RNA interference is not affected by the absence of the QKI-6 isoform in U343 cells.

<p>(A) Control siRNA, siQKI-6 or siAgo2 were transfected in U343 cells and 24 hr later, the pMIR luciferase plasmid was co-transfected with the renilla luciferase vector and an siRNA that targets the pgl2 against firefly luciferase or a control siRNA. Thirty hours later, the cells were lysed and subjected to luciferase assays or immunoblotting analysis panel B. The firefly luciferase activity was normalized with renilla luciferase. (B) Cellular extracts from transfected U343 cells from panel (A) were immunoblotted with anti-Ago2 and anti-QKI antibodies. Immunoblotting against anti-Sam68 antibodies was used as a loading control. Absolute quantified levels of protein bands are presented underneath each blot and were performed using Scion-Image.</p

QKI-6 co-localizes with MBP mRNA in stress granules in primary rat oligodendrocytes.

<p>(A) Primary rat oligodendrocytes were left untreated or treated with 0.5 mM As₂O₃ for 45 min. The cells were fixed and hybridized with a digoxigenin-11-UTP labeled MBP antisense RNA probe (green) and immunostained with anti-QKI-6 antibodies (red). The scale bar represents 5 µm. (B) The quantification of the co-localization between the QKI isoforms and MBP mRNA expressed as a percentage is shown.</p

QKI-6 and QKI-7 co-localize with PABP1 in stress granules in glial cells.

<p>(A) U343 cells were untreated or treated with 0.5 mM As₂O₃ for 45 min. The cells were fixed, permeabilized and immunostained with rabbit anti-QKI-5, -6 and -7 antibodies and a mouse anti-PABP1 followed by secondary goat anti-rabbit Alexa523 (red) and goat anti-mouse Alexa 488 (green) antibodies. The nuclei were stained with DAPI. The scale bar represents 5 µm. (B) The quantification of the co-localization between the QKI isoforms and PABP1 expressed as a percentage is shown. (C) Primary rat oligodendrocytes were untreated or treated with 0.5 mM As₂O₃ for 45 min and analyzed as in panel (A). (D) The co-localization was performed as in panel (B).</p

Mapping the domains required for the QKI-6/Ago2 interaction.

<p>(A) A schematic illustration of the QKI-6 protein showing its regions and its amino acid numbering. Expression vectors encoding GFP-QKI-6 and truncation mutants thereof were transfected in HEK293 cells. The transfected cells were lysed and the cell lysates were subjected to immunoprecipitation with the anti-GFP antibody and the bound proteins separated by SDS-PAGE. The presence of Ago2 was monitored by using anti-Ago2 antibodies as indicated (left panel). Extracts before immunoprecipitation were separated by SDS-PAGE and immunoblotted with anti-GFP antibodies to confirm equivalent expression. The molecular mass markers are shown on the left in kDa. (B) A schematic illustration of Ago2 is shown with its conserved domains and the numbering of its residues. The GFP-QKI-6 expression plasmid was cotransfected with myc-tagged full-length Ago2 or truncation mutants in HEK293 cells, as indicated. Twenty four hours after transfection, the cells were lysed and cell lysates were subjected to immunoprecipitation with anti-Myc antibodies followed by immunoblotting with anti-GFP antibodies. The migration of GFP-QKI-6 is shown (left panel), while the expression of the myc-Ago2 proteins is shown in the right panel. The asterisks denote non-specific proteins recognized by the anti-myc antibodies. The molecular mass markers are shown on the left in kDa.</p

The endogenous QKI isoforms associate with Ago2.

<p>(A) U343 cell lysates were subjected to immunoprecipitations (IP) with control immunoglobulin G (IgG), anti-QKI-5, -QKI-6 and -QKI-7 antibodies. The bound proteins were separated by SDS-PAGE and immunoblotted (IB) with anti-Ago2 antibodies. (B) U343 cell lysates were treated with 1 mg/ml RNase A, 2 U/100 µl RNase V1 or RNase inhibitor as indicated at 37°C for 1 hr and subjected to immunoprecipitation with the anti-QKI-6 antibody. The proteins were separated by SDS-PAGE and immunoblotted with anti-Ago-2 antibodies as indicated (upper panel). The activity of the RNases and the RNase inhibitor was verified by agarose gel electrophoresis with 10 µg of total RNA (lower panel).</p

DCP-1 P bodies are devoid of QKI-7.

<p>Myc-QKI-7 and Flag-DCP1 were co-transfected in HEK293 cells. Thirty hours later, the cells were left untreated or treated with 0.5 mM As₂O₃ for 30 min. The cells were fixed, permeabilized and immunostained with rabbit anti-Myc antibody and mouse anti-Flag antibody followed by goat anti-rabbit Alexa523 (red) and goat anti-mouse Alexa 488 (green). The nuclei were counter-stained with DAPI. The scale bar represents 10 µm.</p

QKI-6 and QKI-7 isoforms co-localize with Ago2 in cytoplasmic granules in glial cells.

<p>(A) U343 cells were untreated or treated with 0.5 mM arsenic oxide (As₂O₃) for 45 min. The cells were fixed, permeabilized and immunostained with rabbit anti-QKI-5, -6 and -7 antibodies and mouse anti-Ago2 antibodies followed by secondary goat anti-rabbit Alexa 488 (green) and goat anti-mouse Alexa 523 (red) antibodies. The nuclei were counter-stained with DAPI. The scale bar represents 5 µm. (B) The quantification of the co-localization between the QKI isoforms and Ago2 expressed as a percentage is shown. (C) Primary rat oligodendrocytes were untreated or treated with 0.5 mM As₂O₃ for 45 min and analyzed as in panel (A). (D) The quantification was performed as in panel (B).</p

Media 1: Enhanced robustness digital holographic microscopy for demanding environment of space biology

Originally published in Biomedical Optics Express on 01 February 2012 (boe-3-2-313

Media 2: Enhanced robustness digital holographic microscopy for demanding environment of space biology

Originally published in Biomedical Optics Express on 01 February 2012 (boe-3-2-313

Media 3: Enhanced robustness digital holographic microscopy for demanding environment of space biology

Originally published in Biomedical Optics Express on 01 February 2012 (boe-3-2-313