Journal: PLoS ONE

Article Title: Host-Pathogen O-Methyltransferase Similarity and Its Specific Presence in Highly Virulent Strains of Francisella tularensis Suggests Molecular Mimicry

doi: 10.1371/journal.pone.0020295

Figure Lengend Snippet: A) Dotplot comparison of pairwise alignments between whole-genome sequences from representatives of various Francisella subspecies is shown. The o-methyltransferase ortholog is highlighted on the Francisella tularensis subsp. holarctica strain 257 in orange (Y-axis) and on the Francisella tularensis subsp tularensis SchuS4 strain (X-axis, reference genome) in red. Blocks of synteny between each of the Francisella genomes compared to the Francisella tularensis subsp tularensis SchuS4 strain are plotted and rearrangements are indicated by breaks in the linearity of the lines and perpendicular orientations.

Article Snippet: Whole genome alignments and dotplot comparisons are provided by the Broad Institute and the methodology for the Francisella comparative genome project is published .

Techniques: Comparison

Journal: Biomolecules

Article Title: [Pt( O , O ′-acac)(γ-acac)(DMS)] Induces Autophagy in Caki-1 Renal Cancer Cells

doi: 10.3390/biom9030092

Figure Lengend Snippet: Cytotoxic effects of [Pt( O , O ′-acac)(γ-acac)(DMS) and cisplatin and effects on the cell cycle of Caki-1 cells. Caki-1 cells were treated with 10 µM [Pt( O , O ′-acac)(γ-acac)(dimethyl sulfide (DMS))] or with 50 µM cisplatin. Cell viability was monitored by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenol tetrazolium bromide (MTT) assay ( A ) and cell death was quantified by fluorescence-activated cell sorter (FACS) after propidium iodide (PI)/annexin V-fluorescein isothiocyanate (FITC) staining ( B ), over a period of 48 h. Data are means ± standard deviation (SD) of five independent experiments with eight replicates in each, and are presented as percent of control. * p < 0.01 between treated and untreated cells (white bar), by Student’s t -test. ( C ) Cells were treated with [Pt( O , O ′-acac)(γ-acac)(DMS)] or cisplatin for 24 h, and cell cycle distribution was analyzed by flow cytometry after staining the cells with PI. Representative FACS histogram from six separate experiments is shown.

Article Snippet: Total cell death was quantified by adding the percentage of cells detected in the upper left (PI), upper right (PI + annexin V-FITC), and lower right (annexin V-FITC) quadrants in the FACS dotplots (Becton-Dickinson, CA, USA).

Techniques: MTT Assay, Fluorescence, Staining, Standard Deviation, Control, Flow Cytometry

Journal: Biomolecules

Article Title: [Pt( O , O ′-acac)(γ-acac)(DMS)] Induces Autophagy in Caki-1 Renal Cancer Cells

doi: 10.3390/biom9030092

Figure Lengend Snippet: [Pt( O , O ′-acac)(γ-acac)(DMS)] does not induce apoptosis in Caki-1 cells. ( A ) Expression of cleaved caspase-9 and -3 in Caki-1 cells. Cell were treated with 10 µM of [Pt( O , O ′-acac)(γ-acac)(DMS)] or 50 µM of cisplatin for the indicated time, and then subjected to Western blotting. Incubation with anti-β-actin confirmed the equal protein loading. The results shown are representative of three different experiments. ( B ) (Up) Caki-1 cells were treated, or not, with cisplatin, or with [Pt( O , O ′-acac)(γ-acac)(DMS)] for 24 h, and then stained with 4,6-diammine-2-phenylindol (DAPI); the representative fields by confocal microscopy (magnification 40×) of one of four independent experiments are shown. (Down) Quantification of the percentage of apoptotic nuclei obtained from cells stained with DAPI (means ± SD; n = 4), after treatment for different times with 10 µM [Pt( O , O ′-acac)(γ-acac)(DMS)] or 50 µM cisplatin. ** p < 0.01 between cisplatin-treated and untreated cells and * p < 0.05 between [Pt( O , O ′-acac)(γ-acac)(DMS)]-treated and untreated cells, by Student’s t -test. ( C ) Visualization of DNA fragmentation in [Pt( O , O ′-acac)(γ-acac)(DMS)]-treated Caki-1 cells. Total DNA was isolated and separated on a 1% agarose gel. A representative example of three independent experiments is shown. ( D , E ) Caki-1 cells were transfected with 30 nM small interfering RNA (siRNA) oligos for apoptosis inducing factor (AIF) and then treated with 10 µM [Pt( O , O ′-acac)(γ-acac)(DMS)]. ( D ) Immunoblotting detection of AIF in cell extracts 48 h after siRNA transfection using polyclonal anti-AIF antibody. Controls were provided by untransfected cells and cells transfected with scrambled siRNA oligos (Scr). Incubation with anti-β-actin confirmed the equal protein loading. A representative example of three independent experiments is shown. ( E ) Cell death was quantified by FACS after propidium iodide (PI)/annexin V-FITC staining, in transfected Caki-1 cells treated with 10 µM of [Pt( O , O ′-acac)(γ-acac)(DMS)] or 50 µM cispltin for 24 h. Data are means ± SD of five independent experiments.

Article Snippet: Total cell death was quantified by adding the percentage of cells detected in the upper left (PI), upper right (PI + annexin V-FITC), and lower right (annexin V-FITC) quadrants in the FACS dotplots (Becton-Dickinson, CA, USA).

Techniques: Expressing, Western Blot, Incubation, Staining, Confocal Microscopy, Isolation, Agarose Gel Electrophoresis, Transfection, Small Interfering RNA