• Author: Sautter, Robert

    College of Arts and Sciences, Florida International University, Miami, United States

  • Author: Ramos, Damaris

    College of Arts and Sciences, Florida International University, Miami, United States

  • Author: Schneper, Lisa

    Herbert Wertheim College of Medicine, Florida International University, Miami, United States

  • Author: Ciofu, Oana

    University of Copenhagen, Denmark

  • Author: Wassermann, Tina

    University of Copenhagen, Denmark

  • Author: Koh, Chong-Lek

    Department of Natural Sciences and Science Education, National Institute of Education, Singapore

  • Author: Heydorn, Arne

    Copenhagen University Hospital, Denmark

  • Author: Hentzer, Morton

    Copenhagen University Hospital, Denmark

  • Author: Høiby, Niels

    University of Copenhagen, Denmark

  • Author: Kharazmi, Arsalan

    Copenhagen University Hospital, Denmark

  • Author: Molin, Søren

    Bacterial Cell Factory, Novo Nordisk Foundation Center for Biosustainability, Center for Systems Microbiology, Department of Systems Biology, Technical University of Denmark, Matematiktorvet, DK-2800, Lyngby, Denmark

  • Author: DeVries, Caroline A.

    University of Tennessee, United States

  • Author: Ohman, Dennis E.

    University of Tennessee, United States

  • Author: Mathee, Kalai

    Herbert Wertheim College of Medicine, Florida International University, Miami, United States

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Infection by the opportunistic pathogen Pseudomonas aeruginosa is a leading cause of morbidity and mortality seen in cystic fibrosis (CF) patients. This is mainly due to the genotypic and phenotypic changes of the bacteria that cause conversion from a typical nonmucoid to a mucoid form in the CF lung. Mucoid conversion is indicative of overproduction of a capsule-like polysaccharide called alginate. The alginate-overproducing (Alg+) mucoid phenotype seen in the CF isolates is extremely unstable. Low oxygen tension growth of mucoid variants readily selects for nonmucoid variants. The switching off mechanism has been mapped to the algT/U locus, and the molecular basis for this conversion was partially attributed to mutations in the algT/U gene itself. To further characterize molecular changes resulting in the unstable phenotype, an isogenic PAO1 derivative that is constitutively Alg+ due to the replacement of the mucA with mucA22 (PDO300) was used. The mucA22 allele is common in mucoid CF isolates. Thirty-four spontaneous nonmucoid variants, or sap (suppressor of alginate production) mutants, of PDO300 were isolated under low oxygen tension. About 40% of the sap mutants were rescued by a plasmid carrying algT/U (Group A). The remaining sap mutants were not (Group B). The members of Group B fall into two subsets: one similar to PAO1, and another comparable to PDO300. Sequence analysis of the algT/U and mucA genes in Group A shows that mucA22 is intact, whereas algT/U contains mutations. Genetic complementation and sequencing of one Group B sap mutant, sap22, revealed that the nonmucoid phenotype was due to the presence of a mutation in PA3257. PA3257 encodes a putative periplasmic protease. Mutation of PA3257 resulted in decreased algT/U expression. Thus, inhibition of algT/U is a primary mechanism for alginate synthesis suppression.
Original languageEnglish
JournalGene
Publication date2012
Volume498
Issue2
Pages242-253
ISSN0378-1119
DOIs
StatePublished
CitationsWeb of Science® Times Cited: 6

Keywords

  • Exopolysaccharide, Alginate, Cystic fibrosis, PalgT/algU, Biosynthetic genes
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