VAAM-Jahrestagung 2011 Karlsruhe, 3.–6. April 2011

MPV009Connecting cell cycle to pathogenic developmentregulatorycascades during pathogenesis of Ustilagomaydis.K. Heimel* 1 , M. Scherer 2 , S. Hassinger 1 , J. Kaemper 11 Institute of Technology and Genetics, Karlsruhe Institute of Technology(KIT), Karlsruhe, Germany2 Qiagen, Hilden, GermanyIn the smut fungus Ustilago maydis, the regulation of cell cycle andmorphogenetic switching during pathogenic and sexual development isorchestrated by the a and b mating-type loci. Activation of either matingtypelocus triggers a G2 cell cycle arrest as a prerequisite for the formationof the infectious dikaryon, which is released only after penetration of thehost plant. The bE/bW heterodimeric transcription factor encoded by the b-mating type locus coordinates a regulatory network consisting of differenttranscription factors. The C2H2 zinc finger transcription factor Rbf1, as amaster regulator, is required for the expression of most (>90%) b-regulatedgenes. Rbf1 is independently from bE/bW sufficient to initiate pathogenicdevelopment, however, further development requires (1) additional factorsas the bE/bW dependently expressed Clp1 protein for cell cycle progressionand (2) additional bE/bW regulated genes to establish the biotrophicinterface. We show that both bW and Rbf1 interact with Clp1. Clp1interaction with bW blocks b-dependent functions, such as the b-dependentG2 cell cycle arrest, dimorphic switching and pathogenic development. Theinteraction of Clp1 with Rbf1 results in the repression of the a-dependentpheromone pathway, conjugation tube formation, and the a-induced G2 cellcycle arrest. Thus, the concerted interaction of Clp1 with Rbf1 and bWcoordinates a- and b-dependent cell cycle control to ensure cell cycle releaseand progression at the onset of biotrophic development.MPV010The molecular basis of symptom formation inSporisorium reilianumY. Zhao*, H. Ghareeb, M.T. Habib, J. SchirawskiMolecular Biology of Plant-Microbe Interactions, Albrecht von HallerInstitute, Göttingen, GermanySporisorium reilianum and Ustilago maydis are closely related biotrophicmaize pathogens that cause different symptoms. S. reilianum infects youngplants, spreads systemically and causes symptoms at the onset of flowering.Symptoms include the formation of spores and leaf-like structures ininflorescences, and an increase in the number of ears formed by the plant. Incontrast, U. maydis can infect the plant via all aerial organs and rapidlyinduces the formation of spore-filled tumors near the site of infection. Thegenomes of S. reilianum and U. maydis are highly syntenic and mostencoded proteins are well conserved. However, a few divergence regionswere identified that encode only weakly conserved proteins[1]. The largestdivergence region is located on chromosome 19 (cluster 19A) and encodesmore than 20 potentially secreted proteins. Deletion of cluster 19A in S.reilianum leads to dramatically reduced virulence, a change in the number ofears per plant, and early senescence of inoculated leaves. By subdeletionanalysis we were able to show that different proteins of cluster 19Acontribute to specific symptoms. Thus, symptom formation is modulated byeffectors of the divergence region cluster 19A of S. reilianum.[1] Schirawski, J. et al (2010): Science 330:1546-1548.MPV011Secreted proteins of the dermatophytic fungusArthroderma benhamiae and their contribution topathogenicityC. Heddergott* 1 , O. Kniemeyer 1 , A.A. Brakhage 1,21 Department of Molecular and Applied Microbiology, Hans-Knöll-Institute(HKI), Jena, Germany2 Institute of Microbiology, Friedrich-Schiller-University, Jena, GermanyTo study the mechanism of keratin degradation and to elucidate hostcolonisation strategies, the secretome of the dermatophytic fungus A.benhamiae was analysed in terms of composition and contribution ofparticular proteins to pathogenesis. Protein secretion is highly up-regulatedduring growth on keratin and the secretome primarily consists of proteasesrepesenting all major functional classes such as fungalysins and subtilisins.By contrast, the hydrofluoric acid - extractable cell wall fraction containedtwo abundant proteins: the constitutively produced hydrophobin Hyp1 andthe keratin-inducible