Roles of extracitoplasmic function (ECF) sigma factors SigX and PA14_21550 in Pseudomonas aeruginosa

Abstract

The gama-proteobacterium Pseudomonas aeruginosa presents a high number of regulatory systems, therefore it is able to colonize different environments and hosts, including immunocompromised humans. It is difficult to eradicate because of its high intrinsic resistance to antibiotics and capacity of growing in biofilm style. This bacterium can direct gene expression through alternative sigma factors, among them about 20 ECF sigma factors, important in detecting and responding to environmental signals. In this work, we intend to understand the function of the ECF sigma factor SigX in the physiology of P. aeruginosa, as well as its relationship with two other sigma factors: the putative ECF sigma PA14_21550, one of the SigX targets, and RpoS, the stationary phase sigma factor that apparently controls sigX expression, characterizing a sigma factors regulation cascade. SigX overexpressing cultures present biphasic growth curve and induction of fatty acids biosynthesis enzymes, accompanied by an increase in membrane fluidity and improved resistance to cold shock and high temperature sensitivity. High levels of SigX overexpression are regulated along the growth curve for one or more regulators that remain unknown. This project also aims at the characterization of this(these) regulator(s), probably proteases, from transposon mutant library PA14NR . The physiological roles of SigX and PA14_21550 will be investigated after construction of mutants for the genes that encode them and assays in various environmental and stress conditions. The regulons of these ECF sigma factors will be determined by transcriptomic analysis via RNA sequencing. This technique will be standardized for use with P. aeruginosa in our laboratory and will constitute a new and efficient tool for gene expression analysis that can be used in several other projects of our group, generating a robust amount of knowledge about gene regulation of this important and versatile opportunistic pathogen.