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Microbiology 155 (2009), 1669-1679; DOI  10.1099/mic.0.024455-0
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Microbiology 155 (2009), 1669-1679; DOI  10.1099/mic.0.024455-0
© 2009 Society for General Microbiology

Identification of the binding site of the {sigma}54 hetero-oligomeric FleQ/FleT activator in the flagellar promoters of Rhodobacter sphaeroides

J. Peña-Sánchez1, S. Poggio1, U. Flores-Pérez1, A. Osorio1, C. Domenzain1, G. Dreyfus2 and L. Camarena1

1 Instituto de Investigaciones Biomédicas, Universidad Nacional Autónoma de México, México D.F. 04510, Mexico
2 Instituto de Fisiología Celular, Universidad Nacional Autónoma de México, México D.F. 04510, Mexico

Expression of the flagellar genes in Rhodobacter sphaeroides is dependent on one of the four sigma-54 factors present in this bacterium and on the enhancer binding proteins (EBPs) FleQ and FleT. These proteins, in contrast to other well-characterized EBPs, carry out activation as a hetero-oligomeric complex. To further characterize the molecular properties of this complex we mapped the binding sites or upstream activation sequences (UASs) of six different flagellar promoters. In most cases the UASs were identified at approximately 100 bp upstream from the promoter. However, the activity of the divergent promoters flhAp-flgAp, which are separated by only 53 bp, is mainly dependent on a UAS located approximately 200 bp downstream from each promoter. Interestingly, a significant amount of activation mediated by the upstream or contralateral UAS was also detected, suggesting that the architecture of this region is important for the correct regulation of these promoters. Sequence analysis of the regions carrying the potential FleQ/FleT binding sites revealed a conserved motif. In vivo footprinting experiments with the motAp promoter allowed us to identify a protected region that overlaps with this motif. These results allow us to propose a consensus sequence that represents the binding site of the FleQ/FleT activating complex.

Correspondence
L. Camarena
rosal{at}servidor.unam.mx

Abbreviations: DMS, dimethyl sulfate; EBP, enhancer binding protein; HTH, helix–turn–helix; IHF, integration host factor; UAS, upstream activation sequence

Two supplementary tables, listing oligonucleotides used in this work and relevant sequences for motif finding, are available with the online version of this paper.






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Copyright © 2009 Society for General Microbiology.