Program Book - 27th Fungal Genetics Conference

CONCURRENT SESSION ABSTRACTSand expressed only during the spore stages of the life cycle which are mitotically quiescent, in contrast to other systems where it is expressedconstitutively. Since transformants overexpressing PiCdc14 exhibit normal nuclear behavior, the protein likely does not play a critical role in mitoticprogression although PiCdc14 is known to complement a yeast Cdc14 mutation that normally arrests mitosis. Further investigation into the role of PiCdc14uncovered a novel role. Subcellular localization studies based on fusions with fluorescent tags showed that PiCdc14 first appeared in nuclei during earlysporulation. During the development of biflagellated zoospores from sporangia, PiCdc14 transits to basal bodies, which are the sites from which flagelladevelop. A connection between Cdc14 and flagella is also supported by their phylogenetic distribution, suggesting an ancestral role of Cdc14 in basalbodies and/or flagellated cells. To help unravel the link between PiCdc14 and the flagella apparatus, searches for its interacting partners using both yeasttwo hybrid and affinity purification are underway. Together with colocalization studies involving known basal body/centrosome markers such as centrinand gamma-tubulin, the location and hence the likely roles of PiCdc14 will be revealed.Molecular Determinants of Sporulation in Ashbya gossypii. Jurgen W. Wendland, Lisa Wasserstroem, Klaus Lengeler, Andrea Walther. Yeast Genetics,Carlsberg Laboratory, Copenhagen V, Kopenhagen V, Denmark.Previously we have analysed the pheromone response MAPK signal transduction cascade in A. gossypii. The major findings were (i) deletion of bothpheromone receptor genes STE2 and STE3 did not inhibit sporulation whereas (ii) deletion of the transcription factor STE12 resulted in hypersporulation(Wendland et al. 2011). Here we present our analysis of key A. gossypii homologs of Saccharomyces cerevisiae sporulation specific genes. We show thatmutants in IME1, IME2, KAR4, and NDT80 are blocked in sporulation. Mutants in IME4, KAR4, and UME6 also confer a vegetative growth defect. IME4expression was found during vegetative growth while IME2 was not detected under these conditions. We performed transcriptional profiling of nonsporulatingstrains and determined a core set of about 50 down-regulated sporulation specific genes in these mutants. Interestingly, this set of downregulatedgenes is upregulated in the A. gossypii ste12 mutant providing regulatory evidence of the hypersporulation phenotype of this mutant. Othergenes identified in the RNAseq data indicated that during development of sporangia metabolic genes for nutrient uptake are active. Therefore weperformed Return-To-Growth assays with mutants inhibited in the sporulation pathway. These strains were kept under conditions in which the wild typeinitiates sporulation. This lead to induction of sporangium formation, a stage at which these strains remained. Supply of new nutrients resulted in hyphaloutgrowth in all mutants indicating that after initiation of the sporulation program A. gossypii can reverted to vegetative growth at different stages. Inaddition we identified differential regulation of two endoglucanases encoded by ENG1 and ENG2. While ENG1 was not differentially regulated, ENG2 wasdown-regulated in e.g. ime1 but strongly up-regulated in ste12. Deletion analysis of ENG2 showed that Eng2 is required for hyphal fragmentation intoindividual sporangia. We can thus provide a detailed overview of the genetic regulation of sporulation in A. gossypii. A comparison with S. cerevisiaehighlights the role of KAR4 in sporulation upstream of IME1. Finally, our study provides further evidence that the pheromone signaling response MAPKcascadein A. gossypii has a regulatory control function over sporulation alongside regulation of sporulation by nutritional cues.THE velvet regulators in Aspergilli. Heesoo Park, JJae-Hyuk Yu. Bacteriology, University of Wisconsin Madison, Madison, WI.The velvet regulators are the key players coordinating fungal growth, differentiation and secondary metabolism in response to various internal andexternal cues. All velvet family proteins contain the conserved velvet homology motif (~190 a.a.), and define a novel class of fungal specific transcriptionfactors with the DNA binding ability. Some velvet regulators form time and/or cell type specific complexes with other velvet regulators or non-velvetproteins. These complexes play differential roles in regulating growth, development, sporogenesis and toxigenesis. Among the velvet complexes, the VelB-VosA hetero-complex acts as a functional unit conferring the completion of sporogenesis (focal trehalose biogenesis and spores wall completion), and thelong-term viability of spore, and the attenuation of conidial germination in the model filamentous fungus Aspergillus nidulans. Both velB and vosA areactivated by AbaA in developing cells, and the VelB-VosA complex plays a dual role in activating genes associated with spore maturation and in exertingnegative feedback regulation of developmental genes. Interestingly, the VelB-VosA complex plays similar yet somewhat distinct roles in spore maturation,dormancy and germination in Aspergillus fumigatus and Aspergillus flavus. A comprehensive model depicting the roles of the velvet regulators in aspergilliis presented.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.94