eIF2α dephosphorylation and cell proliferation in CHO-K1 cells

GADD34 is a member of a growth arrest and DNA damage (GADD)-inducible gene family. Here, we established a novel Chinese hamster ovary (CHO-K1)-K1-derived cell line, CHO-K1-G34M, which carries a nonsense mutation (termed the Q525X mutation) in the GADD34 gene. The Q525X mutant protein lacks the C-terminal 66 amino acids required for GADD34 to bind to and activate protein phosphatase 1 (PP1). We investigated the effects of GADD34 with or without the Q525X mutation on the phosphorylation status of PP1 target proteins, including the α subunit of eukaryotic initiation factor 2 (eIF2α) and glycogen synthase kinase 3β (GSK3β). CHOK1-G34M cells had higher levels of eIF2α phosphorylation compared to the control CHO-K1-normal cells both in the presence and absence of endoplasmic reticulum stress. Overexpression of wild type GADD34 protein in CHOK1-normal cells largely reduced eIF2α phosphorylation, while overexpression of the Q525X mutant did not produce similar reductions. Meanwhile, neither wild type nor Q525X mutation of GADD34 affected the GSK3β phosphorylation status. GADD34 also did not affect the canonical Wnt signaling pathway downstream of GSK3β. Cell proliferation rates were higher, while expression levels of the cyclin dependent kinase inhibitor p21 were lower in CHO-K1-G34M cells compared to the CHO-K1-normal cells. The GADD34 Q525X mutant had a reduced ability to inhibit cell proliferation and enhance p21 expression of the CHO-K1-normal cells compared to the wild type GADD34 protein. These results suggest that the GADD34 protein C-terminal plays important roles in regulating not only eIF2α dephosphorylation but also cell proliferation in CHO-K1 cells

Cell Wall Trapping of Autocrine Peptides for Human G-Protein-Coupled Receptors on the Yeast Cell Surface

G-protein-coupled receptors (GPCRs) regulate a wide variety of physiological processes and are important pharmaceutical targets for drug discovery. Here, we describe a unique concept based on yeast cell-surface display technology to selectively track eligible peptides with agonistic activity for human GPCRs (Cell Wall Trapping of Autocrine Peptides (CWTrAP) strategy). In our strategy, individual recombinant yeast cells are able to report autocrine-positive activity for human GPCRs by expressing a candidate peptide fused to an anchoring motif. Following expression and activation, yeast cells trap autocrine peptides onto their cell walls. Because captured peptides are incapable of diffusion, they have no impact on surrounding yeast cells that express the target human GPCR and non-signaling peptides. Therefore, individual yeast cells can assemble the autonomous signaling complex and allow single-cell screening of a yeast population. Our strategy may be applied to identify eligible peptides with agonistic activity for target human GPCRs

The Discovery of LOX-1, its Ligands and Clinical Significance

LOX-1 is an endothelial receptor for oxidized low-density lipoprotein (oxLDL), a key molecule in the pathogenesis of atherosclerosis.The basal expression of LOX-1 is low but highly induced under the influence of proinflammatory and prooxidative stimuli in vascular endothelial cells, smooth muscle cells, macrophages, platelets and cardiomyocytes. Multiple lines of in vitro and in vivo studies have provided compelling evidence that LOX-1 promotes endothelial dysfunction and atherogenesis induced by oxLDL. The roles of LOX-1 in the development of atherosclerosis, however, are not simple as it had been considered. Evidence has been accumulating that LOX-1 recognizes not only oxLDL but other atherogenic lipoproteins, platelets, leukocytes and CRP. As results, LOX-1 not only mediates endothelial dysfunction but contributes to atherosclerotic plaque formation, thrombogenesis, leukocyte infiltration and myocardial infarction, which determine mortality and morbidity from atherosclerosis. Moreover, our recent epidemiological study has highlighted the involvement of LOX-1 in human cardiovascular diseases. Further understandings of LOX-1 and its ligands as well as its versatile functions will direct us to ways to find novel diagnostic and therapeutic approaches to cardiovascular disease

Volcanic plume measurements using a UAV for the 2014 Mt. Ontake eruption

A phreatic eruption of Mt. Ontake, Japan, started abruptly on September 27, 2014, and caused the worst volcanic calamity in recent 70 years in Japan. We conducted volcanic plume surveys using an electric multirotor unmanned aerial vehicle to elucidate the conditions of Mt. Ontake’s plume, which is flowing over 3000 m altitude. A plume gas composition, sulfur dioxide flux and thermal image measurements and a particle sampling were carried out using the unmanned aerial vehicle for three field campaigns on November 20 and 21, 2014, and June 2, 2015. Together with the results of manned helicopter and aircraft observations, we revealed that the plume of Mt. Ontake was not directly emitted from the magma but was influenced by hydrothermal system, and observed SO2/H2S molar ratios were decreasing after the eruption. High SO2 flux of >2000 t/d observed at least until 20 h after the onset of the eruption implies significant input of magmatic gas and the flux quickly decreased to about 130 t/d in 2 months. In contrast, H2S fluxes retrieved using SO2/H2S ratio and SO2 flux showed significantly high level of 700–800 t/d, which continued at least between 2 weeks and 2 months after the eruption. This is a peculiar feature of the 2014 Mt. Ontake eruption. Considering the trends of the flux changes of SO2 and H2S, we presume that majority of SO2 and H2S are supplied, respectively, from high-temperature magmatic fluid of a deep origin and from hydrothermal system. From the point of view of SO2/H2S ratios and fumarolic temperatures, the plume degassing trend after the 2014 eruption is following the similar course as that after the 1979 eruptions, and we speculate the 2014 eruptive activity will cease slowly similar to the 1979 eruption

Preparation of Templated Mesoporous Soft Carbon from Synthesized Quinoline Pitch

A mesoporous soft carbon has been prepared using a novel method based on aggregation to metal nanoparticles, along with heat treatment of a nitrogen-rich precursor designated as quinolone pitch. From this newly prepared method using quinolone pitch and nickel acetate, mesoporous soft carbon has been prepared. This mesoporous soft carbon exhibits increased capacitance by anodic oxidation in non-aqueous lithium salt electrolyte, even though the activation effect was limited compared with previous reports