integral cell wall protein Kip1. To characterise theseproteins, deletion mutants of the respective genes were generated.The kip1 deletion mutant did not show an apparent phenotype. By contrast,strains constitutively expressing Kip1 induced an increased interleukin 8response in human keratinocytes during infection, indicating a proinflammatoryeffect of this so far uncharacterised cell wall protein. Analysisof the kip1 promoter region suggests a regulation of the gene by the pHresponse transcription factor PacC. In A. benhamiae, deletion of pacC led toa strain severely affected in morphology and retarded in hyphal growth,showing that in this species the transcription factor exhibits globalregulatory functions. The mutant was still able to grow on keratin butshowed prominent alterations of the secretome, suggesting a multiplicity ofsecretory proteins being de-regulated in this mutant. The conidialhydrophobin of A. fumigatus was described to serve as a protectant againstthe immune recognition by host cells [1]. Inspired by this finding, wecharacterised the hydrophobin Hyp1 of A. benhamiae. Analysis of the hyp1gene deletion mutant indicated that also in A. benhamiae hydrophobin servesas a masking protein. Phenotypically Δhyp1 showed a reduced mycelialhydrophobicity and altered adhesivity of conidia to miscellaneous surfaces.Interestingly, the Δhyp1 strain was recognized more effectively bykeratinocyte and macrophage cell lines and by neutrophil granulocytes(PMN). This data was confirmed by cytokine response measurement and, forthe PMN, by neutrophil extracellular trap (NET) - induction.[1] Aimanianda et al (2009): Nature 460:1117-1121.MPV012Generation and functional characterization of truncatedBartonella henselae BadA mutantsP. Kaiser* 1 , D. Linke 2 , H. Schwarz 2 , V. Kempf* 11 Institute for Medical Microbiology and Infection Control, UniversityHospital Frankfuirt am Main, Frankfurt am Main, Germany2 Max Planck Institut for Developmental Biology, Tübingen, GermanyThe humanpathogenic bacterium Bartonella henselae causes cat scratchdisease and vasculoproliferative disorders (e.g., bacillary angiomatosis).Expression of Bartonella adhesin A (BadA) is crucial for bacterialautoagglutination, adhesion to host cells, binding to extracellular matrixproteins and proangiogenic reprogramming. BadA belongs to the class of thelollipop-like structured trimeric autotransporter adhesins (TAAs) and ismodularly constructed consisting out of a head, a long and repetitive neckstalkmodule and a membrane anchor. Until now, the exact biological rolesof these domains in the infection process remains unknown. Here, weanalyzed the functions of certain BadA domains in greater detail. For thispurpose, deletion mutants were produced by truncating the repetitive neckstalkmodule and deleting different head subdomains of BadA. Likewildtype bacteria, a mutant with a nearly completely truncated stalkexhibited autoagglutination, adhesion to collagens and endothelial cells(ECs) and induced the secretion of proangiogenic cytokines (VEGF).Remarkably, B. henselae expressing only parts of the stalk boundfibronectin. Deletions of several head subdomains revealed no specificattribution of domain-function relationships. Our data revealed that thefibronectin binding ability of the BadA is located in the stalk domain. Theadhesion to collagen and ECs and the secretion of proangiogenic cytokinesis mediated by the neck-stalk module and by the single subdomains of theBadA head. These findings suggest overlapping functions of certain BadAdomains in the infection process of the host.MPV013Interaction of Yersinia spp. with invertebratesT. FuchsDepartment of Microbiology, Central Institute for Food and NutritionResearch (ZIEL), Freising, GermanyThe capability of yersiniae to interact with and to kill insect larvae andbacterivorous predators such as nematodes has only recently beenuncovered. Due to the biphasic pathogenicity of Y. enterocolitica and Y.pseudotuberculosis towards invertebrates and humans, these speciesrepresents the transition from entomopathogenic to humanpathogenicbacteria. This is of particular interest because the associations of microbeswith non-vertebrates might be a yet neglected source of human pathogenspektrum | Tagungsband 2011