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Nucleic Acids Research Advance Access originally published online on November 3, 2008
Nucleic Acids Research 2008 36(22):6948-6958; doi:10.1093/nar/gkm499
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Nucleic Acids Research, 2008, Vol. 36, No. 22 6948-6958
© 2008 The Author(s)
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.0/uk/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

Genomics

A whole-genome approach to identifying protein binding sites: promoters in Methanocaldococcus (Methanococcus) jannaschii

Enhu Li1, Claudia I. Reich2,3 and Gary J. Olsen2,4,*

1Division of Biology, California Institute of Technology, 1200 E. California Blvd., Pasadena, CA 91125, 2Department of Microbiology, 3National Center for Supercomputing Applications and 4Institute for Genomic Biology, University of Illinois, Urbana-Champaign, Urbana, IL 61801, USA

*To whom correspondence should be addressed. Tel: +1 217 244 0616; Fax: +1 217 244 6697; Email: gary{at}life.uiuc.edu

Received April 23, 2007. Revised June 6, 2007. Accepted June 7, 2007.

We have adapted an electrophoretic mobility shift assay (EMSA) to isolate genomic DNA fragments that bind the archaeal transcription initiation factors TATA-binding protein (TBP) and transcription factor B (TFB) to perform a genome-wide search for promoters. Mobility-shifted fragments were cloned, tested for their ability to compete with known promoter-containing fragments for a limited concentration of transcription factors, and sequenced. We applied the method to search for promoters in the genome of Methanocaldococcus jannaschii. Selection was most efficient for promoters of tRNA genes and genes for several presumed small non-coding RNAs (ncRNA). Protein-coding gene promoters were dramatically underrepresented relative to their frequency in the genome. The repeated isolation of these genomic regions was partially rectified by including a hybridization-based screening. Sequence alignment of the affinity-selected promoters revealed previously identified TATA box, BRE, and the putative initiator element. In addition, the conserved bases immediately upstream and downstream of the BRE and TATA box suggest that the composition and structure of archaeal natural promoters are more complicated.


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J. Zhang, E. Li, and G. J. Olsen
Protein-coding gene promoters in Methanocaldococcus (Methanococcus) jannaschii
Nucleic Acids Res., June 1, 2009; 37(11): 3588 - 3601.
[Abstract] [Full Text] [PDF]


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