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Journal of Bacteriology, July 2006, p. 4749-4758, Vol. 188, No. 13
0021-9193/06/$08.00+0     doi:10.1128/JB.01937-05
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

A Variation of the Translation Attenuation Model Can Explain the Inducible Regulation of the pBC16 Tetracycline Resistance Gene in Bacillus subtilis

Patricia B. Lodato,{dagger} Elizabeth J. Rogers, and Paul S. Lovett*

Department of Biological Sciences, University of Maryland Baltimore County, Baltimore, Maryland

Received 18 December 2005/ Accepted 21 April 2006

Expression of the tet resistance gene from plasmid pBC16 is induced by the antibiotic tetracycline, and induction is independent of the native promoter for the gene. The nucleotide sequence at the 5' end of the tet mRNA (the leader region) is predicted to assume a complex secondary structure that sequesters the ribosome binding site for the tet gene. A spontaneous, constitutively expressed tet gene variant contains a mutation predicted to provide the tet gene with a nonsequestered ribosome binding site. Lastly, comparable levels of tet mRNA can be demonstrated in tetracycline-induced and uninduced cells. These results are consistent with the idea that the pBC16 tet gene is regulated by translation attenuation, a model originally proposed to explain the inducible regulation of the cat and erm genes in gram-positive bacteria. As with inducible cat and erm genes, the pBC16 tet gene is preceded by a translated leader open reading frame consisting of a consensus ribosome binding site and an ATG initiation codon, followed by 19 sense codons and a stop codon. Mutations that block translation of cat and erm leaders prevent gene expression. In contrast, we show that mutations that block translation of the tet leader result in constitutive expression. We provide evidence that translation of the tet leader peptide coding region blocks tet expression by preventing the formation of a secondary-structure complex that would, in the absence of leader translation, expose the tet ribosome binding site. Tetracycline is proposed to induce tet by blocking or slowing leader translation. The results indicate that tet regulation is a variation of the translation attenuation model.

* Corresponding author. Mailing address: Department of Biological Sciences, University of Maryland Baltimore County, Baltimore, MD 21250. Phone: (410) 455-2249. Fax: (410) 455-3574. E-mail: lovett{at}umbc.edu.

{dagger} Present address: Department of Microbiology and Immunology, University of Maryland School of Medicine, Baltimore, MD 21201.

Journal of Bacteriology, July 2006, p. 4749-4758, Vol. 188, No. 13
0021-9193/06/$08.00+0     doi:10.1128/JB.01937-05
Copyright © 2006, American Society for Microbiology. All Rights Reserved.





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