Improved heterologous protein production by a tripeptidyl peptidase gene (<i>AosedD</i>) disruptant of the filamentous fungus <i>Aspergillus oryzae</i>

Proteolytic degradation is one of the serious bottlenecks limiting the yields of heterologous protein production by Aspergillus oryzae. In this study, we selected a tripeptidyl peptidase gene AosedD (AO090166000084) as a candidate potentially degrading the heterologous protein, and performed localization analysis of the fusion protein AoSedD-EGFP in A. oryzae. As a result, the AoSedD-EGFP was observed in the septa and cell walls as well as in the culture medium, suggesting that AoSedD is a secretory enzyme. An AosedD disruptant was constructed to investigate an effect of AoSedD on the production level of heterologous proteins and protease activity. Both of the total protease and tripeptidyl peptidase activities in the culture medium of the AosedD disruptant were decreased as compared to those of the control strain. The maximum yields of recombinant bovine chymosin (CHY) and human lysozyme (HLY) produced by the AosedD disruptants showed approximately 2.9- and 1.7-fold increases, respectively, as compared to their control strains. These results suggest that AoSedD is one of the major proteases involved in the proteolytic degradation of recombinant proteins in A. oryzae.     

Automatic Context-Specific Subnetwork Discovery from Large Interaction Networks

Genes act in concert via specific networks to drive various biological processes, including progression of diseases such as cancer. Under different phenotypes, different subsets of the gene members of a network participate in a biological process. Single gene analyses are less effective in identifying such core gene members (subnetworks) within a gene set/network, as compared to gene set/network-based analyses. Hence, it is useful to identify a discriminative classifier by focusing on the subnetworks that correspond to different phenotypes. Here we present a novel algorithm to automatically discover the important subnetworks of closely interacting molecules to differentiate between two phenotypes (context) using gene expression profiles. We name it COSSY (COntext-Specific Subnetwork discoverY). It is a non-greedy algorithm and thus unlikely to have local optima problems. COSSY works for any interaction network regardless of the network topology. One added benefit of COSSY is that it can also be used as a highly accurate classification platform which can produce a set of interpretable features.     

ARCS: an aggregated related column scoring scheme for aligned sequences

MOTIVATION: Biologists frequently align multiple biological sequences to determine consensus sequences and/or search for predominant residues and conserved regions. Particularly, determining conserved regions in an alignment is one of the most important activities. Since protein sequences are often several-hundred residues or longer, it is difficult to distinguish biologically important conserved regions (motifs or domains) from others. The widely used tools, Logos, Al2co, Confind, and the entropy-based method, often fail to highlight such regions. Thus a computational tool that can highlight biologically important regions accurately will be highly desired. RESULTS: This paper presents a new scoring scheme ARCS (Aggregated Related Column Score) for aligned biological sequences. ARCS method considers not only the traditional character similarity measure but also column correlation. In an extensive experimental evaluation using 533 PROSITE patterns, ARCS is able to highlight the motif regions with up to 77.7% accuracy corresponding to the top three peaks. AVAILABILITY: The source code is available on http://bio.informatics.indiana.edu/projects/arcs and http://goldengate.case.edu/projects/arcs     

Suppressing IL-32 in monocytes impairs the induction of the proinflammatory cytokines TNFα and IL-1β

Targeting major proinflammatory cytokines such as IL-1β and TNFα is of great interest in patients with chronic inflammatory diseases, including rheumatoid arthritis, colitis, and psoriasis. The cytokine Interleukin (IL)-32 induces proinflammatory cytokines such as TNFα, IL-1β, IL-6, and chemokines. We previously used an IL-32 ligand-affinity column to purify proteinase 3, which is abundantly expressed in neutrophil and monocytic leukocytes but not in other cell types, and found that IL-32 is mainly produced by monocytic leukocytes. This evidence suggested that silencing endogenous IL-32 by short hairpin RNA (shRNA) in monocytic cells might reveal the precise function of endogenous IL-32. Indeed, lipopolysaccharide (LPS)- or phorbol myristate acetate (PMA)-induced proinflammatory cytokine production was significantly inhibited in shRNA/IL-32 stable clones as compared to control clones. Furthermore, macrophages in PMA-differentiated shRNA/IL-32 stable clones displayed remarkably impaired LPS- and IL-1β-induced proinflammatory cytokine production. These data suggest that IL-32 is not only involved in host defense against pathogens, but also might play a role in chronic inflammatory diseases. IL-32 production leads to major proinflammatory cytokine production during the initial immune response.     

