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Regulation of expression of the Pasteurella haemolytica leukotoxin determinant.

Department of Microbiology, University of Guelph, Ontario, Canada.

ABSTRACT

The Pasteurella haemolytica leukotoxin determinant is composed of four contiguous genes encoded on the same DNA strand and denoted lktCABD, in the order of their genetic organization. To gain a better understanding of the expression and regulation of the leukotoxin, the transcripts and promoters of the lkt determinant were mapped. Northern (RNA) blot analysis revealed two sets of transcripts. One set was 3.7 and 3.4 kilobases long, encoded lktCA, and comprised approximately 90% of the transcripts, whereas the other set was 7.4 and 7.1 kilobases long and encoded lktCABD. Two promoters were present, and each had features similar to the Escherichia coli consensus promoter sequences. Both promoters were located upstream from lktC; they were separated by 258 base pairs, as mapped by primer extension analysis. These results suggest a mechanism of expression similar to that of the related E. coli hemolysin. Transcription initiated upstream from lktC at either promoter and continued through lktC and lktA to a rho-independent transcriptional termination signal in the lktA-lktB intercistronic region. This signal attenuated expression by terminating 90% of transcription to generate the 3.7- and 3.4-kilobase lktCA transcripts. The remaining readthrough transcription generated full-length 7.4- and 7.1-kilobase lktCABD transcripts. Expression of the leukotoxin was greatly reduced by growth at 30 degrees C, pH 6.5, and Fe2+ limitation. These conditions also modulated the expression of a number of other secreted proteins, which suggests that all of these secreted proteins are controlled by the same regulatory mechanism.

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