Beclin1-binding UVRAG targets the class C Vps complex to coordinate autophagosome maturation and endocytic trafficking

Autophagic and endocytic pathways are tightly regulated membrane rearrangement processes that are crucial for homeostasis, development and disease. Autophagic cargo is delivered from autophagosomes to lysosomes for degradation through a complex process that topologically resembles endosomal maturation. Here, we report that a Beclin1-binding autophagic tumour suppressor, UVRAG, interacts with the class C Vps complex, a key component of the endosomal fusion machinery. This interaction stimulates Rab7 GTPase activity and autophagosome fusion with late endosomes/lysosomes, thereby enhancing delivery and degradation of autophagic cargo. Furthermore, the UVRAG-class-C-Vps complex accelerates endosome-endosome fusion, resulting in rapid degradation of endocytic cargo. Remarkably, autophagosome/endosome maturation mediated by the UVRAG-class-C-Vps complex is genetically separable from UVRAG-Beclin1-mediated autophagosome formation. This result indicates that UVRAG functions as a multivalent trafficking effector that regulates not only two important steps of autophagy - autophagosome formation and maturation - but also endosomal fusion, which concomitantly promotes transport of autophagic and endocytic cargo to the degradative compartments.     

Antimicrobial treatment of grapes using sodium hypochlorite in winemaking and its effects on the chemical and sensory characteristics of wines

This study was performed to examine the use of NaOCl as an alternative antimicrobial compound in winemaking because of the potential health problems that may arise as a result of the use of SO2. For this, the blank (non-treated), control (SO2-added), and sample (NaOCl-treated) wines were made, and microbial and chemical changes including sensory characteristics were analyzed during the fermentation periods. Treatment of grapes with NaOCl decreased the initial contaminating microbial population in grape must, resulting in higher growth of yeast and lactic acid bacteria. After 200 days of fermentation, the chemical analysis of sample wine revealed that it had higher ethanol content, redness (a*), and concentrations of fruity ester compounds and lower total acidity than the control. In the sensory analyses, the sample wine obtained a higher overall acceptability score (5.70) than the control (4.26). This result reveals that NaOCl can be used as an alternative to SO2 in winemaking for inhibiting the growth of contaminating microorganisms.     

Comparison of Cell Wall Polysaccharide Hydrolysis by a Dilute Acid/Enzymatic Saccharification Process and Rumen Microorganisms

Evaluation of biomass crops for breeding or pricing purposes requires an assay that predicts performance in the bioenergy conversion process. Cell wall polysaccharide hydrolysis was compared for a dilute sulfuric acid pretreatment at 121°C followed with cellulase hydrolysis for 72?h conversion assay (CONV) with in vitro rumen microflora incubation for 72?h (RUMEN) for a set of maize (Zea mays L.) stover samples with a wide range in cell wall composition. Residual polysaccharides from the assays were analyzed for sugar components and extent of hydrolysis calculated. Cell wall polysaccharide hydrolysis was different for all sugar components between the CONV and RUMEN assays. The CONV assay hydrolyzed xylose-, arabinose-, galactose-, and uronic acid-containing polysaccharides to a greater degree than did the RUMEN assay, whereas the RUMEN assay was more effective at hydrolyzing glucose- and mannose-containing polysaccharides. Greater hydrolysis of hemicelluloses and pectins by CONV can be attributed to the acid hydrolysis mechanism of the CONV assay for noncellulosic polysaccharides, whereas the RUMEN assay was dependent on enzymatic hydrolysis. While CONV and RUMEN hydrolysis were correlated for most polysaccharide components, the greatest correlation was only r?=?0.70 for glucose-containing polysaccharides. Linear correlations and multiple regressions indicated that polysaccharide hydrolysis by the RUMEN assay was negatively associated with lignin concentration and ferulate ether cross linking as expected. Corresponding correlations and regressions for CONV were less consistent and occasionally positive. Use of rumen microbial hydrolysis to characterize biomass performance in a conversion process may have some limited usefulness for genetic evaluations, but such assays would be unreliable for biomass pricing.     

Selenoprotein P Status Correlates to Cancer-Specific Mortality in Renal Cancer Patients

Selenium (Se) is an essential trace element for selenoprotein biosynthesis. Selenoproteins have been implicated in cancer risk and tumor development. Selenoprotein P (SePP) serves as the major Se transport protein in blood and as reliable biomarker of Se status in marginally supplied individuals. Among the different malignancies, renal cancer is characterized by a high mortality rate. In this study, we aimed to analyze the Se status in renal cell cancer (RCC) patients and whether it correlates to cancer-specific mortality. To this end, serum samples of RCC patients (n = 41) and controls (n = 21) were retrospectively analyzed. Serum Se and SePP concentrations were measured by X-ray fluorescence and an immunoassay, respectively. Clinical and survival data were compared to serum Se and SePP concentrations as markers of Se status by receiver operating characteristic (ROC) curve and Kaplan-Meier and Cox regression analyses. In our patients, higher tumor grade and tumor stage at diagnosis correlated to lower SePP and Se concentrations. Kaplan-Meier analyses indicated that low Se status at diagnosis (SePP<2.4 mg/l, bottom tertile of patient group) was associated with a poor 5-year survival rate of 20% only. We conclude that SePP and Se concentrations are of prognostic value in RCC and may serve as additional diagnostic biomarkers identifying a Se deficit in kidney cancer patients potentially affecting therapy regimen. As poor Se status was indicative of high mortality odds, we speculate that an adjuvant Se supplementation of Se-deficient RCC patients might be beneficial in order to stabilize their selenoprotein expression hopefully prolonging their survival. However, this assumption needs to be rigorously tested in prospective clinical trials.     

