Program Book - 27th Fungal Genetics Conference

FULL POSTER SESSION ABSTRACTS(PDE H) and a low affinity (PDE L) phosphodiesterases. The Dpde H strain grows slow and does not conidiate; no evident phenotype was reported for Dpde L.We found that PDE L was mainly responsible for the cAMP decrease during the first HO and that hyphal adhesion was retarded in Dpde L. Both PDE H andPDE L were responsible for cAMP decrease during the second HO. H 2O 2 and low Ca ++ activated PDE L and inhibited PDE H. This opposite regulation can explainthe cAMP decrease during the HOs of the N. crassa conidiation process. [1] Toledo I et al. (1986) Aerial growth in Neurospora crassa: characterization of anexperimental model system. Exp Mycol. 10:114-125. [2] Hansberg W; Aguirre J (1990) Hyperoxidant states cause microbial cell differentiation by cellisolation from dioxygen. J Theoret Biol 142:201-221. [3] Hansberg W et al. (2008) Cell differentiation as a response to oxidative stress. In: Stress in Yeasts &Filamentous Fungi (Ed. Avery et al.) Elsevier IBSN 978-0-12-374184-4.124. Ste12 is a negative regulator of conidiation and cell wall lytic enzymes production in response to nitrogen deprivation and light in Trichodermaatroviride. Maria Fernanda Nieto-Jacobo 1 , Alfredo Herrera-Estrella 2 , Alison Stewart 1 , Artemio Mendoza-Mendoza 1 . 1) Bioprotection Research Centre,Lincoln University, Lincoln, Canterbury, New Zealand; 2) Laboratorio Nacional de Genómica para la Biodiversidad, Centro de Investigación y de EstudiosAvanzados del IPN Sede Irapuato, Irapuato 36821, Guanajuato, Mexico.Ste12 is a transcription factor found exclusively in the fungal kingdom. In Saccharomyces cerevisiae, Ste12 regulates mating and invasive/pseudohyphalgrowth, while in saprophytic and parasitic filamentous fungi Ste12-like proteins control mating, plant penetration and invasive growth. Ste12 and Ste12-like proteins are downstream components of the MAPK PMK1 pathway which are capable of regulating several genes encoding fungal virulence factorsinvolved in both plant and animal infection. Among the virulence factors are diverse range of lytic enzymes and cell surface components. Several membersTrichoderma genus are mycoparasites of plant fungal pathogens; so they are widely used as biocontrol agents. In addition, Trichoderma spp. penetrateplant roots and establish beneficial relationships with their host. One crucial element in biocontrol activity and root colonization of Trichoderma is thesynthesis of lytic enzymes. Several lytic enzymes in Trichoderma are regulated by nitrogen metabolite repression. Here we observed that the ste12-liketranscription factor gene is highly up-regulated when Trichoderma is grown on nitrogen depleted medium. To find the role of ste12 in Trichoderma, aste12-like orthologue gene was deleted in T. atroviride and the effects on fungal development and response to different biotic and abiotic stimulievaluated. Our results demonstrate that growth and conidiation of a T. atroviride Ste12-like mutant was only slightly altered in complete media. Weevaluated the ability of the Dste12-like mutant to use a variety of nitrogen sources using Biolog microtiter plates. We noticed that when essential aminoacids are used as the sole nitrogen source, the deletion mutant grew faster than the wild type, however this situation did not occur when the same aminoacids were used as the sole carbon source. In addition, induction of conidiation in response to light or mechanical injury was stronger in the Dste12-likemutants than in the wild type but only when a secondary nitrogen source was used in the medium. Finally we observed that some lytic enzymes aredifferently produced between the wild type and Dste12-like mutants under nitrogen deprivation conditions. We propose that the T. atroviride Ste12-likeorthologue regulates lytic enzymes and conidiation by a mechanism that involves nitrogen catabolite repression.125. Black holes in fungal virulence: loss of RNAi in C. gattii outbreak strains reveals a novel RNAi factor. Marianna Feretzaki, Xuying Wang, BlakeBilmyre, Joseph Heitman. Molecular Genetics and Microbiology, Duke University Medical Center, Durham, NC.Genome instability and mutations provoked by transposon movement are counteracted by novel defense mechanisms in organisms as diverse as fungi,plants, and mammals. In the human fungal pathogen Cryptococcus neoformans we have previously characterized an RNAi silencing