MBE Advance Access published online on January 13, 2007
Molecular Biology and Evolution, doi:10.1093/molbev/msm006
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© 2007 The Authors
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.
Research Article |
Compensatory Evolution in Response to a Novel RNA Polymerase: Orthologous Replacement of a Central Network Gene
a Section of Integrative Biology
b Section of Molecular Genetics and Microbiology
c Institute for Cellular and Molecular Biology, University of Texas, Austin, TX 78712
corresponding authors: Jim Bull, bull{at}mail.utexas.edu, 512-471-5661. Ian Molineux, molineux{at}mail.utexas.edu, 512-471-3143
Accepted for publication December 18, 2006.
A bacteriophage genome was forced to evolve a new system of regulation by replacing its RNA polymerase (RNAP) gene, a central component of the phage developmental pathway, with that of a relative. The experiment used the obligate lytic phage T7 and the RNAP gene of phage T3. T7 RNAP uses 17 phage promoters, which are responsible for all middle and late gene expression, DNA replication and progeny maturation, but the enzyme has known physical contacts with only two other phage proteins. T3 RNAP was supplied in trans by the bacterial host to a T7 genome lacking its own RNAP gene and the phage population was continually propagated on naive bacteria throughout the adaptation. Evolution of the T3 RNAP gene was thereby prevented, and selection was for the evolution of regulatory signals throughout the phage genome. T3 RNAP transcribes from T7 promoters only at low levels, but a single mutation in the promoter confers high expression, providing a ready mechanism for re-evolution of gene expression in this system. When selected for rapid growth, fitness of the engineered phage evolved from a low of 5 doublings/hr to 33 doublings/hr, close to the expected maximum of 37 doublings/hr. However the experiment was terminated before it could be determined accurately that fitness had reached an obvious plateau, and it is not known whether further adaptation could have resulted in complete recovery of fitness. More than 30 mutations were observed in the evolved genome, but changes were found in only 9 of the 16 promoters, and several coding changes occurred in genes with no known contacts with the RNAP. Surprisingly, the T7 genome adapted to T3 RNAP also maintained high fitness when using T7 RNAP, suggesting that the extreme incompatibility of T7 elements with T3 RNAP is not an invariant property of divergence in these expression systems.
Key Words: experimental • evolution • bacteriophage • genomics • adaptation