Program Book - 27th Fungal Genetics Conference

FULL POSTER SESSION ABSTRACTSbroad-host range plant pathogen, only dozens of genes involved in pathogenesis have been identified and characterized. In order to establish a highthroughputplatform,this study delivered the double-stranded RNA(dsRNA) expression cassette into protoplasts to trigger silencing for functional genomicsresearch in C. gloeosporioides.A new silencing vector pSD-SUR1 based on RNA-silencing vector (pSD1) with a convergent dual promoter was introduced. Inthis silencing system, the target gene was proposed to be transcribed as a chimeric RNA which activates the system. As an indicator of gene silencing, GFPfluorescence is used to evaluate efficiency of this silencing system. The fluorescence observation showed GFP fluorescence significantly decreased in someof the silenced strains, comparing with the recipient strain. The GFP mRNA transcript levels in the strains were analyzed using quantitative RT-PCR. Theresults showed that the reduction range of controls in gfp expression was from 30% to 80%,suggesting an effective gene silencing system and a feasibleapproach to generate detectable phenotypes in C. gloeosporioides. In addition, some genes encoding signal transduction pathways and transcriptionalfactor were inserted respectively into the vector pSD-SUR1 and to be silenced. In conclusion, RNA silencing system opens up new opportunity for exploringgene function in the fungus C. gloeosporioides.347. Comparative Genomics of L and S Morphotypes of Aspergillus flavus. Mana Ohkura, Peter Cotty, Marc Orbach. Division of Plant Pathology,University of AZ, Tucson, AZ.Aspergillus flavus is a widely distributed facultative pathogen of plants and animals and the most common causal agent of crop contamination withaflatoxins. Isolates of A. flavus vary widely in aflatoxin producing ability, ranging from atoxigenic to being capable of producing many mg/g. Variability inaflatoxin production makes specific attribution of etiology very complex. Aspergillus flavus exists in two morphotypes the large (L) and small (S) sclerotialproducing strains. The S strains have consistent high aflatoxin-producing ability while the L strains vary greatly in toxin production with atoxigenic strainscommonly found. Some atoxigenic strains are active ingredients in biocontrol products used commercially to prevent contamination. We are applyingcomparative genomics to L and S strains in an attempt to reveal clues to potential differential adaptations associated with the variation in aflatoxinproductionbetween these morphotypes. In addition to aflatoxin producing potential, several characteristics diverge between the morphotypes includingsclerotial morphology as well as spore and hydrolase production and prevalence. We hypothesize there are genomic differences between the L and Smorphotypes that reflect their divergent evolution leading to differential adaptation. To evaluate this, we have sequenced the genomes of three Lmorphotype and three S morphotype isolates from agricultural fields in Arizona belonging to 6 different vegetative compatibility groups. L and S strainisolates from across Arizona were selected. Strains were sequenced to ~40-45 X coverage on the Illumina HiSeq 2000 platform. The genomes wereassembled de novo using VelvetOptimiser and gaps were filled using GapFiller. Preliminary assessment of the assemblies indicate there is ~90% synteniccoverage with the published genome of A. oryzae RIB40, a close relative of A. flavus. The genomes were annotated transitively with RATT using thegenome of A. oryzae RIB40 as a reference. Comparisons of genome statistics, secondary metabolite clusters, and morphotype specific genes will bepresented.348. Searching for Functional Mobile Elements in Coprinopsis cinerea. Kendra Boyd, Madhura Chitnavis, Marilee A. Ramesh. Dept Biol, Roanoke College,Salem, VA.The genome of Coprinopsis cinerea contains both Class I and Class II repetitive elements, making up about 2.5% of the total genomic DNA. Whilebioinformatics techniques were used to identify and classify these elements based on sequence similarity, it is uncertain whether any of the elements arefunctional. Although the nature of repetitive elements is to expand their numbers within the genome, the genome acts to suppress activity throughmutation and methylation. Detailed analysis to survey functionality