Orb6 phosphorylation site mutants impair interaction with Nak1.

<p>Extracts from WT (SP199) cells expressing HA-Orb6 and control vector (lane 1), Myc-Nak1 and control vector (lane 2), HA-Orb6 and Myc-Nak1 (lane 3), HA-Orb6 ^S291A and Myc-Nak1 (lane 4), or HA-Orb6^T456A and Myc-Nak1 (lane 5) from <i>nmt1</i> promoter expression plasmids were analyzed by western-blots using anti-Myc (9E10) or anti-HA (12CA5) monoclonal antibodies (Input panels). HA-Orb6 was immunoprecipitated from cell extracts with anti-HA antibody and equal portions of the immunoprecipitates were probed with anti-HA antibody and anti-Myc antibody (αHA IP panels).</p

Orb6 overexpression partially rescues the <i>nak1</i> ts polarity defect.

<p>HA-Nak1 or HA-Orb6 were expressed in WT (SP199), <i>nak1</i> and <i>orb6</i> ts cells as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0037221#pone-0037221-g001" target="_blank">Figure 1</a>. Cells were grown in thiamine free liquid PMAU media to an O.D.  = 1.0 at 33°C and examined by DIC microscopy.</p

Expression of Nak1 and Orb6 mutants.

<p>HA-Nak1, HA-Orb6, HA-Orb6^T456A, HA-Orb6^S291A, and HA-Orb6^K122A were expressed in WT (SP199), <i>nak1</i> and <i>orb6</i> ts cells using plasmids derived from pREP3X containing the thiamine repressible <i>nmt1</i> promoter (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0037221#s4" target="_blank">Materials and Methods</a>). HA-tagged proteins were detected by Western blots using anti-HA (12CA5) monoclonal antibody.</p

Nak1 and Orb6 interact <i>in vitro</i> and <i>in vivo</i>. A.

<p>Extracts from WT (SP199) cells expressing HA-Orb6 and control vector (lane 1), Myc-Nak1 and control vector (lane 2), or HA-Orb6 and Myc-Nak1 (lane 3) from <i>nmt1</i> promoter expression plasmids were analyzed by western-blots using anti-Myc (9E10) or anti-HA (12CA5) monoclonal antibodies (Input panels). HA-Orb6 was immunoprecipitated from cell extracts with anti-HA antibody and equal portions of the immunoprecipitates were probed with anti-HA antibody and anti-Myc antibody (αHA IP panels). <b>B.</b> Interaction of purified recombinant GST-Orb6 and His₆-HA-Nak1 was assayed by an <i>in vitro</i> binding assay (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0037221#s4" target="_blank">Materials and Methods</a>). The left panel shows Coomassie blue staining of purified recombinant GST (lane 1), GST-Orb6 (lane 2), His₆-vector control (lane 3), His₆-HA-Nak1 (lane 4). Purified His₆-HA-Nak1 (input, lane 7) was incubated with either GST (lane 5) or GST-Orb6 (lane 6) bound to Glutathione Sepharose 4B beads. The right panel shows a Western blot using anti-HA (12CA5) antibody of His₆-HA-Nak1 bound to the beads. <b>C.</b> Schematic diagram of mutant Nak1 expression constructs. The Nak1 N-terminal kinase domain (residues 1–262) and C-terminal (CTR) region (554–652) are indicated. The numbers at the left and the bars at the right indicate the region of Nak1 encoded by the various deletion constructs. <b>D.</b> Extracts from wild-type (SP199) cells co-expressing Myc-Orb6 with the control vector (lane 1), HA-Nak1^1–562 (lane 2), HA-Nak1^1–585 (lane 3), HA-Nak1^1–607 (lane 4), HA-Nak1^1–629 (lane 5), HA-Nak1 (lane6), or HA-Nak1^554–652 (lane 7) were analyzed by western-blots using anti-Myc (9E10) or anti-HA (12CA5) monoclonal antibodies (Input panels). Extracts were immunoprecipitated with anti-HA antibody and equal portions of the immunoprecipitates were probed with anti-HA antibody and anti-Myc antibody (αHA IP panels).</p

Orb6 phosphorylation <i>in vitro</i>.

<p><b>A.</b> Coomassie blue staining of purified recombinant GST-Nak1 (lane 1), GST-Orb6 (lane 2), GST-Orb6^301–469 (GST-Orb6C) (lane 3). <b>B.</b> Kinase assays were performed using purified recombinant GST-Orb6 and immunoprecipitated HA-Nak1 from yeast cell extracts (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0037221#s4" target="_blank">Materials and Methods</a>). Protein A-Sepharose beads bound with HA-Nak1 were removed from samples in lanes 3, 5 following the kinase reaction. HA + Casein (lane 1), HA-Nak1 + Casein (lane 2), HA-Nak1 + Casein (lane 3), HA + GST-Orb6 (lane 4), HA-Nak1 + GST-Orb6 (lane 5). <b>C.</b> Kinase assays were performed using purified recombinant GST-Orb6C and immunoprecipitated HA-Nak1 from yeast cell extracts. HA (lane 1), HA-Nak1 (lane 2), HA-Nak1 + GST-Orb6C (lane 3). <b>D.</b> Kinase assays were performed using purified Orb6 lacking GST and immunoprecipitated HA-Nak1. HA + Orb6 (lane 1), HA-Nak1 + Orb6 (lane 2). <b>E.</b> Kinase assays were performed using purified recombinant GST-Orb6 (lane 1), GST-Orb6^T456A (lane 2), GST-Orb6^S291A (lane 3) and GST-Orb6^K122A (lane 4).</p

