TmpL, a Transmembrane Protein Required for Intracellular Redox Homeostasis and Virulence in a Plant and an Animal Fungal Pathogen

The regulation of intracellular levels of reactive oxygen species (ROS) is critical for developmental differentiation and virulence of many pathogenic fungi. In this report we demonstrate that a novel transmembrane protein, TmpL, is necessary for regulation of intracellular ROS levels and tolerance to external ROS, and is required for infection of plants by the necrotroph Alternaria brassicicola and for infection of mammals by the human pathogen Aspergillus fumigatus. In both fungi, tmpL encodes a predicted hybrid membrane protein containing an AMP-binding domain, six putative transmembrane domains, and an experimentally-validated FAD/NAD(P)-binding domain. Localization and gene expression analyses in A. brassicicola indicated that TmpL is associated with the Woronin body, a specialized peroxisome, and strongly expressed during conidiation and initial invasive growth in planta. A. brassicicola and A. fumigatus ΔtmpL strains exhibited abnormal conidiogenesis, accelerated aging, enhanced oxidative burst during conidiation, and hypersensitivity to oxidative stress when compared to wild-type or reconstituted strains. Moreover, A. brassicicola ΔtmpL strains, although capable of initial penetration, exhibited dramatically reduced invasive growth on Brassicas and Arabidopsis. Similarly, an A. fumigatus ΔtmpL mutant was dramatically less virulent than the wild-type and reconstituted strains in a murine model of invasive aspergillosis. Constitutive expression of the A. brassicicola yap1 ortholog in an A. brassicicola ΔtmpL strain resulted in high expression levels of genes associated with oxidative stress tolerance. Overexpression of yap1 in the ΔtmpL background complemented the majority of observed developmental phenotypic changes and partially restored virulence on plants. Yap1-GFP fusion strains utilizing the native yap1 promoter exhibited constitutive nuclear localization in the A. brassicicola ΔtmpL background. Collectively, we have discovered a novel protein involved in the virulence of both plant and animal fungal pathogens. Our results strongly suggest that dysregulation of oxidative stress homeostasis in the absence of TmpL is the underpinning cause of the developmental and virulence defects observed in these studies

HLAscan: genotyping of the HLA region using next-generation sequencing data

Background Several recent studies showed that next-generation sequencing (NGS)-based human leukocyte antigen (HLA) typing is a feasible and promising technique for variant calling of highly polymorphic regions. To date, however, no method with sufficient read depth has completely solved the allele phasing issue. In this study, we developed a new method (HLAscan) for HLA genotyping using NGS data. Results HLAscan performs alignment of reads to HLA sequences from the international ImMunoGeneTics project/human leukocyte antigen (IMGT/HLA) database. The distribution of aligned reads was used to calculate a score function to determine correctly phased alleles by progressively removing false-positive alleles. Comparative HLA typing tests using public datasets from the 1000 Genomes Project and the International HapMap Project demonstrated that HLAscan could perform HLA typing more accurately than previously reported NGS-based methods such as HLAreporter and PHLAT. In addition, the results of HLA-A, −B, and -DRB1 typing by HLAscan using data generated by NextGen were identical to those obtained using a Sanger sequencing–based method. We also applied HLAscan to a family dataset with various coverage depths generated on the Illumina HiSeq X-TEN platform. HLAscan identified allele types of HLA-A, −B, −C, −DQB1, and -DRB1 with 100% accuracy for sequences at ≥ 90× depth, and the overall accuracy was 96.9%. Conclusions HLAscan, an alignment-based program that takes read distribution into account to determine true allele types, outperformed previously developed HLA typing tools. Therefore, HLAscan can be reliably applied for determination of HLA type across the whole-genome, exome, and target sequences

A Pectate Lyase-Coding Gene Abundantly Expressed during Early Stages of Infection Is Required for Full Virulence in Alternaria brassicicola

