This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrowReprints and Permissions
Right arrow Copyright Information
Right arrow Books from ASM Press
Right arrow MicrobeWorld
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Lee, J. H.
Right arrow Articles by Holmes, R. K.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Lee, J. H.
Right arrow Articles by Holmes, R. K.

 Previous Article  |  Next Article 

Journal of Bacteriology, January 2000, p. 432-438, Vol. 182, No. 2
0021-9193/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

Characterization of Specific Nucleotide Substitutions in DtxR-Specific Operators of Corynebacterium diphtheriae That Dramatically Affect DtxR Binding, Operator Function, and Promoter Strength

John H. Leedagger and Randall K. Holmes*

Department of Microbiology, University of Colorado Health Sciences Center, Denver, Colorado 80262

Received 10 May 1999/Accepted 27 October 1999

The diphtheria toxin repressor (DtxR) of Corynebacterium diphtheriae uses Fe2+ as a corepressor. Holo-DtxR inhibits transcription from the iron-regulated promoters (IRPs) designated IRP1 through IRP5 as well as from the promoters for the tox and hmuO genes. DtxR binds to 19-bp operators with the consensus sequence 5'-TTAGGTTAGCCTAACCTAA-3', a perfect 9-bp palindrome interrupted by a single C · G base pair. Among the seven known DtxR-specific operators, IRP3 exhibits the weakest binding to DtxR. The message (sense) strand of the IRP3 operator (5'-TTAGGTGAGACGCACCCAT-3' [nonconsensus nucleotides underlined]) overlaps by 2 nucleotides at its 5' end with the putative -10 sequence of the IRP3 promoter. The underlined C at position +7 from the center of the IRP3 operator [C(+7)] is unique, because T is conserved at that position in other DtxR-specific operators. The present study examined the effects of nucleotide substitutions at position +7 or -7 in the IRP3 operator. In gel mobility shift assays, only the change of C(+7) to the consensus nucleotide T caused a dramatic increase in the binding of DtxR, whereas other nucleotide substitutions for C(+7) or replacements for A(-7) had only small positive or negative effects on DtxR binding. All substitutions for C(+7) or A(-7) except for A(-7)C dramatically decreased IRP3 promoter strength. In contrast, the A(-7)C variant caused increased promoter strength at the cost of nearly eliminating repressibility by DtxR. The message (sense) strand of the IRP1 operator (5'-TTAGGTTAGCCAAACCTTT-3') includes the -35 region of the IRP3 promoter. A T(+7)C variant of the IRP1 operator was also constructed, and it was shown to exhibit decreased binding to DtxR, decreased repressibility by DtxR, and increased promoter strength. The nucleotides at positions +7 and -7 in DtxR-specific operators are therefore important determinants of DtxR binding and repressibility of transcription by DtxR, and they also have significant effects on promoter activity for IRP3 and IRP1.

* Corresponding author. Mailing address: Department of Microbiology, Campus Box B-175, University of Colorado Health Sciences Center, 4200 East Ninth Ave., Denver, CO 80262. Phone: (303) 315-7903. Fax: (303) 315-6785. E-mail: Randall.Holmes{at}UCHSC.Edu.

dagger Present address: Chonbuk National University, College of Veterinary Medicine, Chonju, 561-756, South Korea.

Journal of Bacteriology, January 2000, p. 432-438, Vol. 182, No. 2
0021-9193/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.


This article has been cited by other articles:

  • Gartemann, K.-H., Abt, B., Bekel, T., Burger, A., Engemann, J., Flugel, M., Gaigalat, L., Goesmann, A., Grafen, I., Kalinowski, J., Kaup, O., Kirchner, O., Krause, L., Linke, B., McHardy, A., Meyer, F., Pohle, S., Ruckert, C., Schneiker, S., Zellermann, E.-M., Puhler, A., Eichenlaub, R., Kaiser, O., Bartels, D. (2008). The Genome Sequence of the Tomato-Pathogenic Actinomycete Clavibacter michiganensis subsp. michiganensis NCPPB382 Reveals a Large Island Involved in Pathogenicity. J. Bacteriol. 190: 2138-2149 [Abstract] [Full Text]  
  • Oram, D. M., Avdalovic, A., Holmes, R. K. (2002). Construction and Characterization of Transposon Insertion Mutations in Corynebacterium diphtheriae That Affect Expression of the Diphtheria Toxin Repressor (DtxR). J. Bacteriol. 184: 5723-5732 [Abstract] [Full Text]  
  • Howard, S. T., Byrd, T. F., Lyons, C. R. (2002). A polymorphic region in Mycobacterium abscessus contains a novel insertion sequence element. Microbiology 148: 2987-2996 [Abstract] [Full Text]  
  • Qian, Y., Lee, J. H., Holmes, R. K. (2002). Identification of a DtxR-Regulated Operon That Is Essential for Siderophore-Dependent Iron Uptake in Corynebacterium diphtheriae. J. Bacteriol. 184: 4846-4856 [Abstract] [Full Text]  
  • Twigg, P. D., Parthasarathy, G., Guerrero, L., Logan, T. M., Caspar, D. L. D. (2001). Disordered to ordered folding in the regulation of diphtheria toxin repressor activity. Proc. Natl. Acad. Sci. USA 98: 11259-11264 [Abstract] [Full Text]  
  • Feese, M. D., Ingason, B. P., Goranson-Siekierke, J., Holmes, R. K., Hol, W. G. J. (2001). Crystal Structure of the Iron-dependent Regulator from Mycobacterium tuberculosis at 2.0-A Resolution Reveals the Src Homology Domain 3-like Fold and Metal Binding Function of the Third Domain. J. Biol. Chem. 276: 5959-5966 [Abstract] [Full Text]  


Copyright © 2009 by the American Society for Microbiology.   For an alternate route to Journals.ASM.org, visit: http://intl-journals.asm.org | More Info»