Program Book - 27th Fungal Genetics Conference

FULL POSTER SESSION ABSTRACTSThe sirtuins are members of the NAD + -dependent histone deacetylase family that contribute to various cellular functions which are affected aging,disease and cancer development. However, physiological roles of the fungal-specific sirtuin family are still poorly understood, especially with regard totheir participation in the genomic stability of yeast. Here, we determined the novel function of the fungal-specific sirtuin HstD, which is homolog of yeastHst4 in Aspergillus oryzae. The deletion of HstD indicated that both conidial development and secondary metabolism were regulated by HstD in A. oryzae.Furthermore, the gene expression of LaeA, which is the most studied coordinator for the regulation of secondary metabolism and development, wasinduced in the DHstD strain, and we found a significant genetic interaction between HstD and LaeA using double-disrupted or overexpression strains. Thus,we concluded the fungal-specific sirtuin HstD coordinates the fungal development and secondary metabolism via the regulation of LaeA gene expression infilamentous fungi. The HstD is fungal-specific, but it is conserved in the vast family of filamentous fungi. Therefore, HstD has great potential as a drugtarget for mycosis or plant disease, because the fungal development and secondary metabolism are virulence determinants of pathogenic fungi. Inaddition, our findings are also important for improving the productivity of useful secondary metabolites and developing an attractive host for theproduction of several heterogeneous secondary metabolites.451. Improved flavor production by manipulation of the Ehrlich pathway in ascomycetes. D. Ravasio, A. Walther, J. Wendland. Carlsberg Laboratory,Copenhagen V, Denmark.The Ehrlich pathway utilizes amino acids to generate higher alcohols with distinctive flavor in three enzymatic steps including a transaminase, adecarboxylase and an aldehyde dehydrogenase. Comparative genomics revealed the absence of key genes of the Ehrlich pathway in Eremotheciumcymbalariae whereas these genes were found to be present in the closely related species Ashbya gossypii. A. gossypii produces a very fruity flavor both inliquid culture and on solid media. The biological significance of this is unknown. Here, we present the functional analysis of A. gossypii key genes of theEhrlich pathway, ARO8a, ARO8b, ARO10, and ARO80. Deletion of any one component resulted in a noticeable reduction of flavor production asdetermined by GC/MS. In Saccharomyces cerevisiae ARO80 has been described as the main transcription factor regulating other genes of the Ehrlichpathway. Therefore, we analyzed the effect of deletion and overexpression of this gene on flavor production in yeast. As expected, overexpressionresulted in a marked increase in flavor production, particularly in isoamyl alcohol, a banana-like flavor. Next to chemical analyses we generated a lacZbasedreporter gene assay using ARO-gene promoters. With such a tool we can determine the status of flavor production under various conditions and in avariety of yeast strains. Initial results will be presented.452. Suppressor mutagenesis of a DlaeA mutant reveals novel regulators of secondary metabolism in Aspergillus nidulans. Alexandra Soukup, Jerry Luo,Jin Woo Bok, Nancy P. Keller. UW-Madison, Madison, WI.Aspergillus nidulans is a filamentous fungus known to produce a variety of complex natural products known as secondary metabolites (SM). Regulation ofthese bioactive SM can occur through cluster specific transcription factors, or through global regulators such as LaeA. Deletion of laeA results in drasticallydecreased amounts of multiple secondary metabolites. A multi-copy suppressor screen for genes capable of phenotypically returning norsolorinic acid(NOR) production to the DlaeA mutant resulted in identification of 17 plasmids containing inserts ranging from one to four genes. Further analysis of thesuppressor plasmids confirmed of a subset to increase SM production both in the original laeA deletion strain and in wild type backgrounds.453. A network of HMG-box transcription factors regulates sexual cycle in the fungus Podospora anserina. J. Aït-Benkhali 1,2 , E. Coppin 1,2 , S. Brun 1,2,3 , T.Martin 4 , C. Dixelius 4 , R. Debuchy 1,2 . 