We thank Fred Brooks for insightful discussions on the roles of PL1332 on pathogenesis mechanisms employed by A. brassicicola.Alternaria brassicicola causes black spot disease of Brassica species. The functional importance of pectin digestion enzymes and unidentified phytotoxins in fungal pathogenesis has been suspected but not verified in A. brassicicola. The fungal transcription factor AbPf2 is essential for pathogenicity and induces 106 genes during early pathogenesis, including the pectate lyase-coding gene, PL1332. The aim of this study was to test the importance and roles of PL1332 in pathogenesis. We generated deletion strains of the PL1332 gene, produced heterologous PL1332 proteins, and evaluated their association with virulence. Deletion strains of the PL1332 gene were approximately 30% less virulent than wild-type A. brassicicola, without showing differences in colony expansion on solid media and mycelial growth in nutrient-rich liquid media or minimal media with pectins as a major carbon source. Heterologous PL1332 expressed as fusion proteins digested polygalacturons in vitro. When the fusion proteins were injected into the apoplast between leaf veins of host plants the tissues turned dark brown and soft, resembling necrotic leaf tissue. The PL1332 gene was the first example identified as a general toxin-coding gene and virulence factor among the 106 genes regulated by the transcription factor, AbPf2. It was also the first gene to have its functions investigated among the 19 pectate lyase genes and several hundred putative cell-wall degrading enzymes in A. brassicicola. These results further support the importance of the AbPf2 gene as a key pathogenesis regulator and possible target for agrochemical development.Yeshttp://www.plosone.org/static/editorial#pee

Transcriptional Responses of the Bdtf1-Deletion Mutant to the Phytoalexin Brassinin in the Necrotrophic Fungus Alternaria brassicicola

Brassica species produce the antifungal indolyl compounds brassinin and its derivatives, during microbial infection. The fungal pathogen Alternaria brassicicola detoxifies brassinin and possibly its derivatives. This ability is an important property for the successful infection of brassicaceous plants. Previously, we identified a transcription factor, Bdtf1, essential for the detoxification of brassinin and full virulence. To discover genes that encode putative brassinin-digesting enzymes, we compared gene expression profiles between a mutant strain of the transcription factor and wild-type A. brassicicola under two different experimental conditions. A total of 170 and 388 genes were expressed at higher levels in the mutants than the wild type during the infection of host plants and saprophytic growth in the presence of brassinin, respectively. In contrast, 93 and 560 genes were expressed, respectively, at lower levels in the mutant than the wild type under the two conditions. Fifteen of these genes were expressed at lower levels in the mutant than in the wild type under both conditions. These genes were assumed to be important for the detoxification of brassinin and included Bdtf1 and 10 putative enzymes. This list of genes provides a resource for the discovery of enzyme-coding genes important in the chemical modification of brassinin

The Effect of Elastic Tape on Lower Extremity Muscle Activity in Squats of Young Female Adults: A Cross-sectional Pilot Study

Background: In terms of physical performance, elastic tape (ET) is known to contribute to injury prevention and performance enhancement. Objects: This study aimed to compare and analyze the effect on lower extremity muscle activity of young adult women with and without ET during squats. Methods: In this study, six healthy, young women were recruited as participants in a university laboratory. Participants were allocated to two groups of three after measuring muscle activity in a pre-test, and the experiment was conducted for a total of two weeks (two sessions). First, 10 half squats were taped once in the first week, and 10 half squats were performed without taping in the second week. The other group did this in reverse and measured muscle activity after the squat was over. Results: As a result of this study, there was no significant difference in the quadriceps with or without ET (Z = –0.11, p > 0.05). Similarly, no significant difference was found in hamstring (Z = –0.31, p > 0.05). Conclusion: No beneficial effect was found on changes in muscle activity following ET application during squats. Further studies require randomized controlled trials that increase the number of participants and the intensity of the intervention, and measure pain, function, and performance rather than muscle properties depending on the biomechanical lifting mechanism

A systematic approach for diagnosing service failure: Service-specific FMEA and grey relational analysis approach

