Temperature sensing in Yersinia pestis: regulation of yopE transcription by lcrF.
In Escherichia coli, a yopE::lacZ fusion was found to be regulated by temperature in the presence of the cloned BamHI G fragment of Yersinia pestis plasmid pCD1, which contains the lcrF locus. Increasing the copy number of lcrF relative to that of the yopE reporter had a negligible effect on the induction ratio (26 versus 37 degrees C) but caused large reductions in the absolute levels of yopE transcription. We localized the lcrF gene by monitoring the induction phenotype of BamHI G deletion derivatives. Sequencing revealed an open reading frame capable of encoding a protein of 30.8 kDa. A protein product of this size was detected in a T7 expression system, and LcrF-dependent yopE-specific DNA binding activity was observed. As expected, LcrF exhibited 98% homology to VirF of Yersinia enterocolitica and significant homology to the carboxy termini of other members of the AraC family of transcriptional regulatory proteins. These proteins could be divided into two classes according to function: those regulating operons involved in catabolism of carbon and energy sources and those involved in regulating virulence genes. lcrF::lacZ transcriptional fusions were constructed and analyzed in Y. pestis and E. coli. The activity of the fusions was not affected by the native pCD1 virulence plasmid, an intact lcrF gene, or temperature. Thus, induction of lcrF transcription is not essential for temperature-dependent activation of yopE transcription. A portion of LcrF was found associated with the membrane fraction in E. coli; however, pulse-chase experiments indicated that this result is an artifact of fractionation.
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Department of Molecular Genetics and Microbiology, University of Massachusetts Medical Center, Worcester 01655.
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Goguen
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D000595:Amino Acid Sequence; D001419:Bacteria / Q000235:genetics; D001425:Bacterial Outer Membrane Proteins / Q000235:genetics* / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D002872:Chromosome Deletion; D003001:Cloning, Molecular / Q000379:methods; D004269:DNA, Bacterial / Q000235:genetics / Q000302:isolation & purification; D004268:DNA-Binding Proteins; D004926:Escherichia coli / Q000235:genetics / Q000502:physiology; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D005838:Genotype; D016350:Leucine Zippers / Q000235:genetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D016366:Open Reading Frames; D010957:Plasmids; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D015183:Restriction Mapping; D012689:Sequence Homology, Nucleic Acid; D013696:Temperature; D015534:Trans-Activators; D014158:Transcription, Genetic; D015010:Yersinia pestis / Q000235:genetics* / Q000502:physiology*; D001616:beta-Galactosidase / Q000235:genetics / Q000378:metabolism
D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; D011993:Recombinant Fusion Proteins; D015534:Trans-Activators; C075515:lcrF protein, Yersinia pestis; C063239:yopE protein, Yersinia; D001616:beta-Galactosidase