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The stringent response promotes biofilm dispersal in Pseudomonas putida
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2017 (English)In: Scientific Reports, ISSN 2045-2322, E-ISSN 2045-2322, Vol. 7, article id 18055Article in journal (Refereed) Published
Abstract [en]

Biofilm dispersal is a genetically programmed response enabling bacterial cells to exit the biofilm in response to particular physiological or environmental conditions. In Pseudomonas putida biofilms, nutrient starvation triggers c-di-GMP hydrolysis by phosphodiesterase BifA, releasing inhibition of protease LapG by the c-di-GMP effector protein LapD, and resulting in proteolysis of the adhesin LapA and the subsequent release of biofilm cells. Here we demonstrate that the stringent response, a ubiquitous bacterial stress response, is accountable for relaying the nutrient stress signal to the biofilm dispersal machinery. Mutants lacking elements of the stringent response – (p)ppGpp sythetases [RelA and SpoT] and/or DksA – were defective in biofilm dispersal. Ectopic (p)ppGpp synthesis restored biofilm dispersal in a ∆relA ∆spoT mutant. In vivo gene expression analysis showed that (p)ppGpp positively regulates transcription of bifA, and negatively regulates transcription of lapA and the lapBC, and lapE operons, encoding a LapA-specific secretion system. Further in vivo and in vitro characterization revealed that the PbifA promoter is dependent on the flagellar σ factor FliA, and positively regulated by ppGpp and DksA. Our results indicate that the stringent response stimulates biofilm dispersal under nutrient limitation by coordinately promoting LapA proteolysis and preventing de novo LapA synthesis and secretion.

Place, publisher, year, edition, pages
Nature Publishing Group, 2017. Vol. 7, article id 18055
National Category
Biochemistry and Molecular Biology
Identifiers
URN: urn:nbn:se:umu:diva-141910DOI: 10.1038/s41598-017-18518-0ISI: 000418644100014PubMedID: 29273811OAI: oai:DiVA.org:umu-141910DiVA, id: diva2:1157222
Note

Originally published in thesis in manuscript form.

Available from: 2017-11-15 Created: 2017-11-15 Last updated: 2018-01-15Bibliographically approved
In thesis
1. Global regulatory factors that impact metabolic and lifestyle choices in Pseudomonas putida
Open this publication in new window or tab >>Global regulatory factors that impact metabolic and lifestyle choices in Pseudomonas putida
2017 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Pseudomonas putida strains have a broad metabolic capacity and are innately resistant to many harmful substances – properties that make them of interest for a number of industrial and biotechnological application. They can rapidly adapt to changes in physico-chemical parameters in the soil and water environments they naturally inhabit. Like other bacteria, they have evolved both specific and cross-acting global regulatory circuits to control endurance traits and life style choices in order to survive. Three such survival tactics are 1) the ability to control flagella-mediated motility to search for metabolically favourable locations, 2) to produce protective biofilm structures to resist environmental insults, and 3) to distinguish the energetically most favourable carbon source amongst an array on offer. These processes are often co-ordinated regulated by intersecting networks that are controlled by global signalling molecules (second messengers) such as the nucleotides ppGpp and c-di-GMP, and globally acting proteins.

In the first part of my thesis I present evidence that the PP4397 protein of P. putida is responsible for slowing down flagella-driven motility in response to c-di-GMP signalling from a dual-functional c-di-GMP turnover protein termed PP2258. This connection is expanded upon to present a potential signal transduction pathway from a surface located receptor to PP2258 and the c-di-GMP responsive PP4397 protein, and from there to the flagella motors to determine flagella performance. The transcriptional regulatory studies that accompany this work suggest a means by which transcriptional control may serve to initiate a co-ordinated blocking of de novo flagella biogenesis and slowing-down flagella rotation – two processes needed to enter the biofilm mode of growth. 

Exiting from a biofilm matrix is also a c-di-GMP elicited behaviour, prompted when nutrients become scarce. In my second piece of work I present evidence that hunger-signals in the form of ppGpp directly control transcription to elevate the levels of a c-di-GMP degrading protein – BifA – which lies at the heart of programed biofilm dispersal. 

The final part of my thesis, concerns how the global regulatory proteins Hfq and Crc act at multiple levels to subvert catabolism of phenolics to favour other preferred sources of carbon. Evidence is presented that this involves a two-tiered translational repression – one at the level of the master regulator of the system, and another at the level of the catabolic enzymes. This study also revealed a hitherto unsuspected role of Crc in maintenance of an IncP-2 plasmid within a bacterial population. This latter finding has implications for a wide variety of processes encoded by the IncP-2 group of Pseudomonas-specific mega-plasmids.

Place, publisher, year, edition, pages
Umeå: Umeå Universitet, 2017. p. 62
Keyword
Pseudomonas putida, c-di-GMP, motility, ppGpp, DksA, biofilm dispersal, transcriptional and translational regulation, dmp-system, phenol catabolism, carbon repression control, plasmid stability
National Category
Biological Sciences Microbiology
Identifiers
urn:nbn:se:umu:diva-141912 (URN)978-91-7601-801-9 (ISBN)
Public defence
2017-12-08, A103 (Astrid Fagraeus), byggnad 6A, Norrlands Universitetssjukhus, Umeå, 09:00 (English)
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Available from: 2017-11-17 Created: 2017-11-15 Last updated: 2017-11-21Bibliographically approved

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