RpmA is required for nonopsonic phagocytosis of Pseudomonas aeruginosa.
Pseudomonas aeruginosa causes severe respiratory tract infections in patients with cystic fibrosis (CF). We have been examining nonopsonic phagocytosis of P. aeruginosa by macrophages. To study the P. aeruginosa-macrophage interaction at the molecular level, we have constructed a transposon Tn5G bank in a clinical isolate of P. aeruginosa (strain 4020) and identified mutants resistant to nonopsonic phagocytosis. Phagocytosis-resistant mutants were enriched by passaging the transposon bank over 18 macrophage monolayers. Of 900 individual mutants isolated from this enriched pool in a nonopsonic phagocytosis assay, we identified 85 putative mutants that were resistant to phagocytosis. In this study, we have characterized one of these transposon mutants, P. aeruginosa 4020 H27A, which was poorly ingested. H27A possessed a Tn5G insertion in a gene encoding a protein with homology to the MotA proteins of several species of bacteria. We have called this gene rpmA for required for phagocytosis by macrophages. RpmA is one of two MotA paralogs in P. aeruginosa. This rpmA::Tn5G mutant was motile both on agar plates and in visual examination of wet mounts. The phagocytosis defect was partially complemented by providing the rpmA gene in trans and fully complemented when both rpmA and rpmB were provided. A rpmA null mutant was ingested by macrophages similar to the H27A transposon mutant. These data suggest that the rpmA and rpmB gene products are required for the efficient ingestion of P. aeruginosa by macrophages.
Simpson
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DA
Division of Infectious and Immunological Diseases, Department of Pediatrics, and the Canadian Bacterial Diseases Network, University of British Columbia, Vancouver, British Columbia, Canada V5Z 4H4. dsimpson@cbdn.ca
Speert
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DP
D001426:Bacterial Proteins / Q000235:genetics / Q000276:immunology*; D001483:Base Sequence; D002648:Child; D003550:Cystic Fibrosis / Q000382:microbiology; D004251:DNA Transposable Elements; D004269:DNA, Bacterial; D005816:Genetic Complementation Test; D006801:Humans; D008264:Macrophages / Q000276:immunology* / Q000382:microbiology; D008565:Membrane Proteins / Q000235:genetics / Q000276:immunology*; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D010587:Phagocytosis / Q000276:immunology*; D011550:Pseudomonas aeruginosa / Q000235:genetics / Q000276:immunology*
D001426:Bacterial Proteins; D004251:DNA Transposable Elements; D004269:DNA, Bacterial; D008565:Membrane Proteins; C409030:RpmA protein, Pseudomonas aeruginosa