Aβ42 induced rough eye phenotype is sensitive to <i>Toll</i> activity.

<p>WT: Wild type flies. Aβ: pGMR-Aβ42 transgenic flies. In the experiments shown in this figure, the flies are hemizygous for the pGMR-Aβ42 transgene. Panel A: WT: eye of a wild type fly. Aβ/+: eye of a wild type fly hemizygous for the pGMR-Aβ42 transgene. Panels B through F show sibling pGMR-Aβ42 transgene flies that are either wild type <i>Tl</i> or heterozygous for the indicated <i>Tl</i> allele. Panel B: Aβ/+: hemizygous for the Aβ transgene; Aβ/<i>Tl^r4</i>: heterozygous for the LOF allele <i>Tl^r4</i>. Panel C: Aβ/+: hemizygous for the Aβ transgene; Aβ/<i>Tl^r3</i>: heterozygous for the LOF allele <i>Tl^r4</i>. Panel D: Aβ/+: hemizygous for the Aβ transgene; Aβ/<i>Tl^KG</i>: heterozygous for the LOF allele <i>Tl^KG03609</i>. Panel E: Aβ/+: hemizygous for the Aβ transgene; Aβ/<i>Tl^rk</i>: heterozygous for the LOF allele <i>Tl^rK343</i>. Panel F: Aβ/+: hemizygous for the Aβ transgene; Aβ/<i>Tl³</i>: heterozygous for the GOF allele <i>Tl^r3</i>.</p

The <i>Drosophila</i> Toll→Dorsal/Dif and mammalian Interleukin Receptor→NFκB innate immunity pathways.

<p>Diagram of the <i>Tl</i>→NFκB signaling pathways in <i>Drosophila</i> and mammals. See text for details.</p

Dorsal mediates Aβ42 dependent neurodegeneration of the eye.

<p>Panels A–E: In the experiments shown in this figure all flies are siblings that are homozygous for the pGMR-Aβ42 transgene. The fly on the left side of each panel is a wild type sib, while the fly on the right side of each panel is a sib that is either heterozygous or homozygous for the indicated <i>dl</i> mutation. Panel A: <i>dl¹</i>/+. Panel B: <i>dl⁴</i>/+ Panel C: <i>dl⁸</i>/+ Panel D: <i>dl¹</i>/<i>dl¹</i>. Panel E: <i>dl⁴/dl⁴</i>. Panel F: In this experiment both flies are hemizygous for the pGMR-Aβ42 transgene. The fly on the left is wild type, while the fly on the right is homozygous <i>dl^`</i>. Arrows in panels A and E point to ommatidia with dead or dying cells.</p

Effect of mutations in the <i>Tl→ΝFκB</i> pathway on the Aβ42 induced rough eye phenotype.

<p>The rough eye phenotype of pGMR-Aβ42 flies was assigned a value of ++++. According to this scoring system, strong, moderate and weak suppression corresponded to +, ++, and +++. Weak and moderate enhancement corresponded to +++++ and ++++++. All flies are hemizygous for the transgene and either wild type (+) or heterozygous for the indicated mutation.</p

Components of the <i>Toll-NF</i>κ<i>B</i> signaling pathway modulate Aβ42 polypetide induced neurodegeneration.

<p>Panels A–F: In the experiments shown in this figure all flies are siblings that are hemizygous for the pGMR-Aβ42 transgene. The fly on the left side of each panel is a wild type sib, while the fly on the right side of each panel is a sib that is heterozygous for the indicated mutation in the <i>Toll-NF</i>κ<i>B</i> signaling pathway mutant. Panel A: <i>dl¹</i>. Panel B: <i>dl⁴</i>. Panel C: <i>dl⁸</i>. Panel D: <i>pll²</i>. Panel E: <i>pll⁷</i>. Panel F: <i>tub</i>.</p

Statistics from data collection and model refinement.

a<p>Values in parentheses refer to the shell with the highest resolution.</p>b<p><i>R</i>_merge = Σ_h Σ_i | <i>I</i>(<i>h</i>)_i –<<i>I</i>(<i>h</i>)>|/Σ_h Σ_i<i>I</i>(<i>h</i>)_i, where <i>I</i>(<i>h</i>) is the intensity for reflection <i>h</i>, Σ_h is the sum for all reflections, and Σ_i is the sum for i measurements of reflection <i>h</i>.</p>c<p><i>R</i> = Σ | |<i>F</i>_obs| – |<i>F</i>_calc| |/Σ |<i>F</i>_obs|, where <i>R</i>_free is calculated for a randomly chosen 5% of reflections, which were not used for structure refinement, and <i>R</i>_work is calculated for the remaining reflections.</p>d<p>Values generated using Refmac <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0070358#pone.0070358-Murshudov1" target="_blank">[20]</a>.</p>e<p>Values generated using MolProbity <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0070358#pone.0070358-Chen1" target="_blank">[21]</a>.</p

The SKI-O-068 binding mode in the Pim1 active site.

<p>(A) SKI-O-068 intermolecular interactions. The two water molecules (W1 and W2) are from the SKI-O-068-bound Pim1 structure. The additional water molecules (W1’ and W2’) are from the native Pim1 structure. The water molecules are depicted as spheres in red. The polar interactions are depicted using gray dashes. (B) The Pim1 surface diagram depicting the active site pocket. Three inhibitors from <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0070358#pone-0070358-g001" target="_blank">Figure 1</a> are superimposed. The molecular surface is depicted in raspberry (hinge), yellow (G-loop), blue (A-loop) and pink (catalytic loop). (C) Detailed view of three inhibitors and two residues (Lys67 and Glu121).</p

The pimtide binding mode at the substrate-binding pocket.

<p>(A) Superposition of pimtide molecules in pimtide-bound structures. PDB codes 2BIL (purple), 2C3I (cyan), 3CY2 (light pink) and 3MA3 (ivory) were used for structural comparison. (B) The pimtide binding mode (green). The <i>Fo</i> – <i>Fc</i> electron density map is contoured at 2.5 σ and colored in gray. The polar interactions are depicted using gray-colored dashes.</p

Overall structure for the complex of Pim1, SKI-O-068, and pimtide.

<p>(A) Ribbon diagram for the monomeric Pim1 structure. The hinge region, G-loop, A-loop and catalytic loop are colored raspberry, yellow, blue and pink, respectively. (B) Electrostatic potential surface diagram for the monomeric Pim1 structure. SKI-O-068 and pimtide are shown in orange and green, respectively. The inhibitor is in the active site, and pimtide is in the active-binding pocket vicinity. The structure figures and surface diagram were drawn using PyMOL (DeLano WL, 2002, The PyMOL Molecular Graphics System. DeLano Scientific, Palo Alto, California).</p

Structures for the ligands analyzed in this study.

<p>(A) Chemical structure of SKI-O-068. Three moieties on the ligand are indicated by blue-colored dashes [(a) the trifluoromethyl benzene moiety, (b) cyclohexanamine moiety and (c) pyrido[4,3-<i>d</i>]pyrimidin-5(6<i>H</i>)-<i>one</i>] moiety. (B) Two chemical scaffolds similar to SKI-O-068 (the triazolopyridazine scaffold from PDB code 3BGQ and triazolopyridine scaffold from PDB code 4A7C).</p