title: ClpXP proteases positively regulate alginate overexpression and mucoid conversion in Pseudomonas aeruginosa.
abstract: Overproduction of the exopolysaccharide alginate and conversion to a mucoid phenotype in Pseudomonas aeruginosa are markers for the onset of chronic lung infection in cystic fibrosis (CF). Alginate production is regulated by the extracytoplasmic function (ECF) sigma factor AlgU/T and the cognate anti-sigma factor MucA. Many clinical mucoid isolates carry loss-of-function mutations in mucA. These mutations, including the most common mucA22 allele, cause C-terminal truncations in MucA, indicating that an inability to regulate AlgU activity by MucA is associated with conversion to the mucoid phenotype. Here we report that a mutation in a stable mucoid strain derived from the parental strain PAO1, designated PAO581, that does not contain the mucA22 allele, was due to a single-base deletion in mucA (DeltaT180), generating another type of C-terminal truncation. A global mariner transposon screen in PAO581 for non-mucoid isolates led to the identification of three regulators of alginate production, clpP (PA1801), clpX (PA1802), and a clpP paralogue (PA3326, designated clpP2). The PAO581 null mutants of clpP, clpX and clpP2 showed decreased AlgU transcriptional activity and an accumulation of haemagglutinin (HA)-tagged N-terminal MucA protein with an apparent molecular mass of 15 kDa. The clpP and clpX mutants of a CF mucoid isolate revert to the non-mucoid phenotype. The ClpXP and ClpP2 proteins appear to be part of a proteolytic network that degrades the cytoplasmic portion of truncated MucA proteins to release the sequestered AlgU, which drives alginate biosynthesis.
authors.affiliation: Department of Biochemistry and Microbiology, Joan C. Edwards School of Medicine at Marshall University, Huntington, WV 25755-9320, USA.
authors.forename: Dongru
authors.initials: D
authors.lastname: Qiu
authors.affiliation: Department of Biochemistry and Microbiology, Joan C. Edwards School of Medicine at Marshall University, Huntington, WV 25755-9320, USA.
authors.forename: Vonya M
authors.initials: VM
authors.lastname: Eisinger
authors.affiliation: Department of Biochemistry and Microbiology, Joan C. Edwards School of Medicine at Marshall University, Huntington, WV 25755-9320, USA.
authors.forename: Nathan E
authors.initials: NE
authors.lastname: Head
authors.affiliation: Channing Laboratory, Department of Medicine, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02115, USA.
authors.forename: Gerald B
authors.initials: GB
authors.lastname: Pier
authors.affiliation: Department of Pediatrics, Joan C. Edwards School of Medicine at Marshall University, Huntington, WV 25701-3655, USA.|Department of Biochemistry and Microbiology, Joan C. Edwards School of Medicine at Marshall University, Huntington, WV 25755-9320, USA.
authors.forename: Hongwei D
authors.initials: HD
authors.lastname: Yu
mesh_terms: D000464:Alginates / Q000378:metabolism*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D003550:Cystic Fibrosis / Q000382:microbiology; D015964:Gene Expression Regulation, Bacterial; D020723:Glucuronic Acid / Q000378:metabolism; D006025:Glycosaminoglycans / Q000378:metabolism*; D006603:Hexuronic Acids / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D010447:Peptide Hydrolases / Q000235:genetics / Q000378:metabolism*; D010641:Phenotype; D017354:Point Mutation; D011552:Pseudomonas Infections / Q000382:microbiology*; D011550:Pseudomonas aeruginosa / Q000235:genetics / Q000378:metabolism*; D012808:Sigma Factor / Q000235:genetics / Q000378:metabolism
chemical_list: C079619:AlgU protein, Pseudomonas aeruginosa; D000464:Alginates; D001426:Bacterial Proteins; D006025:Glycosaminoglycans; D006603:Hexuronic Acids; C107895:MucA protein, Pseudomonas; D012808:Sigma Factor; C046129:mucoid exopolysaccharide (P aeruginosa); D020723:Glucuronic Acid; D010447:Peptide Hydrolases