Abstract
Pseudomonas lipases are well-studied, but few studies have examined the mechanisms of lipase expression regulation. As a global regulatory protein, PmrA controls the expression of multiple genes such as the Dot/Icm apparatus, eukaryotic-like proteins, and secreted effectors. In this study, the effect of PmrA on expression of the lipase lipA in Pseudomonas aeruginosa PAO1 was investigated by knocking out or overexpressing pmrA, rsmY, and rsmA. PmrA regulated the expression of lipA at both the transcriptional and translational level although translation was the pivotal regulatory mechanism for lipA expression. PmrA also regulated the expression of rsmY. Using gel mobility shift assay and pmrA/rsmY double gene knock-out model, we showed that PmrA directly bound to the promoter sequence of rsmY to regulate lipA expression. Translation of lipA was activated by the PmrA/PmrB system via RsmA. Specifically, the Shine-Dalgarno (SD) sequence located at lipA mRNA was overlapped through combination between RsmA and the AGAUGA sequence, subsequently blocking the 30S ribosomal subunit to the SD sequence, leading to translational inhibition of lipA. Transcriptional repression of RsmY initiated translation of lipA through negative translational regulation of rsmA. In conclusion, this study demonstrated that in P. aeruginosa PAO1, PmrA mainly regulated rsmY expression at a translational level to influence lipA expression. RsmY primarily activated lipA translation via negative translational regulation of rsmA.
Highlights
Lipases are ubiquitous enzymes produced by all animals, plants and microbes
Enzyme production is too limited to match the demands of industry because of low lipase expression in original strains
Analyzing associations between RNA-seq [log2 Ratio (PS2/PS1) = 8.8] and proteomic data [Quantition(PS2/PS1) = 2.5] showed that PmrA expression was significantly correlated with lipA expression in P. aeruginosa PAO1
Summary
Lipases (triacylglycerol acylhydrolases, EC 3.1.1.3) are ubiquitous enzymes produced by all animals, plants and microbes. The versatility and plasticity of lipases contribute to their extensive application in the production of fine chemicals (Busto et al, 2006; Chang et al, 2007), detergents (Romdhane et al, 2010; Grbavcicet al., 2011), food (Aravindan et al, 2007; Pan et al, 2012), and biodiesel (Yoo et al, 2011; Jin et al, 2013). Lipases from the genera Pseudomonas and Burkholderia have high tolerance to harsh industrial application environments (Tran et al, 2016; Dwivedee et al, 2017). Enzyme production is too limited to match the demands of industry because of low lipase expression in original strains
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