Article

Global Discovery of Adaptive Mutations

Department of Molecular Biology & Lewis-Sigler Institute for Integrative Genomics, Princeton University, Princeton, New Jersey, USA.
Nature Methods (Impact Factor: 32.07). 09/2009; 6(8):581-3. DOI: 10.1038/nmeth.1352
Source: PubMed

ABSTRACT

Although modern DNA sequencing enables rapid identification of genetic variation, characterizing the phenotypic consequences of individual mutations remains a labor-intensive task. Here we describe array-based discovery of adaptive mutations (ADAM), a technology that searches an entire bacterial genome for mutations that contribute to selectable phenotypic variation between an evolved strain and its parent. We found that ADAM identified adaptive mutations in laboratory-evolved Escherichia coli strains with high sensitivity and specificity.

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Available from: Hani Goodarzi, Aug 22, 2014
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    • "In this study, we address all these questions. Solving these questions is relevant for understanding the adaptive mechanisms prevalent in our experimental conditions and also in other evolutionary experiments, because mutations that affect transcriptional regulators (such as RNAP and the Rho termination factor) typically appear in the early stages of stress adaptation (Applebee et al. 2008; Goodarzi et al. 2009; Kishimoto et al. 2010). Numerous observations point to the possibility that highly pleiotropic mutations in transcriptional regulators could be the first step of a general mechanism of adaptation (Fong et al. 2005). "
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    ABSTRACT: The temporal change of phenotypes during the adaptive process remains largely unexplored, as do the genetic changes that affect these phenotypic changes. Here we focused on three mutations that rose to high frequency in the early stages of adaptation within 12 Escherichia coli populations subjected to thermal stress (42 °C). All the mutations were in the rpoB gene, which encodes the RNA polymerase beta subunit. For each mutation, we measured the growth curves and gene expression (mRNAseq) of clones at 42 °C. We also compared growth and gene expression with their ancestor under unstressed (37 °C) and stressed conditions (42 °C). Each of the three mutations changed the expression of hundreds of genes and conferred large fitness advantages, apparently through the restoration of global gene expression from the stressed toward the prestressed state. These three mutations had a similar effect on gene expression as another single mutation in a distinct domain of the rpoB protein. Finally, we compared the phenotypic characteristics of one mutant, I572L, with two high-temperature adapted clones that have this mutation plus additional background mutations. The background mutations increased fitness, but they did not substantially change gene expression. We conclude that early mutations in a global transcriptional regulator cause extensive changes in gene expression, many of which are likely under positive selection for their effect in restoring the prestress physiology.
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    • "Under this approach, a set of synthetic oligonucleotides representing the mutations found by WGS randomly undergoes recombination with genomic DNA, creating a library of allelic replacement strains. Second, array-based discovery of adaptive mutations (ADAM) uses genetic foot printing to selectively identify mutations that provide a competitive advantage to the cell (Goodarzi et al, 2009). When reconstructing phenotypes through allelic replacement, ADAM allows the researcher to avoid the wasted effort of reconstructing non-beneficial mutations. "
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