Amino acid differences in interferon-tau (IFN-τ) of Bos taurus Coreanae and Holstein

Interferons (IFNs) are commonly grouped into type I and type II IFN. Type I IFNs are known as antiviral IFNs including IFN-α, IFN-β, and IFN-ω whereas type II IFN is referred to immune IFN and IFN-γ is only member of the type II IFN. Type I IFNs are induced by virus invading however type II IFN is produced by mitogenic or antigenic stimuli. IFN-τ was first identified in ruminant ungulates as a pregnancy recognition hormone, trophoblastin. IFN-τ constitutes a new class of type I IFN, which possesses the common features of type I IFN, such as the ability to prevent viral infection and to limit cell proliferation. In addition, IFN-τ is unique in that it is induced by pregnancy unlike other type I IFNs. We cloned Bos taurus (B. T.) Coreanae IFN-τ from peripheral blood mononuclear cells. The amino acid sequence of B. T. Coreanae IFN-τ shares only 90.3% identity with that of Holstein dairy cow. Recombinant B. T. Coreanae and Holstein IFN-τ proteins were expressed in Escherichia coli and the antiviral activity of IFN-τ proteins were examined. Both recombinant proteins were active and protected human WISH and bovine MDBK cells from the cytopathic effect of vesicular stomatitis virus. The recombinant IFN-τ protein of B. T. Coreanae and Holstein properly induced the expression of antiviral genes including 2',5'-oligoadenylate synthetase (OAS) and Mx GTPase 1 (Mx-1).     

Spongiibacter pelagi sp. nov., a marine gammaproteobacterium isolated from coastal seawater

A novel gammaproteobacterium, designated as KMU-158^T, was isolated from seawater collected on the coastline of Dadaepo in the Republic of Korea, and characterized using a polyphasic taxonomic approach. Strain KMU-158^T was Gram-staining-negative, pale beige-colored, strictly aerobic, rod-shaped, motile, and chemoorganoheterotrophic. The novel isolate was able to grow at 0–3% NaCl concentrations (w/v), pH 6.5–9.5, and 15–40 °C. The analysis based on 16S rRNA gene sequences indicated that strain KMU-158^T belongs to the family Spongiibacteraceae and shared the highest similarity with Spongiibacter tropicus CL-CB221^T (96.1%). The major cellular fatty acids were summed feature 3 (C_16:1 ω7c and/or C_16:1 ω6c) and C_17:1 ω8c. The only respiratory quinone was identified as ubiquinone-8. The polar lipids of strain KMU-158^T contained phosphatidylethanolamine, phosphatidylglycerol, diphosphatidylglycerol, and two unidentified lipids. The assembled draft genome size of strain KMU-158^T was 3.29 Mbp with a DNA G?+?C content of 51.3%. The average nucleotide identity, digital DNA–DNA hybridization, and average amino acid identity values of KMU-158^T and the representatives of the genus Spongiibacter were found to be 78.5–79.1%, 13.8–14.1%, and 66.6–66.8%, respectively. From the distinct phenotypic, phylogenetic, genomic, and chemotaxonomic properties, the strain KMU-158^T is considered to represent a novel species of the genus Spongiibacter, for which the name Spongiibacter pelagi sp. nov. is proposed. The type strain of the species S. pelagi sp. nov. is KMU-158^T (=?KCCM 90448^T?=?NBRC 114307^T).