This Article Full Text Full Text (PDF) Supplemental material Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Copyright Information Books from ASM Press MicrobeWorld Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Michalowski, C. B. Articles by Little, J. W. Search for Related Content PubMed PubMed Citation Articles by Michalowski, C. B. Articles by Little, J. W.

Sequence Tolerance of the Phage P_RM Promoter: Implications for Evolution of Gene Regulatory Circuitry

Christine B. Michalowski,¹ Megan D. Short,^1, and John W. Little^1,2*

Department of Biochemistry and Molecular Biophysics,¹ Department of Molecular and Cellular Biology, University of Arizona, Tucson, Arizona²

Received 18 February 2004/ Accepted 1 September 2004

Much of the gene regulatory circuitry of phage centers on a complex region called the O_R region. This 100-bp region is densely packed with regulatory sites, including two promoters and three repressor-binding sites. The dense packing of this region is likely to impose severe constraints on its ability to change during evolution, raising the question of how the specific arrangement of sites and their exact sequences could evolve to their present form. Here we ask whether the sequence of a cis-acting site can be widely varied while retaining its function; if it can, evolution could proceed by a larger number of paths. To help address this question, we developed a cloning vector that allowed us to clone fragments spanning the O_R region. By using this vector, we carried out intensive mutagenesis of the P_RM promoter, which drives expression of CI repressor and is activated by CI itself. We made a pool of fragments in which 8 of the 12 positions in the –35 and –10 regions were randomized and cloned this pool into the vector, making a pool of P_RM variant phage. About 10% of the P_RM variants were able to lysogenize, suggesting that the regulatory circuitry is compatible with a wide range of P_RM sequences. Analysis of several of these phages indicated a range of behaviors in prophage induction. Several isolates had induction properties similar to those of the wild type, and their promoters resembled the wild type in their responses to CI. We term this property of different sequences allowing roughly equivalent function "sequence tolerance " and discuss its role in the evolution of gene regulatory circuitry.

Supplemental material for this article may be found at http://jb.asm.org/.

Present address: Department of Pediatrics, University of Colorado Health Sciences Center, Denver, CO 80262.

This article has been cited by other articles:

• Gedeon, T., Mischaikow, K., Patterson, K., Traldi, E. (2008). Binding Cooperativity in Phage {lambda} is Not Sufficient to Produce an Effective Switch. Biophys. J 94: 3384-3392 [Abstract] [Full Text]  
• Anderson, L. M., Yang, H. (2008). DNA looping can enhance lysogenic CI transcription in phage lambda. Proc. Natl. Acad. Sci. USA 105: 5827-5832 [Abstract] [Full Text]  
• Babic, A. C., Little, J. W. (2007). Cooperative DNA binding by CI repressor is dispensable in a phage {lambda} variant. Proc. Natl. Acad. Sci. USA 104: 17741-17746 [Abstract] [Full Text]  
• Schubert, R. A., Dodd, I. B., Egan, J. B., Shearwin, K. E. (2007). Cro's role in the CI Cro bistable switch is critical for {lambda}'s transition from lysogeny to lytic development. Genes Dev. 21: 2461-2472 [Abstract] [Full Text]  
• Atsumi, S., Little, J. W. (2006). A synthetic phage {lambda} regulatory circuit. Proc. Natl. Acad. Sci. USA 103: 19045-19050 [Abstract] [Full Text]  
• Atsumi, S., Little, J. W. (2006). Role of the lytic repressor in prophage induction of phage {lambda} as analyzed by a module-replacement approach. Proc. Natl. Acad. Sci. USA 103: 4558-4563 [Abstract] [Full Text]  
• Michalowski, C. B., Little, J. W. (2005). Positive Autoregulation of cI Is a Dispensable Feature of the Phage {lambda} Gene Regulatory Circuitry. J. Bacteriol. 187: 6430-6442 [Abstract] [Full Text]