Sequence analysis of genomic DNA (680 Mb) by GS-FLX-Titanium sequencer in the monogonont rotifer, Brachionus ibericus

The monogonont rotifer, Brachionus ibericus (S type), is considered to be a promising model species for developmental biology, evolution, and environmental genomics. In an attempt to accelerate the molecular understanding of B. ibericus, we sequenced 680.5 Mb of genomic DNA using the genome sequencer GS-FLX-Titanium. We obtained 2,062,621 reads (average read length 329.9 bp) and 145,418 contigs (total contigs length 125.7 Mb) after excluding small reads (less than 200 bp) from the assembly, and finally obtained 10,133 unigenes (E value ?? 9.00E?04) after non-redundant (NR) BLAST search. In this article, we summarize the genomic DNA sequences of B. ibericus and discuss their potential use in the study of reproductive biology, endocrinology, environmental genomics, and ecotoxicological studies, and for providing insight into the genetic basis of mechanisms such as egg formation, antioxidant stress defense, and xenobiotic metabolism.     

Down-regulation of aldose reductase renders J774A.1 cells more susceptible to acrolein- or hydrogen peroxide-induced cell death

Aldose reductase (AR) is abundantly expressed in a variety of cell lineages and has been implicated in the cellular response against oxidative stress. However, the exact functional role of AR against oxidative stress remains relatively unclear. This study investigated the role of AR in acrolein- or hydrogen peroxide-induced apoptosis using the J774.A.1 macrophage cell line. Ablation of AR with a small interference RNA or inhibition of AR activity significantly enhanced the acrolein- or hydrogen peroxide-induced generation of reactive oxygen species and aldehydes, leading to increased apoptotic cell death. Blockade of AR activity in J774A.1 cells markedly augmented the acrolein- or hydrogen peroxide-induced translocation of Bax to mitochondria along with reduced Bcl-2 and increased release of cytochrome c from the mitochodria. Taken together, these findings indicate that AR plays an important role in the cellular response against oxidative stress, by sequestering the reactive molecules generated in cells exposed to toxic substances.     

Crystal structure of the serine protease domain of prophenoloxidase activating factor-I

A family of serine proteases (SPs) mediates the proteolytic cascades of embryonic development and immune response in invertebrates. These proteases, called easter-type SPs, consist of clip and chymotrypsin-like SP domains. The SP domain of easter-type proteases differs from those of typical SPs in its primary structure. Herein, we report the first crystal structure of the SP domain of easter-type proteases, presented as that of prophenoloxidase activating factor (PPAF)-I in zymogen form. This structure reveals several important structural features including a bound calcium ion, an additional loop with a unique disulfide linkage, a canyon-like deep active site, and an exposed activation loop. We subsequently show the role of the bound calcium and the proteolytic susceptibility of the activation loop, which occurs in a clip domain-independent manner. Based on biochemical study in the presence of heparin, we suggest that PPAF-III, highly homologous to PPAF-I, contains a surface patch that is responsible for enhancing the catalytic activity through interaction with a nonsubstrate region of a target protein. These results provide insights into an activation mechanism of easter-type proteases in proteolytic cascades, in comparison with the well studied blood coagulation enzymes in mammals.     

Poliovirus entry into human brain microvascular cells requires receptor-induced activation of SHP-2

Viruses use specific receptor molecules to bind selectively to target cells. Receptors have often been considered as mere docking sites, but they may also possess intrinsic signaling capacities that serve to prime the cell for entry and infection. Poliovirus (PV) initiates infection by binding to the PV receptor (PVR) and causes paralytic poliomyelitis by replicating within motor neurons of the brain and spinal cord. We have examined the process by which PV enters cultured human brain microvascular endothelial cells (HBMEC), an in vitro model of the blood-brain barrier. We found that PV enters HBMEC by dynamin-dependent caveolar endocytosis, and that entry depends on intracellular signals triggered by virus attachment to PVR. Tyrosine kinase and RhoA GTPase activation initiated by PVR ligation were both essential. Virus attachment also induced tyrosine phosphorylation of PVR; this permitted the association of PVR with SHP-2, a protein tyrosine phosphatase whose activation was required for entry and infection. The results indicate that receptor-induced signals promote virus entry and suggest a role for tyrosine phosphatases in viral pathogenesis.     

Occurrence and molecular identification of the zoysiagrass mite Aceria zoysiae (Acari: Eriophyidae) in turfgrasses in Korea

Mites are one of the serious pests of turfgrass. Our survey of turfgrass fields from 2013 to 2015 in Korea showed that the occurrence of leaf chlorotic symptom has gradually extended to at least 60% of the examined golf courses. We identified the zoysiae mite Aceria zoysiae in most damaged zoysiagrasses. Artificial infestation of A. zoysiae into zoysiagrasses in pots resulted in symptoms of chlorosis and marginal rolling of the leaves within 3 weeks. We firstly determined the nucleotide sequence of mitochondrial cytochrome c oxidase subunit I (COI) and internal transcribed spacer 2 (ITS2) of the nuclear ribosomal DNA region of A. zoysiae. The variations in COI and ITS2 between A. zoysiae and other species of the genus were 20.9%–43.0% and 7.5%–67.3%, respectively, suggesting significant genetic divergence within the genus. Our study provides valuable information for the rapid diagnosis and infestation monitoring of A. zoysiae in turfgrass fields.