Lin Chao

PERSPECTIVE:EVOLUTION AND DETECTION OF GENETIC ROBUSTNESS

Abstract Robustness is the invariance of phenotypes in the face of perturbation. The robustness of phenotypes appears at various levels of biological organization, including gene expression, protein folding, metabolic flux, physiological homeostasis, development, and even organismal fitness. The mechanisms underlying robustness are diverse, ranging from thermodynamic stability at the RNA and protein level to behavior at the organismal level. Phenotypes can be robust either against heritable perturbations (e.g., mutations) or nonheritable perturbations (e.g., the weather). Here we primarily focus on the first kind of robustness—genetic robustness—and survey three growing avenues of research: (1) measuring genetic robustness in nature and in the laboratory; (2) understanding the evolution of genetic robustness; and (3) exploring the implications of genetic robustness for future evolution.

Prisoner's dilemma in an RNA virus

The evolution of competitive interactions among viruses was studied in the RNA phage φ6 at high and low multiplicities of infection (that is, at high and low ratios of infecting phage to host cells). At high multiplicities, many phage infect and reproduce in the same host cell, whereas at low multiplicities the viruses reproduce mainly as clones. An unexpected result of this study was that phage grown at high rates of co-infection increased in fitness initially, but then evolved lowered fitness. Here we show that the fitness of the high-multiplicity phage relative to their ancestors generates a pay-off matrix conforming to the prisoners dilemma strategy of game theory. In this strategy, defection (selfishness) evolves, despite the greater fitness pay-off that would result if all players were to cooperate. Viral cooperation and defection can be defined as, respectively, the manufacturing and sequestering of diffusible (shared) intracellular products. Because the low-multiplicity phage did not evolve lowered fitness, we attribute the evolution of selfishness to the lack of clonal structure and the mixing of unrelated genotypes at high multiplicity.

Perspective: Sign epistasis and genetic constraint on evolutionary trajectories.

Abstract Epistasis for fitness means that the selective effect of a mutation is conditional on the genetic background in which it appears. Although epistasis is widely observed in nature, our understanding of its consequences for evolution by natural selection remains incomplete. In particular, much attention focuses only on its influence on the instantaneous rate of changes in frequency of selected alleles via epistatic contribution to the additive genetic variance for fitness. Thus, in this framework epistasis only has evolutionary importance if the interacting loci are simultaneously segregating in the population. However, the selective accessibility of mutational trajectories to high fitness genotypes may depend on the genetic background in which novel mutations appear, and this effect is independent of population polymorphism at other loci. Here we explore this second influence of epistasis on evolution by natural selection. We show that it is the consequence of a particular form of epistasis, which we designate sign epistasis. Sign epistasis means that the sign of the fitness effect of a mutation is under epistatic control; thus, such a mutation is beneficial on some genetic backgrounds and deleterious on others. Recent experimental innovations in microbial systems now permit assessment of the fitness effects of individual mutations on multiple genetic backgrounds. We review this literature and identify many examples of sign epistasis, and we suggest that the implications of these results may generalize to other organisms. These theoretical and empirical considerations imply that strong genetic constraint on the selective accessibility of trajectories to high fitness genotypes may exist and suggest specific areas of investigation for future research.