pathway that defendsthe genome against mobile elements and artificially introduced repeats of homologous DNA. Repetitive transgenes and transposons are silenced by anRNAi-dependent pathway during sexual development (sex-induced silencing, SIS) and during vegetative mitotic growth (MIS). RNAi silencing pathways areconserved in the Cryptococcus pathogenic species complex and are mediated by core RNAi components, including an RNA-dependent RNA polymerase(Rdp1), Argonaute (Ago1) and Dicer (Dcr1 and Dcr2). Surprisingly, all of the canonical known RNAi components are missing from all C. gattii VGII strains,the molecular type responsible for the North American Pacific Northwest outbreak. To identify novel components of the RNAi pathway, we surveyed thegenome of the C. gattii R265 isolate for missing genes. One of the most interesting is ZNF3. In previous studies we found that Znf3, a protein with threezinc finger domains, is required for opposite- and same-sex mating in C. neoformans var. neoformans. Surprisingly, in C. neoformans var. grubii ZNF3 is notessential for sexual development. However, it is required for mitotic- and sex-induced silencing via RNAi. SIS is less efficient in znf3D unilateral matings andis abolished in znf3D x znf3D bilateral matings, similar to the phenotypes of rdp1D mutants. Znf3 is also required for transgene-induced mitotic silencing;znf3D mutations abrogate silencing of repetitive transgenes during vegetative growth. Znf3 tagged with mCherry is localized in the cytoplasm in bright,distinct foci. Co-localization of Znf3 with the P-body marker Dcp1-GFP further supports the hypothesis that Znf3 is a novel element of the RNAi pathwayand operates to defend the genome during sexual development and vegetative growth.126. The Crz1/Sp1 transcription factor of Cryptococcus neoformans is activated by calcineurin and regulates cell wall integrity. Sophie Lev 1 , DesmariniDesmarini 1 , Methee Chayakulkeeree 2 , Tania Sorrell 1 , Julianne Djordjevic 1 . 1) Centre for Infectious Diseases and Microbiology, Sydney Medical School andWestmead Millennium Institute, University of Sydney, Westmead 2145 NSW, Australia; 2) Faculty of Medicine, Siriraj Hospital, Mahidol University,Bangkok, Thailand.Cryptococcus neoformans survives host temperature and regulates cell wall integrity via a calcium-dependent phosphatase, calcineurin. However,downstream effectors of C. neoformans calcineurin are largely unknown. In S. cerevisiae and other fungal species, a calcineurin-dependent transcriptionfactor Crz1 translocates to nuclei upon activation and triggers expression of target genes. We now show that the C. neoformans Crz1 ortholog (Crz1/Sp1),previously identified as a protein kinase C target during starvation, is a bona fide target of calcineurin under non-starvation conditions, during cell wallstress and growth at high temperature. Both the calcineurin-defective mutant, Dcna1, and a CRZ1/SP1 mutant (Dcrz1) were susceptible to cell wallperturbing agents. Furthermore, expression of the chitin synthase encoding gene, CHS6, was reduced in both mutants. We tracked the subcellularlocalization of Crz1-GFP in WT C. neoformans and Dcna1 in response to different stimuli, in the presence and absence of the calcineurin inhibitor, FK506.Exposure to elevated temperature (30-37°C vs 25°C) and extracellular calcium caused calcineurin-dependent nuclear accumulation of Crz1-GFP.Unexpectedly, 1M salt and heat shock triggered calcineurin-independent Crz1-GFP sequestration within cytosolic and nuclear puncta. To our knowledge,punctate cytosolic distribution, as opposed to nuclear targeting, is a unique feature of C. neoformans Crz1. We conclude that Crz1 is selectively activatedby calcium/calcineurin-dependent and independent signals depending on the environmental conditions.127. A Fungal Adhesin Guides Community Behaviors by Autoinduction and Paracrinal Signaling. Linqi Wang, Xunyun Tian, Rachana Gyawali, Xiaorong Lin.Biology, Texas A&M University, College Station, TX.Microbes live mostly in a social community rather than in a planktonic state. Such communities have complex spatiotemporal patterns that requireintercellular communication to coordinate gene expression. Here, we demonstrate that Cryptococcus neoformans, a model eukaryotic pathogen, respondsto an extracellular signal in constructing its colony morphology. The signal that directs this community behavior is not a molecule of low molecular weight152