title: Occurrence of mutations impairing sigma factor B (SigB) function upon inactivation of Listeria monocytogenes genes encoding surface proteins.
abstract: Bacteria of the genus Listeria contain the largest family of LPXTG surface proteins covalently anchored to the peptidoglycan. The extent to which these proteins may function or be regulated cooperatively is at present unknown. Because of their unique cellular location, we reasoned that distinct LPXTG proteins could act as elements contributing to cell wall homeostasis or influencing the stability of other surface proteins bound to peptidoglycan. To test this hypothesis, we used proteomics to analyse mutants of the intracellular pathogen Listeria monocytogenes lacking distinct LPXTG proteins implicated in pathogen-host interactions, such as InlA, InlF, InlG, InlH, InlJ, LapB and Vip. Changes in the cell wall proteome were found in inlG and vip mutants, which exhibited reduced levels of the LPXTG proteins InlH, Lmo0610, Lmo0880 and Lmo2085, all regulated by the stress-related sigma factor SigB. The ultimate basis of this alteration was uncovered by genome sequencing, which revealed that these inlG and vip mutants carried loss-of-function mutations in the rsbS, rsbU and rsbV genes encoding regulatory proteins that control SigB activity. Attempts to recapitulate this negative selection of SigB in a large series of new inlG or vip mutants constructed for this purpose were, however, unsuccessful. These results indicate that inadvertent secondary mutations affecting SigB functionality can randomly arise in L. monocytogenes when using common genetic procedures or during subculturing. Testing of SigB activity could be therefore valuable when manipulating genetically L. monocytogenes prior to any subsequent phenotypic analysis. This test may be even more justified when generating deletions affecting cell envelope components.
authors: Centro Nacional de Biotecnología-Consejo Superior de Investigaciones Científicas (CNB-CSIC), 28049 Madrid, Spain.
authors: Juan J
authors: JJ
authors: Quereda
authors: Departamento de Biología Molecular, Universidad Autónoma de Madrid. Centro de Biología Molecular 'Severo Ochoa' (CBMSO-CSIC), 28049 Madrid, Spain.|Centro Nacional de Biotecnología-Consejo Superior de Investigaciones Científicas (CNB-CSIC), 28049 Madrid, Spain.
authors: M Graciela
authors: MG
authors: Pucciarelli
authors: Centro Nacional de Biotecnología-Consejo Superior de Investigaciones Científicas (CNB-CSIC), 28049 Madrid, Spain.
authors: Laura
authors: L
authors: Botello-Morte
authors: Unidad de Proteómica, Centro Nacional Investigaciones Cardiovasculares (CNIC), 28029 Madrid, Spain.
authors: Enrique
authors: E
authors: Calvo
authors: Group of Molecular Microbiology, Instituto de Biologia Molecular e Celular, Universidade do Porto, Porto, Portugal.
authors: Filipe
authors: F
authors: Carvalho
authors: Institut Pasteur, Plate-forme PF1 Génomique, Département Génomes et Génétique, Paris, France.
authors: Christiane
authors: C
authors: Bouchier
authors: Group of Molecular Microbiology, Instituto de Biologia Molecular e Celular, Universidade do Porto, Porto, Portugal.
authors: Ana
authors: A
authors: Vieira
authors: Centro Nacional de Biotecnología-Consejo Superior de Investigaciones Científicas (CNB-CSIC), 28049 Madrid, Spain.
authors: Javier F
authors: JF
authors: Mariscotti
authors: Institute of Medical Microbiology, Justus-Liebig-University, Giessen, D-35392, Germany.
authors: Trinad
authors: T
authors: Chakraborty
authors: Unité des Interactions Bactéries-Cellules, Institut National de la Santé et de la Recherche Médicale (INSERM) U604, Institut Pasteur, and the Institut Scientifique de Recherche Agronomique (INRA) USC2020, Institut Pasteur, Paris F-75015, France.
authors: Pascale
authors: P
authors: Cossart
authors: Institute of Medical Microbiology, Justus-Liebig-University, Giessen, D-35392, Germany.
authors: Torsten
authors: T
authors: Hain
authors: Group of Molecular Microbiology, Instituto de Biologia Molecular e Celular, Universidade do Porto, Porto, Portugal.
authors: Didier
authors: D
authors: Cabanes
authors: Centro Nacional de Biotecnología-Consejo Superior de Investigaciones Científicas (CNB-CSIC), 28049 Madrid, Spain.
authors: Francisco
authors: F
authors: García-Del Portillo
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D002473:Cell Wall / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D054884:Host-Pathogen Interactions; D006801:Humans; D008089:Listeria monocytogenes / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D009154:Mutation; D020543:Proteome; D040901:Proteomics; D012808:Sigma Factor / Q000235:genetics / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D008565:Membrane Proteins; D020543:Proteome; C480945:SigB protein, Listeria monocytogenes; D012808:Sigma Factor