Identification and characterization of a two-component sensor-kinase and response-regulator system (DcuS-DcuR) controlling gene expression in response to C4-dicarboxylates in Escherichia coli.
The dcuB gene of Escherichia coli encodes an anaerobic C4-dicarboxylate transporter that is induced anaerobically by FNR, activated by the cyclic AMP receptor protein, and repressed in the presence of nitrate by NarL. In addition, dcuB expression is strongly induced by C4-dicarboxylates, suggesting the presence of a novel C4-dicarboxylate-responsive regulator in E. coli. This paper describes the isolation of a Tn10 mutant in which the 160-fold induction of dcuB expression by C4-dicarboxylates is absent. The corresponding Tn10 mutation resides in the yjdH gene, which is adjacent to the yjdG gene and close to the dcuB gene at approximately 93.5 min in the E. coli chromosome. The yjdHG genes (redesignated dcuSR) appear to constitute an operon encoding a two-component sensor-regulator system (DcuS-DcuR). A plasmid carrying the dcuSR operon restored the C4-dicarboxylate inducibility of dcuB expression in the dcuS mutant to levels exceeding those of the dcuS+ strain by approximately 1.8-fold. The dcuS mutation affected the expression of other genes with roles in C4-dicarboxylate transport or metabolism. Expression of the fumarate reductase (frdABCD) operon and the aerobic C4-dicarboxylate transporter (dctA) gene were induced 22- and 4-fold, respectively, by the DcuS-DcuR system in the presence of C4-dicarboxylates. Surprisingly, anaerobic fumarate respiratory growth of the dcuS mutant was normal. However, under aerobic conditions with C4-dicarboxylates as sole carbon sources, the mutant exhibited a growth defect resembling that of a dctA mutant. Studies employing a dcuA dcuB dcuC triple mutant unable to transport C4-dicarboxylates anaerobically revealed that C4-dicarboxylate transport is not required for C4-dicarboxylate-responsive gene regulation. This suggests that the DcuS-DcuR system responds to external substrates. Accordingly, topology studies using 14 DcuS-BlaM fusions showed that DcuS contains two putative transmembrane helices flanking a approximately 140-residue N-terminal domain apparently located in the periplasm. This topology strongly suggests that the periplasmic loop of DcuS serves as a C4-dicarboxylate sensor. The cytosolic region of DcuS (residues 203 to 543) contains two domains: a central PAS domain possibly acting as a second sensory domain and a C-terminal transmitter domain. Database searches showed that DcuS and DcuR are closely related to a subgroup of two-component sensor-regulators that includes the citrate-responsive CitA-CitB system of Klebsiella pneumoniae. DcuS is not closely related to the C4-dicarboxylate-sensing DctS or DctB protein of Rhodobacter capsulatus or rhizobial species, respectively. Although all three proteins have similar topologies and functions, and all are members of the two-component sensor-kinase family, their periplasmic domains appear to have evolved independently.
Golby
P
P
The School of Animal and Microbial Sciences, University of Reading, Whiteknights, Reading RG6 6AJ, United Kingdom.
Davies
S
S
Kelly
D J
DJ
Guest
J R
JR
Andrews
S C
SC
D000595:Amino Acid Sequence; D000693:Anaerobiosis; D001426:Bacterial Proteins; D001692:Biological Transport; D002352:Carrier Proteins / Q000235:genetics* / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics*; D029382:Dicarboxylic Acid Transporters; D003998:Dicarboxylic Acids / Q000494:pharmacology*; D004926:Escherichia coli / Q000187:drug effects / Q000235:genetics*; D029968:Escherichia coli Proteins; D005650:Fumarates / Q000494:pharmacology; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D005809:Genes, Regulator; D005816:Genetic Complementation Test; D008565:Membrane Proteins / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D015183:Restriction Mapping; D017386:Sequence Homology, Amino Acid; D013385:Succinate Dehydrogenase / Q000235:genetics / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics*