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

FGP015Comparative genomics and transcriptomics ofPropionibacterium acnesE. Brzuszkiewicz 1 , J. Weiner 2 , A. Wollherr 1 , A. Thürmer 1 , G. Gottschalk 1 ,R. Daniel 1 , T.F. Meyer 3 , H.J. Mollenkopf 4 , H. Brüggemann* 31 Institute of Microbiology and Genetics, Georg-August-University,Göttingen, Germany2 Department of Immunology, Max Planck Institute for Infection Biology,Berlin, Germany3 Department of Molecular Biology, Max Planck Institute for InfectionBiology, Berlin, Germany4 Core Facility Microarray, Max Planck Institute for Infection Biology,Berlin, GermanyThe anaerobic Gram-positive bacterium Propionibacterium acnes is ahuman skin commensal, but is occasionally associated with inflammatorydiseases. Recent work has indicated that evolutionary distinct lineages of P.acnes play etiologic roles in disease while others are associated with health.To shed light on the molecular basis for differential strain properties, wecarried out genomic and transcriptomic analysis of distinct P. acnes strains.We sequenced the genome of the P. acnes strain 266, a type I-1a sequencetype (ST) 18 strain. Comparative genome analysis of strain 266 and fourother P. acnes strains revealed that overall genome plasticity is relativelylow; however, a number of island-like genomic regions, encoding a varietyof putative virulence-associated and fitness traits, differ between phylotypes.Comparative transcriptome analysis revealed that 225 genes of strainKPA171202 (type I-2, ST34) were differentially transcribed in strain 266during exponential growth. 47% of these genes belong to the strain-specificgene content of strain KPA171202, indicating that strain-specific functionsare utilized. Next, we studied differential expression during exponential andstationary growth phases. Genes encoding components of the energyconservingrespiratory chain as well as secreted and virulence-associatedfactors were transcribed during the exponential phase, while the stationarygrowth phase was characterized by up-regulation of genes involved in thestress response and amino acid metabolism. Taken together, our datahighlight the genomic basis for strain diversity and identify, for the firsttime, the transcribed part of the genome, underling the important role activegrowth plays in the inflammatory activity of P. acnes. We argue that thedisease-causing potential of different P. acnes strains is not only determinedby variable genome content but also, and to a greater degree, by variabletranscriptomes.FGP016Deletion analysis reveals essential genes within thegenomic magnetosome island of MagnetospirillumgryphiswaldenseA. Lohße*, S. Ullrich, E. Katzmann, D. SchülerDepartment Biologie I, Ludwig-Maximilians-University, Munich, GermanyThe magnetotactic bacterium M. gryphiswaldense synthesizes intracellularmembrane-enclosed crystals, which consist of the ferrimagnetic mineralmagnetite (Fe 3O 4) referred to as magnetosomes. The biomineralization ofmagnetosomes is controlled by a specific set of genes, which are locatedwithin the conserved magnetosome island (MAI). Beside the mam and mmsgenes, encoding magnetosome proteins, the 130-kb region contains inaddition numerous genes for transposases, pseudogenes and hypotheticalgenes of unknown functions. In order to reveal putative functions inmagnetosome formation, deletion of the mms6-, mamGFDC-, and mamXYoperons lead to severe defects in morphology, size and chain assembly ofmagnetite crystals. However, even multiple deletions including variouscombinations of the mamXY- and mamGFDC operons did not entirelyabolish biomineralization, although only tiny and irregular crystallites wereformed. In contrast, deletion of the 16 kb mamAB operon resulted in thecomplete loss of magnetosomes. This suggests that while several regionswithin the MAI are irrelevant for magnetosome formation, other haveaccessory functions, and only the mamAB operon harbors genes that areabsolutely essential for magnetosome formation. In conclusion, ourapproach will help determining the minimal gene set required formagnetosome synthesis and is promising for future „synthetic biology”approaches.FGP017Functional Networks of Light Controlled Processes:Identification of Regulatory FactorsS. Wolfers* 1,2 , U. Kück 1,21 Department of General and Molecular Botany, Ruhr-University,Bochum,Germany2 Christian Doppler Laboratory for "Biotechnology of Fungi", Ruhr-University, Bochum, GermanyIn previous research, many responses to external and internal stimuli havebeen identified to regulate gene expression in the industrial penicillinproducer Penicillium chrysogenum. Light for instance acts as a major carrierof information, but in case of P. chrysogenum there is little known about theeffect of illumination on regulatory networks. It has been shown, that lighthas an effect on morphology and secondary metabolite production, althoughonly few regulators have been found so far on the molecular level. Toidentify light induced regulatory responses, and the proteins involved, weused microarray-analysis as an experimental approach. The expressionlevels of cultures grown in constant (white) light were compared to those ofcultures grown in darkness for the same time period, thus we were able toidentify genes differently regulated due to illumination. We first looked atthe intersection between genes newly found in this approach and sets ofgenes from previous microarray experiments to reduce the number ofcandidate genes for further analysis. In these experiments expression levelswere compared using wild type, and disruption strains with deleted genesencoding core elements of the velvet complex [1]. To identify light inducedregulatory factors we screened candidate genes for putative transcriptionfactors. Subsequently we have generated deletion strains using the FLP/FRTrecombination system [2] for further characterisation of selected putativetranscription factors.[1] Hoff, B. et al (2010): Two components of a velvet-like complex control hyphal morphogenesis,conidiophore development, and penicillin biosynthesis in Penicillium chrysogenum. Eukaryot Cell 9:1236-50[2] Kopke, K. et al (2010): Application of the Saccharomyces cerevisiae FLP/FRT recombinationsystem in filamentous fungi for marker recycling and construction of knockout strains devoid ofheterologous genes. Appl Environ Microbiol 76: 4664-74FGP018PcchiBI is a target gene of PcVelA in producer strains ofP. chrysogenumJ. Kamerewerd* 1,2 , U. Kück 1,21 Department of General and Molecular Botany, Ruhr-University, Bochum,Germany2 Christian Doppler Laboratory for "Biotechnology of Fungi", Ruhr-University, Bochum, GermanyFungal cell walls are highly dynamic structures with a wide range ofessential roles in fungal development and interaction with the environment.One main component of fungal cell walls is chitin, a β-(1,4)-linkedhomopolymer of N-acetyl-D-glucosamine (GlcNAc) subunits. To maintainthe plasticity of the cell wall, fungi possess a multiplicity of cell wallmodifying enzymes, for example hydrolases involved in the degradation ofcell wall components. Chitinases (EC 3.2.1.14) hydrolyze chitin randomly atinternal sites to generate low molecular mass chitooligomers and can befound in a wide range of organisms. In fungi, only chitinases of glycosylhydrolase family 18 (GH18) with morphogenetic, autolytic and nutritionalroles are described. According to the CAZy-database, 8 ORFs encodingputative chitinases can be found in the genome of Penicillium chrysogenum.Recently we have reported data from microarray analysis showing that genesinvolved in chitin catabolism are strongly downregulated in a ∆PcVelAmutant of P. chrysogenum lacking the global regulator protein PcVelA, ahomologue of the VeA protein from Aspergillus nidulans. In order toanalyze the biological function of a target gene of PcVelA encoding aputative class V chitinase, a disruption strain was generated. The sum of ouranalysis indicates functional similarities and differences of this chitinase incomparison to homologous proteins from different Aspergillus species,illustrating the plasticity of class V chitinases in filamentous fungi.spektrum | Tagungsband 2011