RESOURCE-LIMITED GROWTH, COMPETITION, AND PREDATION: A MODEL AND EXPERIMENTAL STUDIES WITH BACTERIA AND BACTERIOPHAGE

We present a model of resource-limited population growth, competition, and predation based on what we believe to be biologically realistic assumptions about the relationship between resources and the growth of primary consumers and about the interaction of the primary consumers with the predators that prey upon them. Consideration is given to an equable habitat in which resources are continually supplied and wastes are continually removed. The general properties of this model are examined and two specific cases are studied in some detail: (i) one resource, one prey, and one predator, and (ii) one resource, two prey, and one predator. Particular consideration is given to the conditions which will permit the continued coexistence of the interacting populations. In conceiving this model we were guided by the specific case of bacteria and their virulent viruses. To study its validity we compare the theoretical predictions with the experimental results from continuous-culture populations of the bacterium E. coli and the phage T2. Structurally stable equilibria with all populations coexisting are possible when the number of distinct predator populations is not more than the number of distinct prey populations and the number of the latter is not more than the sum of the number of resources and the number of predator populations. For one resource, one prey, and one predator there are stable states of coexistence. Given specific resource utilization functions, the levels of these equilibria or oscillations and the range of parameter values required for stability can be determined. In situations where a stable equilibrium exists for the one predator-one prey system, a second population of primary consumers which is totally resistant to predation can also be maintained. Sufficient conditions for this to occur are that the resistant population have a lower intrinsic growth rate than the sensitive but that the former can, nevertheless, survive and multiply living on the resources present in the one predator-one prey system. If, however, this second species of primary consumer becomes slightly sensitive to predation, it may then entirely displace the original sensitive population. Stable equilibria were observed in glucose minimal continuous cultures containing T2 sensitive E. coli B and T2 phage. The equilibrium concentration of glucose and the densities of the populations were similar to those predicted by the model. However, with the estimated values of the parameters, the experimental system fell in the range where the model predicted that the oscillations would increase to the point where the populations would eventually become extinct. Stable equilibria were also observed for a glucose-limited chemostat culture containing T2 phage together with a T2-sensitive clone of E. coli B and a T2-resistant strain of E. coli K12. This system fulfilled the conditions under which the theory predicts that stable coexistence will occur. We discuss the validity of this model as a general analogue of resource limited growth, competition, and predation in planktonic species. We also consider the implications of these theoretical and experimental results for the general theory of competition and predation and for the specific problem of coexistence for bacteria and their virulent viruses.

Competition between high and low mutating strains of escherichia coli

The dynamics of bacterial populations are often characterized by several distinctive features: under optimal growth conditions they double every few hours; they usually contain in excess of 106 individuals; higher fitness mutants have a good chance of arising in a population since average mutation rates are approximately 10-6 to 10-7 per gene replication; new favorable mutations, in the absence of genetic recombination, always increase to fixation in linkage with the genome of the parent clone in which they originally occurred; and higher fitness mutants often exhibit 10% to 20% higher growth rates than their parental clones. Consequently, when populations of such organisms are exposed to a new environment, a series of replacement cycles rapidly ensues, each cycle corresponding to the fixation of a higher fitness mutation in linkage with the genome of its parent clone. The linkage between the new mutation and the genome of the parent clone, and the rapidity of these clonal replacements, are two features that distinguish such asexual populations from ones that reproduce sexually. The existence of such cycles in asexual populations was first studied systematically by Atwood et al. (1951) in a study with laboratory populations of Escherichia coli in long-term cultures. By comparing the relative fitness of a series of bacterial clones isolated from these cultures, Atwood et al. were able to show that populations underwent a succession of clonal changes, each clonal replacement

A Complex Community in a Simple Habitat: An Experimental Study with Bacteria and Phage

Continuous culture populations of the bacterium especially coli and its virulent virus T7 have been studied as a model of a predator—prey in a simple habitat. These organisms maintain apparently stable states of coexistence in: (1) a phage—limited situation where all of the bacteria are sensitive to the coexisting virus and the sole, and potentially limiting carbon source, glucose, is present in excess; and (2) a resource—limited situation where the majority of the bacteria are resistant to these phage and in which there is little free glucose. The composition of these interacting populations is examined in detail and evidence indicating that this simple experimental culture system can support relatively complex communities is presented. In the predator—limited situation, two populations at each of two trophic levels can be maintained; the wild—type bacterial and phage strains, denoted B0 and T70, a mutant bacterial clone which is resistant to T70, denote B1 and a host range mutant phage, T71 which is capable of growth on both B0 and B1. In the resource—limited situation, three populations of bacteria and two populations of phage can coexist. The include the above described clones and a third bacterial strain, B2, which is resistant to both T70 and T71. In phage—free competition, the wild—type B0 bacterial clone has a marked advantage relative to both B1 and B2 while no difference is detected between B1 and B2. When competing for a B0 host, the wild—type T70 phage clone has a marked advantage over T71. The fit of these observations to some previously developed theory of resource—limited growth, competition and predation is discussed and a mechanism to account for the persistence of these communities is presented. The latter assumes that their stability can be attributed solely to intrinsic factors, i.e., the population growth and interaction properties of the organisms in this continuous culture habitat.

Evolvability of an RNA virus is determined by its mutational neighbourhood

The ubiquity of mechanisms that generate genetic variation has spurred arguments that evolvability, the ability to generate adaptive variation, has itself evolved in response to natural selection. The high mutation rate of RNA viruses is postulated to be an adaptation for evolvability, but the paradox is that whereas some RNA viruses evolve at high rates, others are highly stable. Here we show that evolvability in the RNA bacteriophage φ6 is also determined by the accessibility of advantageous genotypes within the mutational neighbourhood (the set of mutants one or a few mutational steps away). We found that two φ6 populations that were derived from a single ancestral phage repeatedly evolved at different rates and toward different fitness maxima. Fitness measurements of individual phages showed that the fitness distribution of mutants differed between the two populations. Whereas population A, which evolved toward a higher maximum, had a distribution that contained many advantageous mutants, population B, which evolved toward a lower maximum, had a distribution that contained only deleterious mutants. We interpret these distributions to measure the fitness effects of genotypes that are mutationally available to the two populations. Thus, the /evolvability of φ6 is constrained by the distribution of its mutational neighbours, despite the fact that this phage has the characteristic high mutation rate of RNA viruses.

RAPID EVOLUTIONARY ESCAPE BY LARGE POPULATIONS FROM LOCAL FITNESS PEAKS IS LIKELY IN NATURE

Abstract Fitness interactions between loci in the genome, or epistasis, can result in mutations that are individually deleterious but jointly beneficial. Such epistasis gives rise to multiple peaks on the genotypic fitness landscape. The problem of evolutionary escape from such local peaks has been a central problem of evolutionary genetics for at least 75 years. Much attention has focused on models of small populations, in which the sequential fixation of valley genotypes carrying individually deleterious mutations operates most quickly owing to genetic drift. However, valley genotypes can also be subject to mutation while transiently segregating, giving rise to copies of the high fitness escape genotype carrying the jointly beneficial mutations. In the absence of genetic recombination, these mutations may then fix simultaneously. The time for this process declines sharply with increasing population size, and it eventually comes to dominate evolutionary behavior. Here we develop an analytic expression for Ncrit, the critical population size that defines the boundary between these regimes, which shows that both are likely to operate in nature. Frequent recombination may disrupt high‐fitness escape genotypes produced in populations larger than Ncrit before they reach fixation, defining a third regime whose rate again slows with increasing population size. We develop a novel expression for this critical recombination rate, which shows that in large populations the simultaneous fixation of mutations that are beneficial only jointly is unlikely to be disrupted by genetic recombination if their map distance is on the order of the size of single genes. Thus, counterintuitively, mass selection alone offers a biologically realistic resolution to the problem of evolutionary escape from local fitness peaks in natural populations.

Understanding the evolutionary fate of finite populations: the dynamics of mutational effects.

The most consistent result in more than two decades of experimental evolution is that the fitness of populations adapting to a constant environment does not increase indefinitely, but reaches a plateau. Using experimental evolution with bacteriophage, we show here that the converse is also true. In populations small enough such that drift overwhelms selection and causes fitness to decrease, fitness declines down to a plateau. We demonstrate theoretically that both of these phenomena must be due either to changes in the ratio of beneficial to deleterious mutations, the size of mutational effects, or both. We use mutation accumulation experiments and molecular data from experimental evolution to show that the most significant change in mutational effects is a drastic increase in the rate of beneficial mutation as fitness decreases. In contrast, the size of mutational effects changes little even as organismal fitness changes over several orders of magnitude. These findings have significant implications for the dynamics of adaptation.

Epistasis and its consequences for the evolution of natural populations

Throughout the neodarwinian synthesis, theorists have debated the role of gene interactions, or epistasis, in the evolutionary process. Unfortunately, empirical measurement of the role of epistasis in the evolution of natural populations has, until now, been difficult. Two developments in empirical approaches have occurred: (1) application of theory to the evolution of natural populations, and (2) the concurrent development of molecular marker-assisted techniques to understand the architecture of quantitative genetic variation. Thus, exciting developments in both theory and empirical data collection provide the stimulus for documenting the role of epistasis in the evolutionary process.