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- The evolution of a key innovation in an experimental population of Escherichia coli : a tale of opportunity, contingency, and co-option
- Blount, Zachary David
- Electronic Theses & Dissertations
The importance of historical contingency in evolution has been extensively debated over the last few decades, but direct empirical tests have been rare. Twelve initially identical populations of
E. coliwere founded in 1988 to investigate this issue. They have since evolved for more than 50,000 generations in a glucose-limited medium that also contains a citrate. However, the inability to use citrate as a carbon source under oxic conditions is a species-defining trait of ...
Show moreThe importance of historical contingency in evolution has been extensively debated over the last few decades, but direct empirical tests have been rare. Twelve initially identical populations of
E. coliwere founded in 1988 to investigate this issue. They have since evolved for more than 50,000 generations in a glucose-limited medium that also contains a citrate. However, the inability to use citrate as a carbon source under oxic conditions is a species-defining trait of E. coli. A weakly Cit +variant capable of aerobic citrate utilization finally evolved in one population just prior to 31,500 generations. Shortly after 33,000 generations, the population experienced a several-fold expansion as strongly Cit +variants rose to numerical dominance (but not fixation). The Cit +trait was therefore a key innovation that increased both population size and diversity by opening a previously unexploited ecological opportunity.The long-delayed and unique evolution of the Cit +innovation might be explained by two possible hypotheses. First, evolution of the Cit +function may have required an extremely rare mutation. Alternately, the evolution of Cit +may have been contingent upon one or more earlier mutations that had accrued over the population's history. I tested these hypotheses in a series of experiments in which I "replayed" evolution from different points in the population's history. I observed no Cit +mutants among 8.4 x 10 12ancestral cells, nor among 9 x 10 12cells from 60 clones sampled in the first 15,000 generations. However, I observed a significantly greater tendency to evolve Cit +among later clones. These results indicate that one or more earlier mutations potentiated the evolution of Cit +by increasing the rate of mutation to Cit +to an accessible, though still very low, level. The evolution of the Cit +function was therefore contingent on the particular history of the population in which it occurred.I investigated the Cit +innovation's history and genetic basis by sequencing the genomes of 29 clones isolated from the population at various time points. Analysis of these genomes revealed that at least 3 distinct clades coexisted for more than 10,000 generations prior to the innovation's evolution. The Cit +trait originated in one clade by a tandem duplication that produced a new regulatory module in which a silent citrate transporter was placed under the control of an aerobically-expressed promoter. Subsequent increases in the copy number of this new regulatory module refined the initially weak Cit +phenotype, leading to the population expansion. The 3 clades varied in their propensity to evolve the novel Cit +function, though genotypes able to do so existed in all 3, implying that potentiation involved multiple mutations.My findings demonstrate that historical contingency can significantly impact evolution, even under the strictest of conditions. Moreover, they suggest that contingency plays an especially important role in the evolution of novel innovations that, like Cit +, require prior construction of a potentiating genetic background, and are thus not easily evolved by gradual, cumulative selection. Contingency may therefore have profoundly shaped life's evolution given the importance of evolutionary novelties in the history of life. Finally, the genetic basis of the Cit +function illustrates the importance of promoter capture and altered gene regulation in mediation the exaptation events that often underlie evolutionary innovations.
- Experimental evolution and ecological consequences : new niches and changing stoichiometry
- Turner, Caroline B.
- Electronic Theses & Dissertations
Evolutionary change can alter the ecological conditions in which organisms live and continue to evolve. My dissertation research used experimental evolution to study two aspects of evolutionary change with ecological consequences: the generation of new ecological niches and evolution of the elemental composition of biomass. I worked with the long-term evolution experiment (LTEE), which is an ongoing experiment in which E. coli have evolved under laboratory conditions for more than 60,000...
Show moreEvolutionary change can alter the ecological conditions in which organisms live and continue to evolve. My dissertation research used experimental evolution to study two aspects of evolutionary change with ecological consequences: the generation of new ecological niches and evolution of the elemental composition of biomass. I worked with the long-term evolution experiment (LTEE), which is an ongoing experiment in which E. coli have evolved under laboratory conditions for more than 60,000 generations. The LTEE began with extremely simple ecological conditions. Twelve populations were founded from a single bacterial genotype and growth was limited by glucose availability. In Chapter 1, I focused on a population within the LTEE in which some of the bacteria evolved the ability to consume a novel resource, citrate. Citrate was present in the growth media throughout the experiment, but E. coli is normally unable to consume it under aerobic conditions. The citrate consumers (Cit+) coexisted with a clade of bacteria which were unable to consume citrate (Cit-). Specialization on glucose, the standard carbon source in the LTEE, was insufficient to explain the frequency-dependent coexistence of Cit- with Cit+. Instead Cit– evolved to cross-feed on molecules released by Cit+. The evolutionary innovation of citrate consumption led to a more complex ecosystem in which two co-existing ecotypes made use of five different carbon sources.After 10,000 generations of coexistence, Cit- went extinct from the population (Chapter 2). I conducted replay experiments, re-evolving for 500 generations 20 replicate populations from prior to extinction. Cit- was retained in all populations, indicating that the extinction was not deterministic. Furthermore, when I added small numbers of Cit- to the population after extinction, Cit- was able to reinvade. It therefore appears that the Cit- extinction was not due to exclusion by Cit+, but rather to unknown laboratory variation.Chapter 3 shifts focus to studying evolutionary changes in stoichiometry, the ratio of different elements within organisms’ biomass. Variation in stoichiometry between organisms has important ecological consequences, but the evolutionary origin of that variation had not previously been studied experimentally. Growth in the LTEE is carbon limited and nitrogen and phosphorus are abundant. Additionally, daily transfer to fresh media selects for increased growth rate, which other research has suggested correlates to higher phosphorus content. Consistent with our predictions based on this environment, clones isolated after 50,000 generations of evolution had significantly higher nitrogen and phosphorus content than ancestral clones. There was no change in the proportion of carbon in biomass, but the total amount of carbon retained in biomass increased, indicating that the bacteria also evolved higher carbon use efficiency.To test whether the increases in nitrogen and phosphorus observed in the LTEE were a result of carbon limitation or were side effects of other selective factors in the experiment, I evolved clones from the LTEE for 1000 generations under nitrogen rather than carbon limitation (Chapter 4). The stoichiometry of the bacteria did change over the course of 1000 generations, indicating that evolution of stoichiometry can occur over relatively short time frames. Unexpectedly however, the evolved bacteria had higher nitrogen and phosphorus content. It appears that the bacteria were initially poor at incorporating nitrogen into biomass, but evolved improved nitrogen uptake.