Program Book - 27th Fungal Genetics Conference

FULL POSTER SESSION ABSTRACTSlimited to the production of cellulolytic enzymes. The continuing characterization of these mutants will enhance our understanding of the ability of N.crassa to utilize the complex carbon sources present in its natural environment.721. Identification and Functional analysis of New Neurospora crassa Nonself Recognition Loci. Jiuhai Zhao, Charles Hall, Elizabeth Hutchison, DavidKowbel, Juliet Welch, N. Louise Glass. Department of Plant and Microbial Biology, University of California, Berkeley USA 94720.Self/nonself recognition is a ubiquitous and essential function for many organisms. In filamentous fungi, self/nonself recognition is conferred by geneticdifferences at het (heterokaryon incompatibility) loci. The genes that mediate HI (heterokaryon incompatibility) exhibit characteristic evolutionarysignatures, including balancing selection and trans-species polymorphisms. Recent analyses show that genes containing a HET domain are involved in HI,making HET domain genes good candidates for identifying new het loci. In this study, we utilized RNA-seq data from a population of 110 Neurospora crassastrains to look for HET domain genes that were highly polymorphic, have multiple alleles, and show balancing selection, and trans-species polymorphisms.Using this approach, we identified 19 of the 62 HET domain genes in N. crassa that fit the criteria for a het locus. Further, we showed that one of these HETdomain genes, NCU09037, functions as a het locus.722. Combinatorial cationic and oxidative stresses promote the killing of Candida albicans cells by human neutrophils. Alistair J P Brown 1 , DespoinaKaloriti 1 , Mette Jacobsen 1 , Zhikang Yin 1 , Anna Tillmann 1 , Miranda Patterson 2 , Deborah A Smith 2 , Emily Cook 3 , Tao You 4 , Iryna Bohovych 1 , Celso Grebogi 4 ,Neil A R Gow 1 , Janet Quinn 2 , Ken Haynes 3 . 1) School of Medical Sciences, University of Aberdeen, Aberdeen, United Kingdom; 2) Institute for Cell andMolecular Biosciences, Faculty of Medical Sciences, Newcastle University, United Kingdom; 3) School of Biosciences, College of Life & EnvironmentalSciences, University of Exeter, United Kingdom; 4) Institute for Complex Systems and Mathematical Biology, School of Natural and Computing Sciences,University of Aberdeen, United Kingdom.Candida albicans is an opportunistic pathogen of humans. It is thought to have evolved as a relatively harmless commensal. C. albicans is a frequentcause of mucosal and skin infections (thrush). However, our immune system normally blocks potentially lethal systemic infections of the bloodstream andinternal organs. Neutropenic patients are prone to systemic candidiasis because they lack the effective defenses provided by circulating neutrophils. Wehave shown that the efficient killing of C. albicans by human neutrophils is mediated by the potent combinations of stresses they impose on the invadingfungus, rather than specific individual stresses. In particular, exposure to the combination of reactive oxygen species and cation fluxes kills C. albicanssynergistically. We have explored the mechanistic basis for this synergistic killing using genomic exploration and molecular dissection in C. albicanscombined with dynamic mathematical modeling. Signalling via the Hog1 stress activated protein kinase and the Cap1 AP-1-like transcription factor isinhibited by combinatorial oxidative and cationic stresses, and as a result, their downstream gene targets are not induced. This prevents the activation ofnormal oxidative and cationic stress adaptation and repair mechanisms. In particular, hydrogen peroxide detoxification mechanisms are inhibited byelevated salt concentrations. This leads to the accumulation of intracellular reactive oxygen species, and ultimately to accelerated necrotic death. Ectopicexpression of key detoxification mechanisms in C. albicans cells decreases the efficacy of killing by human neutrophils.723. Phosphoproteomic analysis of the aquatic fungus Blastocladiella emersonii during germination. J. Crestani, S. Lopes Gomes. Biochemistry, IQ, USP,Sao Paulo, Brazil.The aquatic fungus Blastocladiella emersonii presents an interesting life cycle with two cell differentiation stages, the germination and the