adjacent to the superintendent's house at the west end of the complex near Highway 101. Detail of Holloshaft pump. View to the south. - Prairie Creek Fish Hatchery, Hwy. 101, Orick, Humboldt County, CA

  4. VIEW OF NORTHERN AND EASTERN SIDES FROM PARKING LOT ADJACENT ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    VIEW OF NORTHERN AND EASTERN SIDES FROM PARKING LOT ADJACENT TO BUILDING 199 (POLICE STATION) - U.S. Naval Base, Pearl Harbor, Post Office, Avenue A near Eleventh Avenue, Pearl City, Honolulu County, HI

  5. VIEW OF CONCRETE CHANNEL ADJACENT TO TUMALO FEED CANAL INTAKE ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    VIEW OF CONCRETE CHANNEL ADJACENT TO TUMALO FEED CANAL INTAKE STRUCTURE (DOWNSTREAM SIDE). LOOKING EAST/NORTHEAST - Tumalo Irrigation District, Tumalo Project, West of Deschutes River, Tumalo, Deschutes County, OR

  6. Adjacent Segment Disease Perspective and Review of the Literature

    PubMed Central

    Saavedra-Pozo, Fanor M.; Deusdara, Renato A. M.; Benzel, Edward C.

    2014-01-01

    Background Adjacent segment disease has become a common topic in spine surgery circles because of the significant increase in fusion surgery in recent years and the development of motion preservation technologies that theoretically should lead to a decrease in this pathology. The purpose of this review is to organize the evidence available in the current literature on this subject. Methods For this literature review, a search was conducted in PubMed with the following keywords: adjacent segment degeneration and disease. Selection, review, and analysis of the literature were completed according to level of evidence. Results The PubMed search identified 850 articles, from which 41 articles were selected and reviewed. The incidence of adjacent segment disease in the cervical spine is close to 3% without a significant statistical difference between surgical techniques (fusion vs arthroplasty). Authors report the incidence of adjacent segment disease in the lumbar spine to range from 2% to 14%. Damage to the posterior ligamentous complex and sagittal imbalances are important risk factors for both degeneration and disease. Conclusion Insufficient evidence exists at this point to support the idea that total disc arthroplasty is superior to fusion procedures in minimizing the incidence of adjacent segment disease. The etiology is most likely multifactorial but it is becoming abundantly clear that adjacent segment disease is not caused by motion segment fusion alone. Fusion plus the presence of abnormal end-fusion alignment appears to be a major factor in creating end-fusion stresses that result in adjacent segment degeneration and subsequent disease. The data presented cast further doubt on previously established rationales for total disc arthroplasty, at least with regard to the effect of total disc arthroplasty on adjacent segment degeneration pathology. PMID:24688337

  7. Approximating the largest eigenvalue of network adjacency matrices

    NASA Astrophysics Data System (ADS)

    Restrepo, Juan G.; Ott, Edward; Hunt, Brian R.

    2007-11-01

    The largest eigenvalue of the adjacency matrix of a network plays an important role in several network processes (e.g., synchronization of oscillators, percolation on directed networks, and linear stability of equilibria of network coupled systems). In this paper we develop approximations to the largest eigenvalue of adjacency matrices and discuss the relationships between these approximations. Numerical experiments on simulated networks are used to test our results.

  8. The phylogeny of Aerococcus and Pediococcus as determined by 16S rRNA sequence analysis: description of Tetragenococcus gen. nov.

    PubMed

    Collins, M D; Williams, A M; Wallbanks, S

    1990-08-01

    The phylogenetic interrelationships of the genera Pediococcus and Aerococcus were investigated using reverse transcriptase sequencing of 16S rRNA. The genus Pediococcus was found to be phylogenetically heterogeneous. The four species P. acidilactici, P. damnosus, P. parvulus and P. pentosaceus formed a phylogenetically distinct group. Within this pediococcal cluster, P. acidilactici was closely related to P. pentosaceus whereas P. damnosus showed a specific relationship with P. parvulus. The species P. dextrinicus, although showing significant sequence relatedness with these pediococcal species, was peripheral to the genus. Pediococcus halophilus exhibited low sequence homology with all of the species examined and formed a distinct line of descent. Pediococcus halophilus exhibited a closer affinity with enterococci and carnobacteria than with the other lactic acid bacteria. Pediococcus urinae-equi was phylogenetically very closely related to Aerococcus viridans. The 16S rRNA sequences of the type strains of these species differed by only two nucleotides (99.9% sequence homology) and clearly demonstrate that P. urinae-equi is a member of the genus Aerococcus. PMID:2227360

  9. Trichopodiella faurei n. sp. (Ciliophora, Phyllopharyngea, Cyrtophoria): morphological description and phylogenetic analyses based on SSU rRNA and group I intron sequences.

    PubMed

    Gong, Jun; Gao, Shan; Roberts, David McL; Al-Rasheid, Khaled A S; Song, Weibo

    2008-01-01

    A new marine cyrtophorian ciliate Trichopodiella faurei n. sp., which belongs to the order Dysteriida, family Hartmannulidae, was investigated at the morphological and molecular levels. A combination of morphological features of the organism including the oval body shape, 2-3 contractile vacuoles, 22-28 nematodesmal rods in the cytopharyngeal basket, and 31-39 somatic kineties, distinguishes it from all other known congeners. In reconstructed small subunit (SSU) rRNA phylogenies, T. faurei groups with Isochona, a representative genus of the subclass Chonotrichia. The similarity of the infraciliature between hartmannulids and several chonotrichian examples also suggests that these taxa should be closely related. A new S943 intron belonging to group IC1 was identified in the SSU rRNA gene of this species. This intron is phylogenetically related to the S891 introns previously found in the suctorians Acineta sp. and Tokophrya lemnarum, and their internal guide sequences share four nucleotides, indicating that these introns were vertically inherited from a common phyllopharyngean ancestor and that reverse splicing might have been involved in the transposition. PMID:19120794

  10. Differentiation of Listeria monocytogenes and Listeria innocua by 16S rRNA genes and intraspecies discrimination of Listeria monocytogenes strains by random amplified polymorphic DNA polymorphisms.

    PubMed Central

    Czajka, J; Bsat, N; Piani, M; Russ, W; Sultana, K; Wiedmann, M; Whitaker, R; Batt, C A

    1993-01-01

    Differences in the 16S rRNA genes (16S rDNA) which can be used to discriminate Listeria monocytogenes from Listeria innocua have been detected. The 16S rDNA were amplified by polymerase chain reaction with a set of oligonucleotide primers which flank a 1.5-kb fragment. Sequence differences were observed in the V2 region of the 16S rDNA both between L. monocytogenes Scott A and L. innocua and between different L. monocytogenes serotypes. Although L. monocytogenes SLCC2371 had the same V2 region sequence as L. innocua, the two species were different within the V9 region at nucleotides 1259 and 1292, in agreement with previous studies (R.-F. Wang, W.-W. Cao, and M.G. Johnson, Appl. Environ. Microbiol. 57:3666-3670, 1991). Intraspecies discrimination of L. monocytogenes strains was achieved by using the patterns generated by random amplified polymorphic DNA primers. Although some distinction can be made within the L. monocytogenes species by their 16S rDNA sequence, a far greater discrimination within species could be made by generating random amplified polymorphic DNA patterns from chromosomal DNA. By using a number of 10-bp primers, unique patterns for each isolate which in all cases examined differentiate between various L. monocytogenes serotypes, even though they may have the same 16S rRNA sequences, could be generated. Images PMID:8439157