Issues and Prospects of the Library in Japan : From the Answer of the Examination in Asistant Librarian Training

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papyrifera

Betula papyrifera Marshallpaper birch;white birch;canoe birchbouleau à papier;bouleau blanc;bouleau à canotBetula papyriferaMackenzie District, Reindeer Station, east branch of Mackenzie River, on middle slope of Caribou HillsTree with 6 trunks to 18 ft. in height, dbh 2 to 4 inche

La Petite presse : journal quotidien... / [rédacteur en chef : Balathier Bragelonne]

29 janvier 18851885/01/29 (A19,N6841)

Additional file 3: Table S2. of The PfAlba1 RNA-binding protein is an important regulator of translational timing in Plasmodium falciparum blood stages

Gene Ontology (GO) analysis of transcripts that co-immunoprecipitated with PfAlba1-Ty1 complexes in trophozoites. (XLS 192 kb

CsrA interacts directly with <i>fleQ</i>-mRNA <i>in vitro</i>.

<p><b>A</b>) Electromobility shift assays (EMSA) with 200nM of biotinylated <b><i>fleQ</i>-mRNA</b> combined with varying concentrations of purified CsrA-His were undertaken in a 6% Native Tris-PAGE gel. Lane 1: no CsrA, lane 2: 0.2 μM CsrA, lane 3: 0.5 μM CsrA, lane 4: 1.0 μM CsrA, lane 5: 2.0 μM CsrA, lane 6: 5.0 μM CsrA, lane 7: 5.0 μM CsrA + 2.0 μM unlabeled RsmZ. <b>B</b>) Mfold secondary structure prediction of the <b><i>fleQ</i>-mRNA</b> fragment used for the EMSA. Red, two potential CsrA-binding sites, which are mutated in C. Blue, transcriptional start codon. <b>C</b>) EMSA with recombinant CsrA and 200 nM of non-mutated RNA (pFleQ) or mutated in the indicated regions (mFleQ). AGGA motifs were replaced by an AAAA sequence using PCR mutagenesis. Lane 1: no CsrA + non-mutated <b><i>fleQ</i>-mRNA</b>, lane 2: 5.0 μM CsrA + non-mutated <b><i>fleQ</i>-mRNA</b>, lane 3: 5.0 μM CsrA + m<b><i>fleQ</i>-mRNA</b> mutated in region 1, lane 4: 5.0 μM CsrA + m<b><i>fleQ</i>-mRNA</b> mutated in region 2, lane 5: 5.0 μM CsrA + m<b><i>fleQ</i>-mRNA</b> mutated in both region 1 and 2.</p

CsrA modulates the expression of a thiamine pyrophosphate (TPP) riboswitch element.

<p><b>A</b>) Schematic representation of the <i>thi</i>-operon in <i>L</i>. <i>pneumophila</i> including the transcriptional start site (TSS), the CsrA-binding region, the thi element of the predicted TPP riboswitch and a predicted transcription termination site upstream of the start codon (AUG). The CsrA-binding site is overlapping the <i>thi</i> element of the TPP riboswitch. This organization suggests that CsrA is implicated in the fine-tuning of the expression of the downstream <i>thi</i>-operon most probably due to conformational changes in the secondary RNA structure. <b>B</b>) EMSA with 200nM of biotinylated thi-element (TPP) RNA and purified CsrA: Lane 1: no CsrA, lane 2: 1.0 μM CsrA, lane 3: 2.0 μM CsrA, lane 4: 5.0 μM CsrA, lane 5: 5.0 μM CsrA + 2.0 μM unlabeled RsmZ. <b>C)</b> Beta-lactamase (BlaM) assay in minimal medium grown <i>Legionella</i> without, with 1 mM and with 2 mM of TPP. BlaM activity in 10μg total protein of wt and <i>csrA</i>^<i>-</i> strain containing the 5'UTR of the <i>thi</i>-operon in a pXDC61 plasmid was measured. Each value represents the mean +/- SD of three independent experiments. BlaM activity is significantly decreased in the mutant at the different conditions indicating a positive effect of CsrA on the <i>thi</i>-operon expression in <i>L</i>. <i>pneumophila</i>. <b>D</b>) Model of the TPP riboswitch modulated by CsrA. Mfold prediction of the secondary structure of the 5'UTR <i>thi</i>-region: When TPP is bound, the OFF state of the riboswitch is favored in which the expression of the operon is inhibited (most likely due to premature termination at the predicted termination site). The presence of CsrA in contrast might stabilize the ON state where the structure of the thi-element is dispersed, hence, higher amounts of TPP would be necessary to shift the element back to the OFF state leading to the down-regulation of the <i>thi</i>-genes expression.</p

CsrA interacts directly with <i>lqsR</i> mRNA <i>in vitro</i>.

