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Expression analysis of the AdnA regulon in Pseudomonas fluorescens Pf0-1

ABSTRACT: Investigation of whole genome gene expression AdnA in Pseudomonas fluorescens, an ortholog of FleQ in P. aeruginosa, regulates both motility and flagella-mediated attachment to various surfaces. A whole genome microarray determined the AdnA transcriptome by comparing the gene expression pattern of wild-type Pf0-1 to that of Pf0-2x (adnA-) in broth culture. In the absence of AdnA, expression of 92 genes was decreased, while 11 genes showed increased expression. Analysis of 16 of these genes fused to lacZ confirmed the microarray results. Several genes were further evaluated for their role in motility and biofilm formation. Two genes, Pfl01_1508 and Pfl01_1517, affected motility and had different effects on biofilm formation in Pf0-1. These two genes are predicted to encode proteins similar to the glycosyl transferases FgtA1 and FgtA2, which have been shown to be involved in virulence and motility in P. syringae. Three other genes, Pfl01_1516, Pfl01_1572, and Pfl01_1573, not previously associated with motility and biofilm formation in Pseudomonas had similar affects on biofilm formation in Pf0-1. Deletion of each of these genes led to different motility defects. Our data revealed an additional level of complexity in the control of flagella function beyond the core genes known to be required, and may yield insights into processes important for environmental persistence of P. fluorescens Pf0-1.

ORGANISM(S): Pseudomonas fluorescens Pseudomonas fluorescens Pf0-1

PROVIDER: GSE33865 | GEO | 2011/11/30

SECONDARY ACCESSION(S): PRJNA148155

REPOSITORIES: GEO

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Project description:Pseudomonas fluorescens is implicated in food spoilage especially under cold storage, causing the increase of food waste and the reduction of environmental (and nutritional) sustainability. In food industry, Pseudomonas biofilm on pipe surfaces and facilities creates a persistent source of contamination and increases resistance to common disinfection strategies. Even though gene expression and protein pattern changes under environmental stimuli have been suggested, cellular mechanisms involved in biofilm production are poorly investigated for this species. In the present work, the cheese blue pigmenting P. fluorescens ITEM 17298 produced increased biofilm biomass and motility at 15 °C in comparison to 30 °C. Proteomic analysis revealed major differences of proteins involved in biofilm formation. In particular, at 15 °C we found increased amount of the c-di-GMP regulator PleD involved in motility as well as AlgB and the carbon storage regulator directly involved biofilm formation. Carbon metabolism, protein and fatty acid synthesis were also stimulated under cold incubation. Moreover, for the first time, enzymes correlated to indigoidine synthesis (blue pigment) were identified in the same condition. The differences in expression of regulatory proteins could therefore be the major determinant of the biofilm formation ability of P. fluorescens under refrigerated condition. Genomic and proteomic results also revealed virulence factors mostly detected at 30 °C. In order to counteract P. fluorescens persistence, this study also proposed a control strategy for biofilm development. Thus, a sub-lethal concentration of bovine lactoferrin hydrolysate (HLF) was assayed at both temperatures. Treatment caused a significant reduction in biofilm biomass, also confirmed by changes in proteome pattern. At each condition proteins involved regulatory functions and transcription factors were downregulated after HLF-treatment. Among these, at 15 °C we found that PleD was absent in treated samples, whilst negative regulators of alginate biosynthesis were detected; likewise, indigoidine related proteins were repressed. The cyclic-di-GMP-binding biofilm dispersal mediator was instead detected in all treated samples.

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Expression analysis of the AdnA regulon in Pseudomonas fluorescens Pf0-1

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