Regulation of Expression of the Nonhemolytic Phospholipase C of Burkholderia cepacia

Abstract

Burkholderia cepacia is an opportunistic pathogen that causes serious pulmonary infections in cystic fibrosis patients. We have purified and partially characterized one potential virulence factor for the organism-a nonhemolytic phospholipase C-and we studied the effect of iron restriction and choline and phosphate concentrations on the expression of phospholipase C. Iron limitation did not affect expression, the effect of choline was variable, and high phosphate concentrations repressed expression. Experiments with heat-treated spent culture supernatants suggested that autoinducers affected the expression of the phospholipase and two other potential virulence factors, a protease and a lipase. We screened 26 B. cepacia isolates for autoinducer activity: 11 induced violacein production in the autoinducer-deficient mutant Chromobacterium violaceum CV026. Spent supernatants from two strains, one that was positive in the C. violaceum assay and one that was negative, were tested for inducing early expression of phospholipase C, protease, and lipase in homologous and heterologous cultures. Expression of all three enzymes was increased or induced at an earlier stage in the growth curve in every case, suggesting not only that autoinducers were involved in the regulation of the expression of these enzymes, but also that the autoinducers were of two different classes.

Legionella pneumophila genes that encode lipase and phospholipase C activities a aThe GenBank accession numbers for the L. pneumophila lipA, lipB and plcA sequences are AF454863, AF454864 and AF454865, respectively.

“…First, bacterial lipases have been implicated in a variety of pathogenic processes, including the inhibition of phagocyte function (Gribbon et al, 1993 ;Jaeger et al, 1994 ;Konig et al, 1996 ;Pratt et al, 2000 ; Rollof et al, 1988). Second, pNPPC hydrolysis has been associated with the production of phospholipase C (PLC) enzymes in L. pneumophila and others, and in some of those other bacteria, the PLC is implicated in virulence (Baine, 1988 ;Dowling et al, 1992 ;Filloux et al, 1987 ;Jepson et al, 1999 ; Merino et al, 1999 ;Schmiel & Miller, 1999 ;Songer, 1997 ;Strom et al, 1991 ;Swanson & Hammer, 2000 ;Terada et al, 1999 ;Titball, 1998 ;Weingart & Hooke, 1999). Third, lipase-and PLC-like activities are conserved among clinical isolates of L. pneumophila as well as other pathogenic Legionella species (Baine, 1985 ;Muller, 1981 ;Nolte et al, 1982 ;Thorpe & Miller, 1981).…”

Type II Secretion and Pathogenesis

“…In a B. cepacia cepR mutant, no extracellular protease can be detected, but it is not yet known if CepR regulates protease production and/or biogenesis of the type II secretion apparatus (80). Another recent report suggests that the expression of lipase and phospholipase C is also under quorum-sensing control in B. cepacia (156). In addition to intraspecies regulation, there is also apparent cross talk and interspecies signaling between P. aeruginosa and B. cepacia (97).…”

Legionella pneumophila Mip, a Surface-Exposed Peptidylproline cis-trans -Isomerase, Promotes the Presence of Phospholipase C-Like Activity in Culture Supernatants

“…In many bacteria, p-NPPC hydrolysis is attributed to phospholipase C (PLC) enzymes, many of which promote virulence (46,54,83,86). Early reports described p-NPPC hydrolase activities for various Legionella species (6,7,56,57,84).…”