and expression was conducted on two families of repetitive elements, the Gypsy-likeretrotransposons (Class I) and the hAT Transposons (Class II). The potential for functionality was determined based on size, structure and flanking repeatsequence integrity. Evidence for expression was determined based on reviewing EST, SAGE and methylation data for these elements. Of the 31 largestgypsy elements analyzed, all appear to be inactive. However, one of the nine hAT elements appears to be structurally intact and shows evidence ofexpression. The dimerization domain of this element is being studied as a potential region to assay for activity.349. Genome-wide analysis of small RNA machineries in fungal kingdom. Jiayao Wu 1 , Jaeyoung Choi 2 , Fred O. Asiegbu 1 , Jari P.T. Valkonen 1 , Yong-HwanLee 1,2 . 1) Forest Science, Helsinki University, Helsinki, Finland; 2) Department of Agricultural Biotechnology, Seoul National University, Seoul, Korea.RNA interference (RNAi) is a phenomenon widely conserved in eukaryotes to regulate gene expression through diverse pathways at transcriptional (TGS)or post-transcriptional level (PTGS). In fungi, the RNAi pathways are found with three major functions: genomeic defence, heterochromatin formation, andgene regulation. The mechanisms of RNAi in fungi seem to be unique and highly differentiated from plant and animal kingdoms, although the coremechanisms are relatively similar. We identified 3 key genes such as Argonaute, Dicer and RNA-dependent RNA Polymerase (RdRP) in the pathway from143 fungal and 66 other genomes. They were found in most genomes with very different gene numbers, while some of fungal genomes appear to be lackof all the components indicating the absent of the whole pathways. In general, fungi have the same domains in Argonauts with plant and animal, butlonger in the length and less in the number. Compared to plant and animal, fungi have more Dicers, but they do not contain PAZ domain, which is essentialfor RNAi in plant and animal. Phylogentic analysis indicates that most fungal Argonuates belong to AGO-like subfamily. However, fungal Dicers could to bedivided into two subfamilies; one is closely related to plant and animal Dicers and the other only exists within fungal kingdom. Further analysis usingcodonW shows RNAi proteins are evolved into different subfamilies under natural selection not due to random mutation. Taken together fungi RNAipathway is likely to be much complex than we expected with multiple functions in diverse regulatory pathways. All information on proteins analyzed isarchived in Fungal Small RNA Machinery Database (http://funrna.riceblast.snu.ac.kr/).350. Comparative Analysis of Malassezia Mating Loci. Jun Xu 1 , Wenjun Li 2 , Anastasia Giotia 3 , Björn Nystedt 4 , Anna Averettec 2 , Charles Saunders 1 , ThomasDawson 1 , Joseph Heitman 2 , Annika Scheyniuse 5 . 1) Procter & Gamble Co., Mason Business Center, USA; 2) Department of Molecular Genetics andMicrobiology, Duke University Medical Center, USA; 3) Science for Life Laboratory, Translational Immunology Unit, Department of Medicine Solna,Karolinska Institutet, Stockholm, Sweden; 4) Science for Life Laboratory, Department of Biochemistry and Biophysics, Stockholm University, Stockholm,Sweden; 5) Translational Immunology Unit, Department of Medicine Solna, Karolinska Institutet, Stockholm, Sweden.Malassezia fungi are naturally found on the skin surfaces of many animals and are associated with skin disorders such as dandruff and atopicdermatitis/eczema. Whole genome sequence analysis showed that M. globosa has a bipolar mating type with two MAT alleles encoding the homeodomain(HD) and pheromone/receptor (P/R) loci separated by 167 kb of intervening sequence. We compared the M. globosa mating locus with the newlysequenced M. sympodialis genome. Our analysis showed that the M. sympodialis MAT region has extensive well-conserved synteny with M. globosa andthe HD and P/R mating subloci are physically linked with a 141 kb separating the two. Interestingly, MAT sequences derived from a population of M.sympodialis isolates suggests that M. sympodialis does not fit a traditional bipolar or tetrapolar system. Instead, it is more similar to a pseudo-bipolarmodel previously reported for Sporidiobolus salmonicolor in which the HD and P/R genes are physically linked similar to bipolar mating type configurations.206