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Journal of Bacteriology, January 2006, p. 103-114, Vol. 188, No. 1
0021-9193/06/$08.00+0     doi:10.1128/JB.188.1.103-114.2006
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

Genetic Transplantation: Salmonella enterica Serovar Typhimurium as a Host To Study Sigma Factor and Anti-Sigma Factor Interactions in Genetically Intractable Systems

Joyce E. Karlinsey* and Kelly T. Hughes{dagger}

Department of Microbiology, University of Washington, Seattle, Washington 98195

Received 18 July 2005/ Accepted 28 September 2005

In Salmonella enterica serovar Typhimurium, {sigma}28 and anti-sigma factor FlgM are regulatory proteins crucial for flagellar biogenesis and motility. In this study, we used S. enterica serovar Typhimurium as an in vivo heterologous system to study {sigma}28 and anti-{sigma}28 interactions in organisms where genetic manipulation poses a significant challenge due to special growth requirements. The chromosomal copy of the S. enterica serovar Typhimurium {sigma}28 structural gene fliA was exchanged with homologs of Aquifex aeolicus (an extreme thermophile) and Chlamydia trachomatis (an obligate intracellular pathogen) by targeted replacement of a tetRA element in the fliA gene location using {lambda}-Red-mediated recombination. The S. enterica serovar Typhimurium hybrid strains showed {sigma}28-dependent gene expression, suggesting that {sigma}28 activities from diverse species are preserved in the heterologous host system. A. aeolicus mutants defective for {sigma}28/FlgM interactions were also isolated in S. enterica serovar Typhimurium. These studies highlight a general strategy for analysis of protein function in species that are otherwise genetically intractable and a straightforward method of chromosome restructuring using {lambda}-Red-mediated recombination.

* Corresponding author. Mailing address: Department of Microbiology, University of Washington, Seattle, WA 98195. Phone: (206) 616-9925. Fax: (206) 543-8297. E-mail: joya{at}u.washington.edu.

{dagger} Present address: Department of Biology, University of Utah, Salt Lake City, UT 84112.

Journal of Bacteriology, January 2006, p. 103-114, Vol. 188, No. 1
0021-9193/06/$08.00+0     doi:10.1128/JB.188.1.103-114.2006
Copyright © 2006, American Society for Microbiology. All Rights Reserved.



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