sporulation,during which drastic morphological and biochemical changes are observed. During germination, protein synthesis is inhibited, a transient increase of cAMPlevels is observed, activation of PKA is detected, as well as mobilization of cellular glycogen and an efflux of calcium. These results suggest a possiblephosphorylation/dephosphorylation control, probably through cell signaling networks, which were not characterized up to now. Therefore, the presentwork aims to elucidate these signaling mechanisms by using a phosphoproteomic analysis of B. emersonii during the germination. Firstly, we compared thephosphoproteomic profile from two distinct cell types of B. emersonii, the zoospores and the germling cell (at 45min of germination) by using twodimensionalgels stained with Pro-Q Diamond phosphoprotein dye. The comparison revealed about 82 phosphoproteins from germling cells and 44phosphoproteins from zoospores. These preliminary results suggest that phosphorylation events may be involved during early germination. To detect thesignaling processes and the proteins involved in these events we utilized IMAC-IMAC phosphoproteomic methodology to obtain an enrichedphosphopeptide sample from each stage of germination and early vegetative growth (0, 25, 45, 60 and 90 min). Enriched phosphopeptide samples will bedetected by using a LTQ Velos Orbitrap mass spectrometer; filtered using DTASelect and analyzed using Ascore and Debunker. The results of this work willcontribute to improve the knowledge of the cellular regulatory processes in this early diverging fungus. Financial support: FAPESP and CNPq.724. Towards an accurate genome: high-throughput proteogenomic validation of Stagonospora nodorum genes via sub-cellular proteomics. KejalDodhia 1 , Robert Syme 1 , Thomas Stoll 2 , Marcus Hastie 2 , James Hane 3 , Angela Williams 3 , Eiko Furuki 1 , Jeffrey Gorman 2 , Richard Oliver 1 , Kar-Chun Tan 1 . 1) TheAustralian Centre for Necrotrophic Fungal Pathogens, Environment & Agriculture, Curtin University, Perth, Bentley 6102, Australia; 2) Protein DiscoveryCentre, Queensland Institute of Medical Research, Herston, Qld 4029, Australia; 3) Plant Industry, Commonwealth Scientific and Industrial ResearchOrganisation, Private Bag No, 5, Wembley WA 6913, Australia.Stagonospora nodorum is the causal agent of stagonospora nodorum blotch on wheat. S. nodorum was the first of the Pleosporales fungi to have itsgenome sequence published and genes annotated. However, in silico gene annotation can be erroneous. Therefore, experimental evidence is oftenneeded to refine gene annotations. Proteogenomics is an emerging high-throughput technique, which is a “direct-to-genome mapping” techniquewhereby the mass spectra from protein analyses are mapped onto the predicted gene set and/or the 6-frame whole genome translation. In this study, weperformed a comprehensive proteogenomic analysis of the secreted, intracellular and cell-wall/membrane sub-proteomes of S. nodorum using a twodimensionalliquid chromatography (2D-LC) LTQ Orbitrap MS approach. This study has verified a total of 3580 genes from all sub-proteomes. Of these, 113had not been experimentally verified previously. When combined with previous proteomic data, 4377 (35% of the total predicted gene set) genes wereverified. In addition, all mass spectra were matched to a 6-frame genome translation database to identify evidence of gene model conflicts. The study hasfound that 2629 genes showed evidence of frame conflicts and extensions of coding exons into annotated introns or untranslated regions. At least 43potential new genes were identified.725. Evolutionary Imprint of Fungal PKS-NRPS Catalytic Domains. Daniela Boettger 1 , Holger Bergmann 2 , Barbara Kühn 1 , Ekaterina Shelest 2 , ChristianHertweck 1 . 1) Department Biomolecular Chemistry, Leibniz Institute for Natural Product Research and Infection Biology - Hans Knöll Institute,Beutenbergstrasse 11a, 07743 Jena, Germany; 2) Department of Systems Biology/Bioinformatics, Leibniz Institute for Natural Product Research andInfection Biology - Hans Knöll Institute, Beutenbergstrasse 11a, 07743 Jena, Germany.Fungal polyketide synthase-nonribosomal peptide synthetase (PKS-NRPS) hybrid enzymes produce a broad array of ecologically and medicinally relevant298