Involvement Of Quorum-sensing In Biosynthesis Of Polyhydroxyalkanoates In Pseudomonas Aeruginosa

Quorum-sensing is a regulatory mechanism with which bacteria regulate the gene expression according to the density of their population. Pseudomonas aeruginosa regulates expression of multiple genes via a hierarchical quorum-sensing cascade through LasR and RhlR and their cognate signal molecules N-(3-oxododecanoyl)-L-homoserine lactone (3O-C12-HSL) and N-(butanoyl)-L-homoserine lactone (C4-HSL).Under nutrient-limited conditions and in the presence of excess of carbon sources, Polyhydroxyalkanoates (PHAs) are synthesized by a class of bacteria as intracellular carbon and energy storage biopolymers. Sun et al. found that the signaling molecules receptor LuxR is necessary for PHB synthesis in Vibrio harveyi, this implies that QS participated in PHA synthesis in intracellular bacterial. Some studies analyzed QS can only indirectly involved in the regulation of PHA. Thus, according to the relevant literature, P. aeruginosa PAO1 are used to research the relationship between quorum-sensing and the PHA synthesis .It aims to explore the regulation of QS on biosynthesis of polyhydroxyalkanoates in P.aeruginosa. Wild-type P. aeruginosa PAO1 and its QS mutants are used to research the effects of quorum-sensing on biosynthesis of PHA by GC and real-time PCR at physiological and molecular level. A reporting vecter is constructed by fusining the promoter region of phaC1 gene with a promoterless lacZ gene on plasmid pRK970Km. The resulting plasmid is introduced into QS mutants and the corresponding effects on phaC1 transcription are investigated. At the same time in order to further confirm the biological synthesis of PHA is caused by the QS system, the shuttle vector contain the lasI/lasR genes were introduced into QS mutant PAO55, PAO56. It has been found that 1) The content of PHA in C4-HSL synthase gene rhlI mutant strain PAO210 is comparable no significant difference with that of the wild type. However, the PHA contents were significantly affected in 3OC12-HSL synthase gene lasI mutant strain PAO55, 3OC12-HSL transcriptional regulator gene lasR mutant strain PAO56 and lasI / lasR double mutant strain PAO57. 2) PHA synthase gene phaC1 expression exhibited a significant reduction in lasI mutant and lasR mutant strains. 3OC12-HSL signaling molecules complementary experiment shows that the expression of phaC1 can be recovered to the level of the wild type, but the synthesis of PHA is only partially restored in lasI mutant strain. 3)β-Galactosidase expression from a translational pphaC1-lacZ fusion was determined in PAO2, PAO552. Theβ-galactosidase activity of PAO552 was significantly lower than the wild type, It shows that the level of phaC1 transcription reduce significantly comparing with wild-type. The result indicate that the level of phaC1 transcription is positively regulated by lasI gene from the QS systerm. 4)The complemented strain containing the plasmid-bearing lasI/lasR gene could restore partially of the synthesis of PHA.Our study suggests that QS system might be involved in the regulation of intracellular PHA biosynthesis in P. aeruginosa PAO1.