Activation of multiple antibiotic resistance and binding of stress-inducible promoters by Escherichia coli Rob protein.
Multiple antibiotic resistance in Escherichia coli can be mediated by induction of the SoxS or MarA protein, triggered by oxygen radicals (in the soxRS regulon) or certain antibiotics (in the marRAB regulon), respectively. These small proteins (SoxS, 107 residues; MarA, 127 residues) are homologous to the C terminus of the XylS-AraC family of proteins and are more closely related to a approximately 100-residue segment in the N terminus of Rob protein, which binds the right arm of the replication origin, oriC. We investigated whether the SoxS-MarA homology in Rob might extend to the regulation of some of the same inducible genes. Overexpression of Rob indeed conferred multiple antibiotic resistance similar to that known for SoxS and MarA (against chloramphenicol, tetracycline, nalidixic acid, and puromycin), as well as resistance to the superoxide-generating compound phenazine methosulfate. The Rob-induced antibiotic resistance depended only partially on the micF antisense RNA that down-regulates the OmpF outer membrane porin to limit antibiotic uptake. Similar antibiotic resistance was conferred by expression of a Rob fragment containing only the N-terminal 123 residues that constitute the SoxS-MarA homology. Both intact Rob and the N-terminal fragment activated expression of stress genes (inaA, fumC, sodA) but with a pattern distinct from that found for SoxS and MarA. Purified Rob protein bound a DNA fragment containing the micF promoter (50% bound at approximately 10(-9) M Rob) as strongly as it did oriC, and it bound more weakly to DNA containing the sodA, nfo, or zwf promoter (50% bound at 10(-8) to 10(-7) M). Rob formed multiple DNA-protein complexes with these fragments, as seen previously for SoxS. These data point to a DNA-binding gene activator module used in different protein contexts.
Ariza
R R
RR
Department of Molecular and Cellular Toxicology, Harvard School of Public Health, Boston, Massachusetts 02115.
Li
Z
Z
Ringstad
N
N
Demple
B
B
D001426:Bacterial Proteins / Q000235:genetics / Q000502:physiology*; D001483:Base Sequence; D004268:DNA-Binding Proteins / Q000235:genetics / Q000502:physiology*; D004352:Drug Resistance, Microbial; D004926:Escherichia coli / Q000187:drug effects* / Q000235:genetics; D029968:Escherichia coli Proteins; D015964:Gene Expression Regulation, Bacterial; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D010446:Peptide Fragments / Q000502:physiology; D011401:Promoter Regions, Genetic
D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; D010446:Peptide Fragments; C080267:Rob protein, E coli