In any organization, the importance of failure management cannot be mentioned by a single word. However, most failure analysis is dominated by the manufacturing sector, despite the increasing importance of the service sector. In response, this paper proposes a systematic approach for identifying and evaluating potential failures using a service-specific failure mode and effect analysis (service-specific FMEA) and grey relational analysis. The proposed approach consists of two stages: construction of service-specific FMEA and application of grey relational analysis. The first stage, construction of service-specific FMEA, aims at incorporating the service specific characteristics to the traditional FMEA, providing 3 dimensions and 19 sub-dimensions, encompassing the service characteristics. At the second stage, grey relational analysis is applied to calculate the risk priority of each failure mode to deal with the necessities of a flexible evaluation framework under these interrelated multi-dimensions. The proposed approach is expected to help the service managers to manage the service failure within the systematic framework. This paper contributes to the field in that it incorporates the service-specific characteristics to the traditional FMEA, as well as providing the appropriate evaluation framework using grey relational analysis. (C) 2011 Elsevier Ltd. All rights reserved.GEUM Y, 2010, SERV IND J 0928, DOI DOI 10.1080/02642069.2010.503876NARAYANAGOUNDER S, 2009, P WORLD AC SCI ENG TKuo Y, 2008, COMPUT IND ENG, V55, P80, DOI 10.1016/j.cie.2007.12.002Li GD, 2007, MATH COMPUT MODEL, V46, P573, DOI 10.1016/j.mcm.2006.11.021CHEN JK, 2007, J FAILURE ANAL PREVE, V7, P321Chuang PT, 2007, SERV IND J, V27, P91, DOI 10.1080/02642060601122587DONG C, 2007, INT J QUAL RELIAB MA, V24, P958TAY KM, 2006, INT J QUALITY RELIAB, V23, P1047TENG SG, 2006, INT J QUALITY RELIAB, V23, P179, DOI 10.1108/02656710610640943YANG CC, 2006, J MANUFACTURING TECH, V17, P926Chen MF, 2004, MATH COMPUT MODEL, V40, P1473, DOI 10.1016/j.mcm.2005.01.006Fliess S, 2004, J BUS RES, V57, P392, DOI 10.1016/S0148-2963(02)00273-4LEWIS BR, 2004, INT J CONT HOSPITALI, V16, P6, DOI 10.1108/09596110410516516SYSON F, 2004, J SERV MARK, V18, P255Rhee SJ, 2003, ADV ENG INFORM, V17, P179, DOI 10.1016/j.aei.2004.07.002Hess RL, 2003, J ACAD MARKET SCI, V31, P127, DOI 10.1177/0092070302250898MUELLER RD, 2003, HOSP MANAGE, V22, P395Pillay A, 2003, RELIAB ENG SYST SAFE, V79, P69STAMATIS DH, 2003, FAILURE MODE EFFECTGoldstein SM, 2002, J OPER MANAG, V20, P121, DOI 10.1016/S0272-6963(01)00090-0PUENTE J, 2002, INT J QUALITY RELIAB, V19, P137WU HH, 2002, QUAL ENG, V15, P209CHANG CL, 2001, INTEGRATED MANUFACTU, V12, P211Colgate M, 2001, INT J SERV IND MANAG, V12, P215LEWIS BR, 2001, INT J BANK MARKETING, V19, P37MATTILA AS, 2001, J SERV MARK, V15, P583Michel S, 2001, INT J SERV IND MANAG, V12, P20Miller JL, 2000, J OPER MANAG, V18, P387Stewart DM, 1999, PROD OPER MANAG, V8, P240CHANG CL, 1999, KYBERNETES, V28, P1072ROOS I, 1999, J SERV RES-US, V2, P68SMITH A, 1999, J MARKETING RES, V34, P356Vandenbrande WW, 1998, QUAL PROG, V31, P97Tax SS, 1998, SLOAN MANAGE REV, V40, P75Boshoff C, 1998, INT J SERV IND MANAG, V9, P24*DAIML CHRYSL CORP, 1998, QSA QUAL SYST ASSJOHNE A, 1998, EUR J MARKETING, V32, P184MCDOUGALL G, 1998, J HOSPITALITY LEISUR, V5, P27VANLOOY B, 1998, SERVICES MANAGEMENTBoshoff C, 1997, INT J SERV IND MANAG, V8, P110HOFFMAN KD, 1997, ESSENTIALS SERVICE MEdvardsson B, 1996, SERV IND J, V16, P140Reichheld FF, 1996, HARVARD BUS REV, V74, P56BENDAYA M, 1996, INT J QUALITY RELIAB, V13, P43ARMISTEAD CG, 1995, MANAGING SERVICE RECBOWLES JB, 1995, RELIAB ENG SYST SAFE, V50, P203HOFFMAN KD, 1995, J SERV MARK, V9, P49KELLOGG DL, 1995, J OPERATIONS MANAGEM, V13, P323TAYLOR S, 1994, J MARKETING, V58, P56BAILEY D, 1994, MANAG SERV QUAL, V4, P25FITZSIMMONS JA, 1994, SERVICE MANAGEMENT CKELLEY SW, 1994, J ACADEMY MARKETING, V22, P52KELLEY SW, 1993, J RETAILING, V69, P429BOULDING W, 1993, J MARKETING RES, V30, P7ZEITHAML VA, 1993, J ACADEMY MARKETING, V21, P1BERRY LL, 1992, ORGAN DYN, V20, P5BITNER MJ, 1990, J MARKETING, V54, P71DALE BG, 1990, QUALITY RELIABILITY, V6, P179GRONROOS C, 1990, SERVICE MANAGEMENT MDENG JL, 1989, J GREY SYSTEM, V1, P1JONES P, 1989, MANAGEMENT SERVICE IBERRY L, 1991, MARKETING SERVICESBELL CR, 1987, MANAGE REV, V76, P32SHOSTACK GL, 1987, J MARKETING, V51, P34SURPRENANT CF, 1987, J MARKETING, V51, P73AGBONIFOH BA, 1986, EUR J MARKETING, V20, P43PARASURAMAN A, 1985, J MARKETING, V49, P41SHOSTACK GL, 1985, SERVICE ENCOUNTER, P243SHOSTACK GL, 1984, HARVARD BUS REV, V62, P133CHASE RB, 1981, OPER RES, V29, P698

Dynamic Evolution Of Plant Mitochondrial Genomes: Mobile Genes And Introns And Highly Variable Mutation Rates

We summarize our recent studies showing that angiosperm mitochondrial (mt) genomes have experienced remarkably high rates of gene loss and concomitant transfer to the nucleus and of intron acquisition by horizontal transfer. Moreover, we find substantial lineage-specific variation in rates of these structural mutations and also point mutations. These findings mostly arise from a Southern blot survey of gene and intron distribution in 281 diverse angiosperms. These blots reveal numerous losses of mt ribosomal protein genes but, with one exception, only rare loss of respiratory genes. Some lineages of angiosperms have kept all of their mt ribosomal protein genes whereas others have lost most of them. These many losses appear to reflect remarkably high (and variable) rates of functional transfer of mt ribosomal protein genes to the nucleus in angiosperms. The recent transfer of cox2 to the nucleus in legumes provides both an example of interorganellar gene transfer in action and a starting point for discussion of the roles of mechanistic and selective forces in determining the distribution of genetic labor between organellar and nuclear genomes. Plant mt genomes also acquire sequences by horizontal transfer. A striking example of this is a homing group I intron in the mt cox1 gene. This extraordinarily invasive mobile element has probably been acquired over 1,000 times separately during angiosperm evolution via a recent wave of cross-species horizontal transfers. Finally, whereas all previously examined angiosperm mtDNAs have low rates of synonymous substitutions, mtDNAs of two distantly related angiosperms have highly accelerated substitution rates

Fungal-specific transcription factor AbPf2 activates pathogenicity in Alternaria brassicicola

Alternaria brassicicola is a successful saprophyte and necrotrophic plant pathogen. To identify molecular determinants of pathogenicity, we created non-pathogenic mutants of a transcription factor-encoding gene, AbPf2. The frequency and timing of germination and appressorium formation on host plants were similar between the non-pathogenic abpf2 mutants and wild-type A. brassicicola. The mutants were also similar in vitro to wild-type A. brassicicola in terms of vegetative growth, conidium production, and responses to a phytoalexin, reactive oxygen species and osmolites. The hyphae of the mutants grew slowly but did not cause disease symptoms on the surface of host plants. Transcripts of the AbPf2 gene increased exponentially soon after wild-type conidia contacted their host plants . A small amount of AbPf2 protein, as monitored using GFP fusions, was present in young, mature conidia. The protein level decreased during saprophytic growth, but increased and was located primarily in fungal nuclei during pathogenesis. Levels of the proteins and transcripts sharply decreased following colonization of host tissues beyond the initial infection site. When expression of the transcription factor was induced in the wild-type during early pathogenesis, 106 fungal genes were also induced in the wild-type but not in the abpf2 mutants. Notably, 33 of the 106 genes encoded secreted proteins, including eight putative effector proteins. Plants inoculated with abpf2 mutants expressed higher levels of genes associated with photosynthesis, the pentose phosphate pathway and primary metabolism, but lower levels of defense-related genes. Our results suggest that AbPf2 is an important regulator of pathogenesis, but does not affect other cellular processes in A. brassicicola