Quorum Sensing in Burkholderia cepacia : Identification of the LuxRI Homologs CepRI

Abstract

Burkholderia cepacia has emerged as an important pathogen in patients with cystic fibrosis. Many gram-negative pathogens regulate the production of extracellular virulence factors by a cell density-dependent mechanism termed quorum sensing, which involves production of diffusible N -acylated homoserine lactone signal molecules, called autoinducers. Transposon insertion mutants of B. cepacia K56-2 which hyperproduced siderophores on chrome azurol S agar were identified. One mutant, K56-R2, contained an insertion in a luxR homolog that was designated cepR . The flanking DNA region was used to clone the wild-type copy of cepR . Sequence analysis revealed the presence of cepI , a luxI homolog, located 727 bp upstream and divergently transcribed from cepR . A lux box-like sequence was identified upstream of cepI . CepR was 36% identical to Pseudomonas aeruginosa RhlR and 67% identical to SolR of Ralstonia solanacearum . CepI was 38% identical to RhlI and 64% identical to SolI. K56-R2 demonstrated a 67% increase in the production of the siderophore ornibactin, was protease negative on dialyzed brain heart infusion milk agar, and produced 45% less lipase activity in comparison to the parental strain. Complementation of a cepR mutation restored parental levels of ornibactin and protease but not lipase. An N -acylhomoserine lactone was purified from culture fluids and identified as N -octanoylhomoserine lactone. K56-I2, a cepI mutant, was created and shown not to produce N -octanoylhomoserine lactone. K56-I2 hyperproduced ornibactin and did not produce protease. These data suggest both a positive and negative role for cepIR in the regulation of extracellular virulence factor production by B. cepacia.