Genetic analysis of an important oxidative stress locus in the anaerobe Bacteroides fragilis.
The obligate anaerobe, Bacteroides fragilis, is a highly aerotolerant intestinal tract organism that has evolved a complex oxidative stress response (OSR). The redox regulator OxyR controls several OSR genes (katB, dps, and ahpC), but there is little else known about other genes it regulates. To identify additional genes in the OxyR regulon, two-dimensional gel electrophoresis was used to isolate proteins from a mutant that constitutively expresses genes in the regulon. The 28,500 Da protein thioredoxin peroxidase (Tpx) was identified. Two additional genes induced during oxidative stress were identified adjacent to tpx, a putative RNA-binding protein (rbpA) and a cytochrome-c peroxidase (ccp). Transcriptional analyses showed that tpx and rbpA were transcribed as monocistronic mRNA species or as a bicistronic operon. Transcription of tpx was induced by exposure to air or H(2)O(2) from an OxyR-dependent promoter and to a lesser extent from a second OxyR-independent promoter. Expression of the rbpA gene during oxidative stress was regulated by the OxyR-dependent tpx promoter resulting in the bicistronic tpx/rbp mRNA. The ccp gene was expressed only as a monocistronic message and induction was only observed after exposure to H(2)O(2) in an OxyR-independent manner. Disruption of the tpx operon or ccp resulted in sensitivity to the organic peroxides cumene hydroperoxide (CHP) and t-butyl hydroperoxide (TBHP) but not to H(2)O(2). This work brings the total of oxyR-controlled genes in B. fragilis to five and suggests the existence of a second peroxide response regulator that controls ccp expression.
Herren
Christopher D
CD
Department of Microbiology and Immunology, The Brody School of Medicine, East Carolina University, 600 Moye Boulevard, Greenville, NC 27858-4354, USA.
Rocha
Edson R
ER
Smith
C Jeffrey
CJ
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D001441:Bacteroides fragilis / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D015152:Blotting, Northern; D003578:Cytochrome-c Peroxidase / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins; D015180:Electrophoresis, Gel, Two-Dimensional; D015964:Gene Expression Regulation, Bacterial; D008969:Molecular Sequence Data; D009154:Mutation; D009363:Neoplasm Proteins; D009876:Operon / Q000235:genetics; D018384:Oxidative Stress / Q000235:genetics*; D010544:Peroxidases / Q000235:genetics / Q000378:metabolism; D054464:Peroxiredoxins; D011401:Promoter Regions, Genetic / Q000235:genetics; D012329:RNA, Bacterial / Q000235:genetics / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D018085:Regulon / Q000235:genetics; D012097:Repressor Proteins / Q000235:genetics; D014157:Transcription Factors / Q000235:genetics; D024363:Transcription Initiation Site; D014158:Transcription, Genetic
D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; D009363:Neoplasm Proteins; D012329:RNA, Bacterial; D016601:RNA-Binding Proteins; D012097:Repressor Proteins; D014157:Transcription Factors; D010544:Peroxidases; D054464:Peroxiredoxins; D003578:Cytochrome-c Peroxidase