Program Book - 27th Fungal Genetics Conference

FULL POSTER SESSION ABSTRACTSA screen of mutants in Neurospora crassa single gene deletion library identified 24 cell fusion genes. Bioinformatics studies indicate that 14 of thesegenes are likely to function in signal transduction pathways, 4 genes are transcription factors, 3 genes are likely to be involved in the process of vesiculartrafficking, and 3 genes are highly conserved in fungal species with unknown functions. GFP and RFP fusion proteins were constructed for 2 vesiculartrafficking proteins AMPH-1 and HAM-10, and 1 conserved hypothetical protein HAM-8 to study their functions during cell fusion process. Fluorescentprotein markers for cellular organelles (including nucleus, mitochondria, golgi apparatus, endoplasmic reticulum, vacuole and vesicle), and for cytoskeleton(including actin filament and microtubule) were obtained from Fungal Genetic Stock Center. Strains expressing individual fluorescent protein marker wereused to study the cellular localizations of AMPH-1, HAM-10 and HAM-8 by using fluorescent confocal microscopy. The fluorescent protein marker strainswere also used to study the dynamics of organelle movements during cell fusion by using time-lapse fluorescence microscopy. Fluorescent signals fromAMPH-1, HAM-10 and HAM-8 were compared with two signaling molecules MAK-2 and SO to study their potential involvement in signal transduction.Results shown AMPH-1, HAM-8 and HAM-10 all colocalize with vesicle marker. One of the conserved hypothetical proteins, HAM-6, was modified with aFLAG tag to study its functions during cell fusion.154. Identification of novel Neurospora crassa genes involved in hyphal fusion by tanscriptomic analysis. Wilfried Jonkers, Abigail C. Leeder, N. LouiseGlass. Department of Plant and Microbial Biology, University of California, Berkeley, CA.Hyphal fusion of Neurospora crassa germlings is a highly regulated process involving -among others- the conserved MAP kinase MAK-2 and the SOprotein of unknown biochemical function. During chemotrophic interactions between two genetically identical germlings, MAK-2 and SO alternatelylocalize at the conidial anastomosis tubes (CATs) every 4 minutes, perfectly out of phase of each other. How this process is initiated, maintained and whatother proteins are involved is still unknown. One conserved fungal target of MAK-2 is the yeast Ste12-like transcription factor, named PP-1. Similar to mak-2, pp-1 is also required for hyphal fusion and normal mycelial growth. To identify downstream targets of MAK-2 and PP-1 that may play a role in germlingfusion, micro-array and RNAseq analyses were performed on wild type (WT) and Dpp-1 strains. Combining the micro-array and RNAseq data, 32 geneswere identified that showed at least 2-fold differential expression in WT as compared to Dpp-1. These include six genes, which are homologs of yeastSte12 targets. To test the involvement of these genes in hyphal fusion, a deletion strain was obtained or constructed and assayed for germling fusionphenotype. Three deletion strains were completely devoid of fusion: Dham-7, Dasm-1 and Dham-11, and one deletion strain, Dham-12 showed reducedfusion frequencies when compared to WT. ham-7 was previously identified as fusion gene while asm-1 was shown to be involved in meiosis. When Dham-7 + Dham-7 or Dham-7 + WT germlings are confronted which each other, chemotropic interactions are not initiated, CATs are not observed and MAK-2 andSO are localized predominantly to the cytoplasm. ham-11 is a newly identified gene involved in germling fusion; Dham-11 + Dham-11 germlings do notshow chemotropic interactions or cell fusion. However, in contrast to Dham-7, Dham-11 germling fuse normally with WT germlings. MAK-2 and SO alsoshow normal oscillation in WT and Dham-11 germlings undergoing chemotropic growth. The observations suggest that HAM-11 might be involved in theproduction or proper release of a signal capable of inducing cell recognition and the germling fusion process.155. N-acetylglucosamine (GlcNAc) Triggers a Morphogenetic Program in Systemic Dimorphic Fungi. Sarah A. Gilmore 1 , Shamoon Naseem 2 , James B.Konopka 2 , Anita Sil 1 . 