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Journal of Bacteriology, April 1999, p. 2217-2224, Vol. 181, No. 7
0021-9193/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

Genes Coding for Phosphotransacetylase and Acetate Kinase in Sinorhizobium meliloti Are in an Operon That Is Inducible by Phosphate Stress and Controlled by PhoB

Michael L. Summers,dagger Michael C. Denton, and Timothy R. McDermott*

Department of Land Resources and Environmental Sciences, Montana State University, Bozeman, Montana 59717

Received 27 October 1998/Accepted 27 January 1999

Recent work in this laboratory has shown that the gene coding for acetate kinase (ackA) in Sinorhizobium meliloti is up-regulated in response to phosphate limitation. Characterization of the region surrounding ackA revealed that it is adjacent to pta, which codes for phosphotransacetylase, and that these two genes are part of an operon composed of at least two additional genes in the following order: an open reading frame (orfA), pta, ackA, and the partial sequence of a gene with an inferred peptide that has a high degree of homology to enoyl-ACP reductase (fabI). Experiments combining enzyme assays, a chromosomal lacZ::ackA transcriptional fusion, complementation analysis with cosmid subclones, and the creation of mutations in pta and ackA all indicated that the orfA-pta-ackA-fabI genes are cotranscribed in response to phosphate starvation. Primer extension was used to map the position of the phosphate starvation-inducible transcriptional start sites upstream of orfA. The start sites were found to be preceded by a sequence having similarity to PHO boxes from other phosphate-regulated genes in S. meliloti and to the consensus PHO box in Escherichia coli. Introduction of a phoB mutation in the wild-type strain eliminated elevated levels of acetate kinase and phosphotransacetylase activities in response to phosphate limitation and also eliminated the phosphate stress-induced up-regulation of the ackA::lacZ fusion. Mutations in either ackA alone or both pta and ackA did not affect the nodulation or nitrogen fixation phenotype of S. meliloti.

* Corresponding author. Mailing address: Department of Land Resources and Environmental Sciences, Montana State University, Bozeman, MT 59717. Phone: (406) 994-2190. Fax: (406) 994-3933. E-mail: timmcder{at}gemini.oscs.montana.edu.

dagger Present address: Department of Biology, California State University---Northridge, Northridge, CA 91330-8303.

Journal of Bacteriology, April 1999, p. 2217-2224, Vol. 181, No. 7
0021-9193/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.


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