title: A complex multilevel attack on Pseudomonas aeruginosa algT/U expression and algT/U activity results in the loss of alginate production.
abstract: 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.
authors.affiliation: Department of Biological Sciences, College of Arts and Sciences, Florida International University, Miami, FL 33199, USA.
authors.forename: Robert
authors.initials: R
authors.lastname: Sautter
authors.forename: Damaris
authors.initials: D
authors.lastname: Ramos
authors.forename: Lisa
authors.initials: L
authors.lastname: Schneper
authors.forename: Oana
authors.initials: O
authors.lastname: Ciofu
authors.forename: Tina
authors.initials: T
authors.lastname: Wassermann
authors.forename: Chong-Lek
authors.initials: CL
authors.lastname: Koh
authors.forename: Arne
authors.initials: A
authors.lastname: Heydorn
authors.forename: Morton
authors.initials: M
authors.lastname: Hentzer
authors.forename: Niels
authors.initials: N
authors.lastname: Høiby
authors.forename: Arsalan
authors.initials: A
authors.lastname: Kharazmi
authors.forename: Søren
authors.initials: S
authors.lastname: Molin
authors.forename: Caroline A
authors.initials: CA
authors.lastname: Devries
authors.forename: Dennis E
authors.initials: DE
authors.lastname: Ohman
authors.forename: Kalai
authors.initials: K
authors.lastname: Mathee
mesh_terms: D000464:Alginates / Q000378:metabolism*; D000595:Amino Acid Sequence; D001425:Bacterial Outer Membrane Proteins / Q000235:genetics / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D004251:DNA Transposable Elements; D015964:Gene Expression Regulation, Bacterial; D005816:Genetic Complementation Test; D020723:Glucuronic Acid / Q000378:metabolism; D006603:Hexuronic Acids / Q000378:metabolism; D008826:Microbial Sensitivity Tests; D008969:Molecular Sequence Data; D009154:Mutation; D010447:Peptide Hydrolases / Q000235:genetics / Q000378:metabolism; D011550:Pseudomonas aeruginosa / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D012808:Sigma Factor / Q000235:genetics* / Q000378:metabolism; D001618:beta-Lactamases / Q000235:genetics / Q000378:metabolism
chemical_list: C079619:AlgU protein, Pseudomonas aeruginosa; D000464:Alginates; D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D004251:DNA Transposable Elements; D006603:Hexuronic Acids; C107895:MucA protein, Pseudomonas; D012808:Sigma Factor; D020723:Glucuronic Acid; D010447:Peptide Hydrolases; D001618:beta-Lactamases
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