<p><b>A)</b> Electromobility shift assay (EMSA) with 200nM of biotinylated <i>lqsR</i> mRNA combined with varying concentrations of purified CsrA-His <i>lqsR</i> mRNA and recombinant CsrA in 6% Native Tris-PAGE. Lane 1: no CsrA, lane 2: 0.2 μM CsrA, lane 3: 0.5 μM CsrA, lane 4: 1.0 μM CsrA, lane 5: 2.0 μM CsrA, lane 6: 5.0 μM CsrA, lane 7: 5.0 μM CsrA + 2.0 μM unlabled RsmZ. <b>B)</b> Mfold secondary structure prediction of the <i>lqsR</i> mRNA fragment used for the EMSA. Red, two potential CsrA-binding sites, which are mutated in C. Blue, transcriptional start codon. <b>C</b>) EMSA with recombinant CsrA and 200 nM of non-mutated RNA (pLqsR) or mutated in the indicated regions (mLqsR). AGGA motifs were replaced by an AAAA sequence using PCR mutagenesis. Lane 1: no CsrA + non-mutated <i>lqsR</i> mRNA, lane 2: 5.0 μM CsrA + non-mutated <i>lqsR</i> mRNA, lane 2: 5.0 μM CsrA + m<i>lqsR</i> mRNA mutated in region 1, lane 3: 5.0 μM CsrA + m<i>lqsR</i> mRNA mutated in region 2, lane 4: 5.0 μM CsrA + m<i>lqsR</i> mRNA mutated in both region 1 and 2.</p

Half of the <i>L</i>. <i>pneumophila</i> proteins are differentially expressed upon CsrA deletion.

<p>Protein intensities in the wt and <i>csrA</i>^<i>-</i> strains (three biological replicates, <i>csrA</i>^<i>-</i> = Mut1-3; wt = WT1-3) were measured by differential shotgun proteomics and visualized in a heat map (left) and a profile plot (right) after non-supervised hierarchical clustering. Every row represents a quantified protein (n = 1448) for which the normalized (LFQ) intensity in each biological replicate is color indicated in the columns.</p

Summary of the genes influenced by CsrA and discussed in detail.

<p>Summary of the genes influenced by CsrA and discussed in detail.</p

Glyceraldehyde 3-phosphate (Gap) and transketolase (Tkt) transcription is regulated differently by CsrA.

<p><b>A</b>) Schematic organization of the PPP/Glycolysis-operon in <i>L</i>. <i>pneumophila</i> Paris. TSS indicates the transcriptional start site under the control of an RpoD-dependent promoter. Green, bold arrows show the CsrA-binding region and black arrows highlight the region where qRT-PCR was conducted. <b>B</b>) EMSA with 200nM of biotinylated RNA demonstrating the interaction of purified CsrA and <i>gap</i> mRNA: Lane 1: no CsrA, lane 2: 0.5 μM CsrA, lane 3: 1.0 μM CsrA, lane 4: 2.0 μM CsrA, lane 5: 5.0 μM CsrA, lane 6: 5.0 μM CsrA + 2.0 μM unlabled RsmZ. Right side, run-off transcript produced under optimal <i>in vitro</i> transcription conditions performed with the MEGAshortscript Kit (ambion) to show transcript length compared to the low range ssRNA ladder (NEB). <b>C</b>) qRT-PCR results of the <i>gap</i> and the <i>tkt</i> transcripts at different growth stages (OD) between wt and <i>csrA</i>^- show lower expression levels of the <i>gap</i> gene in E-phase (OD1-3) in absence of CsrA whereas <i>tkt</i> is not affected. No differences are noticed during transition (OD3) and PE-phase. Complementation of the <i>csrA</i>^- strain restored the wt transcript levels</p