Modulation of bft expression by the Bacteroides fragilis pathogenicity island and its flanking region.
To establish a recombinant system for high-level expression of biologically active Bacteroides fragilis toxin (BFT), we studied the expression of bft in non-toxigenic B. fragilis (NTBF) strains. The bft gene and the B. fragilis pathogenicity island (BfPAI) were cloned into NTBF strains with two distinct genetic patterns: (i) pattern II, strains lacking the BfPAI and its flanking region; and (ii) pattern III, strains lacking the BfPAI but containing its flanking region. Analysis of BFT activity of these recombinant strains on HT29/C1 cells showed that both the BfPAI and its flanking regions are important to optimal BFT activity. Reverse transcription polymerase chain reaction (RT-PCR) analysis indicated that the BfPAI and its flanking regions modulate bft expression. Further experiments demonstrated that the approximately 700 bp region upstream of bft is the BfPAI region critical for optimal bft expression. We conclude that both the region flanking the BfPAI and approximately 700 bp region upstream of bft are crucial to maximal BFT production by ETBF strains.
Division of Infectious Diseases, Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.
Augusto A
AA
Franco
Rodney K
RK
Cheng
Alan
A
Goodman
Cynthia L
CL
Sears
D001441:Bacteroides fragilis / Q000472:pathogenicity*; D001483:Base Sequence; D015153:Blotting, Western; D004269:DNA, Bacterial; D004591:Electrophoresis, Polyacrylamide Gel; D015964:Gene Expression Regulation, Bacterial / Q000235:genetics*; D005798:Genes, Bacterial; D018398:Homeodomain Proteins / Q000235:genetics*; D008969:Molecular Sequence Data; D051761:Paired Box Transcription Factors; D011401:Promoter Regions, Genetic; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014157:Transcription Factors / Q000235:genetics*; D014774:Virulence / Q000235:genetics*
D004269:DNA, Bacterial; D018398:Homeodomain Proteins; D051761:Paired Box Transcription Factors; D014157:Transcription Factors; C101485:homeobox protein PITX1