1) Department of Microbiology and Immunology, University of California, San Francisco, San Francisco, CA; 2) Department ofMolecular Genetics and Microbiology, Stony Brook University, Stony Brook, NY.Cellular differentiation is an essential process for the development and growth of multicellular eukaryotic organisms. Similarly, many unicellularorganisms undergo a program of cellular differentiation to produce a new cell type specialized for survival in a distinct environmental niche. Systemicdimorphic fungal pathogens, such as Histoplasma capsulatum (Hc) and Blastomyces dermatitidis (Bd), can switch between a unicellular parasitic yeast formadapted for growth within mammals and an infectious soil-growing filamentous form as part of their natural life cycles. Temperature is thought to be thepredominant environmental cue that promotes cellular differentiation of systemic dimorphic fungi; however, work with other fungi indicates thatadditional environmental cues including CO2, light, and nutrient availability can influence how an organism responds to its environment. Recent worksuggests that the ubiquitous monosaccharide N-acetylglucosamine (GlcNAc) can play a role in cell signaling in fungi. We identified GlcNAc as a potentinducer of the yeast-to-filament transition in Hc and Bd. Micromolar concentrations of exogenous GlcNAc were sufficient to induce a robust morphologicaltransition of Hc yeast cells to filamentous cells at room temperature, indicating that dimorphic fungal cells may be sensing GlcNAc, or one of its catabolicbyproducts, to promote filamentation. Using GlcNAc as a tool to induce a robust and more synchronous phase transition of Hc yeast cells to filaments, weexamined the temporal regulation of the Hc transcriptome during morphogenesis to reveal candidate genes involved in establishing the filamentousgrowth program. Two genes we identified during transcriptome analysis included NGT1 and NGT2, which encode GlcNAc major facilitator superfamilytransporters. RNAi depletion of NGT1 or NGT2 rendered Hc cells unable to respond to exogenous GlcNAc. Furthermore, wild type levels of NGT1 and NGT2transcripts were important for efficient Hc yeast-to-filament conversion even in the absence of exogenously added GlcNAc. These data suggest that Ngt1and Ngt2 may monitor endogenous GlcNAc as part of an autoregulatory system that allows Hc to regulate its filamentous growth.156. How water influences fungal growth on "real" materials. H.P. Huinink 1 , K.A. Laarhoven, van 1 , M. Bekker 1 , J. Dijksterhuis 2 , O.C.G. Adan 1 . 1) AppliedPhysics, Eindhoven University of Technology, Eindhoven, Netherlands; 2) CBS - KNAW, Utrecht, Netherlands.Understanding fungal growth on construction materials is important to control problems with mould growth in buildings. The indoor environment isgenerally a harsh environment for a fungus. The climate is relatively dry and only during certain events at specific locations in the building (cooking,showering, etc.) there are peaks in the humidity. The porous nature of construction materials seems to play an important role in the survival of organisms,because it buffers the climate at the surface of materials by storing water. A model has been developed that describes the thermodynamic state and flowof water inside porous materials in connection to the growth of the organism. The model shows that the activity of water in a material is the keyparameter controlling growth. However, the model also proves that growth cannot be predicted on the basis of experiments performed on idealizedmicrobiological media (agar) with a well defined water activity. In those media water is always abundantly present irrespective of the activity. In porousmaterials however the amount of water dramatically reduces with the water activity. It is shown that porous materials with small pores in general containmore water than materials with big pores. A drop in the amount of water due to a decreasing activity has direct consequences for the food supply.Whereas in idealized media the amount of water is very high and therefore the mobility of nutrients, in porous materials the mobility of nutrients willdecrease with decreasing water activity. To understand the behavior of a fungus on materials, its growth has to be really studied on these materials.157. Identification and characterization of two genes required in the control of a cell degeneration in the filamentous fungi Podospora anserina. HerveLalucque 1,2 , Fabienne Malagnac 1,2 , Pierre Grognet 1,2 , Philippe Silar 1,2 . 1) Univ Paris Diderot, Sorbonne Paris Cité, Laboratoire Interdisciplinaire des Energiesde demain (LIED), 75205 Paris France; 2) Institut de Génétique et Microbiologie (IGM), UMR 8621 CNRS Univ Paris Sud, 91405 Orsay France.For several years, we use the coprophilous fungus Podospora anserina to study a cell degeneration called Crippled Growth (CG) triggered by an27th Fungal Genetics Conference | 159