Program Book - 27th Fungal Genetics Conference

CONCURRENT SESSION ABSTRACTSWednesday, March 13 3:00 PM–6:00 PMHeatherMembrane Trafficking and Molecular OrganizationCo-chairs: Vicky Sophianopoulou and Gero SteinbergDistinct secretion systems operate during biotrophic invasion by the rice blast fungus, Magnaporthe oryzae. Barbara Valent 1 , Martha Giraldo 1 , ChangHyun Khang 1,4 , Yasin Dagdas 2 , Yogesh Gupta 2 , Thomas Mentlak 2,5 , Mihwa Yi 1 , Melinda Dalby 1 , Hiromasa Saitoh 3 , Ryohei Terauchi 3 , Nicholas Talbot 2 . 1) DeptPlant Pathology, Kansas State Univ, Manhattan, KS; 2) School of Biosciences, Univ of Exeter, Exeter, U.K; 3) Iwate Biotechnology Research Center, Kitakami,Iwate, Japan; 4) Current Address: Dept of Plant Biology, Univ of Georgia, Athens, GA; 5) Current Address: Cambridge Consultants Ltd, Cambridge, U.K.During biotrophic invasion, Magnaporthe oryzae secretes cytoplasmic effectors, which preferentially accumulate in biotrophic interfacial complexes(BICs) and are translocated into the cytoplasm of the rice cells, and apoplastic effectors, which remain in the extracellular space between the fungal cellwall and the rice plasma membrane. BICs localize in front of the tips of filamentous hyphae that enter rice cells, and remain subapically beside the firstbulbous invasive hyphal cells after hyphal differentiation. In contrast, secreted apoplastic effectors uniformly outline the entire bulbous invasive hypha.We have determined that cytoplasmic effector genes were highly up-regulated in the BIC-associated cells at early invasion stages, and that effectorpromoters played the major role in determining preferential BIC localization of cytoplasmic effectors. Subapical BIC-associated hyphal cells continued toexpress protein secretion machinery components while invasive hyphae grew elsewhere in the host cell, suggesting that these subapical invasive hyphalcells are involved in active secretion. Disruption of the conventional ER-Golgi secretion pathway by Brefeldin A treatment blocked secretion of apoplasticeffectors, but not secretion of cytoplasmic effectors. Pathogen mutants that failed to express exocyst complex components or a t-SNARE were defective insecretion of cytoplasmic effectors, as well as defective in pathogenicity. In contrast, secretion of apoplastic effectors was not impaired in these mutants.Our data suggest that M. oryzae possesses distinct secretory mechanisms for targeting cytoplasmic and apoplastic effectors during rice invasion.The cellular role of early endosome motility in Ustilago maydis. Yujiro Higuchi 1 , Peter Ashwin 2 , Gero Steinberg 1 . 1) Biosciences, University of Exeter,Exeter EX4 4QD, UK; 2) Mathematics Research Institute, University of Exeter, Exeter EX4 4QF, UK.Early endosomes (EEs) are dynamic organelles that move along microtubules, which is mediated by the motor proteins kinesin-3 and dynein. Despite ourgrowing knowledge about the mechanistics of motion, the physiological significance of EE motility remains elusive. A recent study suggested that RNAbindingproteins travel on EEs, which might support local protein translation at the hyphal tip. However, evidence for apical translation is missing. Here, weinvestigate the distribution of ribosomes, using native levels of ribosomal proteins. We will summarize our findings on protein translation and will discussthe role of EE-dependent transport of RNA-binding proteins in the light of our findings.The arrestin-like protein ArtA is essential for ubiquitylation and endocytosis of the UapA transporter in response to both broad-range and specificsignals. George Diallinas, Mayia Karachaliou, Sotiris Amillis, Minos Evangelinos, Alexandros Kokotos. Faculty of Biology, University of Athens, Athens,Greece.We investigated the role of all arrestin-like proteins of Aspergillus nidulans in respect to growth, morphology, sensitivity to drugs and specifically for theendocytosis and turnover of the uric acid-xanthine transporter UapA. All arrestin null mutants are viable showing wild-type growth and morphology,except one which is affected in conidiospore production, but several have modified profiles in respect to N or C source utilization and drug sensitivity. Asingle arrestin, ArtA, is essential for HulARsp5-dependent ubiquitination and endocytosis of UapA in response to ammonium or substrates. Geneticanalysis further showed that residues 545-561 of the UapA C-tail, which includes a critical di-acidic motif, is required for efficient UapA endocytosis.Mutational analysis of ArtA shows that the N-terminal region (2-123) and both PY elements are essential for its function. ArtA undergoes HulA-dependentubiquitination at residue Lys343 and this modification is critical for the efficiency of UapA ubiquitination and endocytosis, especially in response toammonium. Lastly, we show that ArtA is essential for vacuolar turnover of transporters specific for purines (AzgA) or L-proline (PrnB), but not for anaspartate/glutamate transporter (AgtA). Our results are discussed within the frame of recently proposed mechanisms on how arrestins are activated andrecruited for ubiquitination of transporters in response to broad range signals, but also put the basis for understanding how arrestins, such as ArtA,regulate the turnover of a specific transporter in the presence of its substrates.Escaping the hustle - zones of differential protein turnover in the yeast plasma membrane. Guido Grossmann 1,2 , Vendula Stradalova 3 , MichaelaBlazikova 3 , Miroslava Opekarová 4 , Jan Malinsky 3 , Widmar Tanner 5 . 1) Center for Organismal Studies, University of Heidelberg, Heidelberg, Germany; 2)Department for Plant Biology, Carnegie Institution for Science, Stanford, CA; 3) Institute of Experimental Medicine, Academy of Sciences of the CzechRepublic, Prague, Czech Republic; 4) Institute of Microbiology, Academy of Sciences of the Czech Republic, Prague, Czech Republic; 5) Cell biology andplant biochemistry, University of Regensburg, Regensburg, Germany.The plasma membrane (PM) consists of specialized domains that differ in their protein distribution, lipid composition and structure, and that areessential for PM functions such as membrane transport or signal perception and transduction. The mechanisms that establish and maintain thisheterogeneity are still largely unknown but involve interactions between membrane constituents, local modification of the membrane structure, tetheringto other cellular components like ECM/cell wall or cytoskeleton, and polarized exo- and endocytosis. In the PM of bakers yeast, stable rod-shapedmembrane invaginations exist, called membrane compartment of Can1 (MCC), that exhibit a specific composition of lipids and proteins, and are stabilizedby a protein structure called eisosome. Chemical and genetic screens, revealed important roles of the membrane potential, lipid composition, and proteinscaffolds in organizing the PM into specialized domains. The distribution of MCC domains further determines the distribution of the PM-associated corticalER that dynamically covers large areas of the PM. Mapping of endocytic events revealed very low rates of clathrin-mediated endocytosis in PM areascovered by cER and within MCC. The formation of such "quiet zones" provides a mechanism for membrane domain formation through local confinementof membrane turnover.27th Fungal Genetics Conference | 35