Very Bright and Efficient Microcavity Top-Emitting Quantum Dot Light-Emitting Diodes with Ag Electrodes

The microcavity effect in top-emitting quantum dot light-emitting diodes (TQLEDs) is theoretically and experimentally investigated. By carefully optimizing the cavity length, the thickness of the top Ag electrode and the thickness of the capping layer, very bright and efficient TQLEDs with external quantum efficiency (EQE) of 12.5% are demonstrated. Strong dependence of luminance and efficiency on cavity length is observed, in good agreement with theoretical calculation. By setting the normal-direction resonant wavelength around the peak wavelength of the intrinsic emission, highest luminance of 112 000 cd/m² (at a driving voltage of 7 V) and maximum current efficiency of 27.8 cd/A are achieved, representing a 12-fold and a 2.1-fold enhancement compared to 9000 cd/m² and 13.2 cd/A of the conventional bottom emitting devices, respectively, whereas the highest EQE of 12.5% is obtained by setting the resonant wavelength 30 nm longer than the peak wavelength of the intrinsic emission. Benefit from the very narrow spectrum of QDs and the low absorption of silver electrodes, the potential of microcavity effect can be fully exploited in TQLEDs

Performance of Inverted Quantum Dot Light-Emitting Diodes Enhanced by Using Phosphorescent Molecules as Exciton Harvesters

Exciton harvesters based on blue phosphorescent molecules bis­(4,6-difluorophenylpyridinato-<i>N</i>,C2)­picolinatoiridium (FIrpic) doped in 4,4′,4″-tris­(carbazol-9-yl)­triphenylamine (TCTA) are used to enhance the performance of inverted quantum dot light-emitting diodes (QD-LEDs). In the proposed device structures, electrons that leak to the TCTA layer can be effectively captured by FIrpic and subsequently can recombine in the TCTA:FIrpic layer. The harvested energy is then nonradiatively transferred to the adjacent QDs via Förster dipole–dipole coupling mechanism. Because of effective harvest of leaked electrons and complete energy transfer from FIrpic to the adjacent QDs, the demonstrated QD-LEDs exhibit pure QD emission, higher efficiency (1.62-fold improvement), and longer lifetime

Enhanced Oxygen and Hydroxide Transport in a Cathode Interface by Efficient Antibacterial Property of a Silver Nanoparticle-Modified, Activated Carbon Cathode in Microbial Fuel Cells

A biofilm growing on an air cathode is responsible for the decreased performance of microbial fuel cells (MFCs). For the undesired biofilm to be minimized, silver nanoparticles were synthesized on activated carbon as the cathodic catalyst (Ag/AC) in MFCs. Ag/AC enhanced maximum power density by 14.6% compared to that of a bare activated carbon cathode (AC) due to the additional silver catalysis. After operating MFCs over five months, protein content on the Ag/AC cathode was only 38.3% of that on the AC cathode, which resulted in a higher oxygen concentration diffusing through the Ag/AC cathode. In addition, a lower pH increment (0.2 units) was obtained near the Ag/AC catalyst surface after biofouling compared to 0.8 units of the AC cathode, indicating that less biofilm on the Ag/AC cathode had a minor resistance on hydroxide transported from the catalyst layer interfaces to the bulk solution. Therefore, less decrements of the Ag/AC activity and MFC performance were obtained. This result indicated that accelerated transport of oxygen and hydroxide, benefitting from the antibacterial property of the cathode, could efficiently maintain higher cathode stability during long-term operation

ILTox: A Curated Toxicity Database for Machine Learning and Design of Environmentally Friendly Ionic Liquids

A comprehensive online database on the toxicity of ionic liquids (ILs) is urgently needed to facilitate machine learning to design environmentally friendly ILs. In this direction, we present ILTox, a manually curated database of 1183 ILs with over 6700 pieces of toxicity data across different living organisms, including mammalian cells, bacteria, and plants. All the toxicity values and structural information on ILs have been rigorously assessed to ensure data quality. Using this database, various machine learning models have been constructed to quantitatively analyze the relationship between the ILs’ structures and their toxicities. Furthermore, the optimized models were used for a virtual screening of desired properties from 8 million ILs. Our results demonstrated that the ILTox database could accelerate the transformation of toxicity data into critical structure–toxicity relationship knowledge. As far as we know, ILTox is the only available database on IL toxicity and is now openly accessible at http://www.iltox.com

ILTox: A Curated Toxicity Database for Machine Learning and Design of Environmentally Friendly Ionic Liquids

A comprehensive online database on the toxicity of ionic liquids (ILs) is urgently needed to facilitate machine learning to design environmentally friendly ILs. In this direction, we present ILTox, a manually curated database of 1183 ILs with over 6700 pieces of toxicity data across different living organisms, including mammalian cells, bacteria, and plants. All the toxicity values and structural information on ILs have been rigorously assessed to ensure data quality. Using this database, various machine learning models have been constructed to quantitatively analyze the relationship between the ILs’ structures and their toxicities. Furthermore, the optimized models were used for a virtual screening of desired properties from 8 million ILs. Our results demonstrated that the ILTox database could accelerate the transformation of toxicity data into critical structure–toxicity relationship knowledge. As far as we know, ILTox is the only available database on IL toxicity and is now openly accessible at http://www.iltox.com