Activation of the leu-500 promoter: a topological domain generated by divergent transcription in a plasmid.
The Salmonella typhimurium leu-500 promoter is active only in topA strains. In an earlier study (Chen, D., Bowater, R., Dorman, C., & Lilley, D. M. J. (1992) Proc. Natl. Acad. Sci. U.S.A. 89, 8784-8788), we showed that the activity of this promoter on a circular plasmid is a function of the transcription and translation of an adjacent tetA gene, and we suggested that the effect arises because of increased local negative superhelix density due to transcription (Liu, L. F., & Wang, J. C. (1987) Proc. Natl. Acad. Sci. U.S.A. 84, 7024-7027) initiated at the tetA promoter. In this study we show that translation of the 5' (N-terminal) section of tetA is required for activity of the leu-500 promoter, consistent with a requirement for membrane association of TetA. We have also shown the importance of a second transcription unit, the ampicillin resistance gene bla, in the activation of the leu-500 promoter. Thus the activity of the leu-500 promoter was reduced by partial deletion or premature termination of bla and was increased when the transcription of bla was boosted by the insertion of the stronger tac promoter. However, even in the latter situation the role of the tetA gene is dominant, and deletion of the tetA gene reduced activity of the leu-500 promoter to very low levels. These results suggest the existence of a topological domain defined by the divergent bla and tetA transcription units. Membrane insertion at tetA is essential to provide an anchorage point.(ABSTRACT TRUNCATED AT 250 WORDS)
Chen
D
D
Department of Biochemistry, The University, Dundee, U.K.
Bowater
R P
RP
Lilley
D M
DM
D017920:Antiporters; D001426:Bacterial Proteins / Q000235:genetics; D001483:Base Sequence; D004264:DNA Topoisomerases, Type I / Q000378:metabolism; D004270:DNA, Circular / Q000235:genetics; D015964:Gene Expression Regulation, Bacterial; D007930:Leucine; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D009838:Oligodeoxyribonucleotides / Q000737:chemistry; D009876:Operon; D010957:Plasmids; D011401:Promoter Regions, Genetic; D012486:Salmonella typhimurium / Q000235:genetics*; D013329:Structure-Activity Relationship; D014158:Transcription, Genetic; D001618:beta-Lactamases / Q000235:genetics
D017920:Antiporters; D001426:Bacterial Proteins; D004270:DNA, Circular; D009838:Oligodeoxyribonucleotides; C079051:tetA protein, Bacteria; D001618:beta-Lactamases; D004264:DNA Topoisomerases, Type I; D007930:Leucine