1) Univ Paris-Sud, Institut de Génétique et Microbiologie, Orsay, France; 2) CNRS, Institut de Génétique etMicrobiologie, Orsay, France; 3) UFR des Sciences du Vivant, Université Paris-7 Diderot, Paris, France; 4) Department of Plant Biology and Forest Genetics,Uppsala BioCenter, Swedish University of Agricultural Sciences and Linnean Center for Plant Biology, Uppsala, Sweden.High-mobility group B proteins are eukaryotic DNA-binding proteins characterized by the HMG-box functional motif. These transcription factors play apivotal role in global genomic functions and in the control of genes involved in specific developmental or metabolic pathways. The filamentous ascomycetePodospora anserina contains 12 HMG-box genes. Of these, four have been previously characterized; three are mating-type genes that control fertilizationand development of the fruiting-body, whereas the last one encodes a factor involved in mitochondrial DNA stability. Systematic deletion analysis of theeight remaining uncharacterized HMG-box genes indicated that none were essential for viability, but that seven were involved in the sexual cycle. TwoHMG-box transcription factors display striking features. Pa_1_13940, an ortholog of SpSte11 from Schizosaccharomyces pombe, is a pivotal activator ofmating-type genes in P. anserina, whereas Pa_7_7190 is a repressor of several phenomena specific to the stationary phase, most notably hyphalanastomoses. Constitutive expression of mating-type genes in a DPa_1_13940 strain did not restore fertility, indicating that Pa_1_13940 has additionalfunctions related to sexual reproduction besides activating mating-type genes. RT-qPCR analyses of HMG-box genes in different HMG-box deletion strainsindicated that Pa_1_13940 is at the hub of a network of several HMG-box factors that regulate the sexual cycle. Complementation experiments with astrain deleted for mating-type genes revealed that this network control fertility genes in addition to mating-type target genes. This study points to thecritical role of the HMG-box members in sexual reproduction in fungi, as 11 out of 12 members were involved in the sexual cycle in P. anserina.Pa_1_13940 and SpSte11 are conserved transcriptional regulators of mating-type genes, although P. anserina and S. pombe have diverged 1.1 billion yearsago. Two HMG-box genes, SOX9 and its upstream regulator SRY, play also an important role in sex determination in mammals. The mating-type genes andtheir upstream regulatory factor form a module of HMG-box genes similar to the SRY/SOX9 module, suggesting it may be ancestral in Opisthokonta.454. Sclerotinia sclerotiorum MAT genes function in fertility and apothecial morphogenesis. Benjamin Doughan, Jeffrey Rollins. Plant Pathology,University of Florida, Gainesville, FL.Sclerotinia sclerotiorum (Lib.) de Bary is an omnivorous, polyphagus, phytopathogenic fungus that relies on the completion of the sexual cycle to initiatemost new disease cycles. The sexual cycle is characterized by the development of apothecia that forcibly discharge ascospores for local and, under suitableconditions, long distance dissemination. A strategy for understanding the regulation of apothecial multicellular development is being pursued throughfunctional characterization of the mating type genes in S. sclerotiorum. These genes are hypothesized to encode master regulatory proteins required foraspects of sexual development ranging from fertilization through fertile fruiting body development. Experimentally, gene deletion strategies wereperformed to create loss-of-function mutants in the two conserved “core” mating type genes common to most ascomycete fungi as well for two lineagespecificgenes found only in S. sclerotiorum and closely related fungi. mat 1-1-1 and mat 1-2-1 mutants are able to form ascogonia but are blocked in allaspects of apothecia development. These mutants also exhibit defects in secondary sexual characters including the production of smaller sclerotia andlower numbers of spermatia. mat 1-2-4 mutants are delayed in apothecia production and form apothecia with aberrant disc morphogenesis and ascosporeproduction. They too produce lower numbers of spermatia and smaller sclerotia and additionally, exhibit a slower hyphal growth rate. Phenotypes of themat 1-1-5 gene knockouts are under evaluation and will be reported. Our findings demonstrate that mat genes are involved in both sexual fertility anddevelopment in S. sclerotiorum.232