Laboratory of Genetics, Wageningen University, Dreijenlaan 2, 6703 HA Wageningen, The Netherlands1
Tel: +31 317 483144. Fax: +31 317 483146. e-mail: arjan.devisser{at}genetics.dpw.wau.nl
Keywords: mutator, mutation rate, evolution, clonal interference
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Mutation rate evolution |
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The best known class of mutation-rate mutants is that of the so-called general mutators, which have increased mutation rates throughout their genome due to defects in DNA proofreading and repair functions. These mutants have constitutively higher mutation rates, in contrast to transient mutators, where the increase of the mutation rate depends on environmental conditions (Taddei et al., 1995 ; Rosenberg et al., 1998
; Foster, 1999
). For the sake of simplicity and because their fate depends on very much the same evolutionary costs and benefits as those of transient mutators, I will concentrate on the evolutionary fate of genetically stable mutators only.
Mutators are found among both natural and laboratory populations of bacteria (Jyssum, 1960 ; Gross & Siegel, 1981
; LeClerc et al., 1996
; Mao et al., 1997
; Sniegowski et al., 1997
; Matic et al., 1997
; Boe et al., 2000
; Oliver et al., 2000
; Giraud et al., 2001a
; Denamur et al., 2002
). Most spontaneous mutator mutants appear defective in mismatch repair pathways, often because the mutS gene is inactivated (Miller, 1996
; LeClerc et al., 1996
; Matic et al., 1997
; Sniegowski et al., 1997
; Funchain et al., 2001
). Depending on the type of function that is defective, mutators can have mutation rates that are moderately (
10-fold) to strongly (10010000-fold) increased (Miller, 1996
). Anti-mutator mutants with lower than normal mutation rates have also been observed (reviewed by Sniegowski et al., 2000
), albeit at much lower frequencies than mutators (Drake et al., 1998
). However, within mutator populations, mutants with lower mutation rates (but higher than wild-type) can evolve (Tröbner & Piechocki, 1984
; Giraud et al., 2001a
). These studies show that evolution of the mutation rate is possible in evolving populations of bacteria. Here, I review a number of recent studies that shed light on the short- and long-term fate of microbial mutators, and provide new insights into the mechanisms and dynamics of mutation-rate evolution.
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Short-term fate |
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Hitch-hiking with beneficial mutations
How might such indirect benefits of an increased mutation rate be realized? Because mutator mutants introduce more mutations in their offspring than the rest of the population, they will also generate more beneficial mutations. In asexually reproducing populations, a mutator allele (the modifier) and the mutations it produces (the effect) remain physically linked; their fates are thus tied. Once natural selection causes a beneficial mutation in a mutator background to increase in frequency, the mutator allele will hitch-hike to high frequency along with it (Maynard-Smith & Haigh, 1974 ; Chao & Cox, 1983
). Natural selection favouring mutators is thus indirect.
For a mutator to increase in frequency within a population of finite size, it needs only to produce a successful beneficial mutation in one of its descendants before it occurs in the remaining wild-type majority of the population. To compensate for its numerical disadvantage, a mutator subpopulation needs a mutation rate which is increased by more than the inverse of its numerical disadvantage to have a better chance to produce the next beneficial mutation (Chao & Cox, 1983 ). Given that the relative size of a mutator subpopulation is smaller than 10-3, a more than 1000-fold increase of the mutation rate would be needed to win the race to produce the next beneficial mutation. The fact is, however, that a mutator subpopulation has an increased per capita probability of producing beneficial mutations, which is equal to its strength. Therefore, if adaptation involves the sequential selection of several beneficial mutations, these chance events add up to a robust benefit for the mutator subpopulation to win the competition and hitch-hike to high frequency (Mao et al., 1997
; Miller et al., 1999
; Tenaillon et al., 1999
).
Conditions for hitch-hiking
The indirect selective benefit of mutator mutants thus depends on opportunities for adaptation. The fraction of mutations that improve adaptation depends on the evolutionary history of the population in the present environment. If the population is already well adapted, then most if not all mutations will have negative or at best neutral effects on fitness, and the mutator subpopulation lacks opportunities for hitch-hiking. Instead, it will pay the cost of the increased production of deleterious mutations. However, if the environment is in some respect novel, adaptation is not perfect and the mutator subpopulation may outcompete the wild-type majority by its association with a higher per-cell number of beneficial mutations. Novel environmental conditions are abundant if environments (i) change rapidly in time, as for pathogens coping with host defence mechanisms such as the immune system, or (ii) are heterogeneous and consist of multiple ecological niches. For example, Pseudomonas aeruginosa living in the lungs of cystic fibrosis patients is thought to cope with rapidly changing conditions due to host defence mechanisms and therapeutic treatment, as well as with a heterogeneous environment (Oliver et al., 2000 ). Consistent with the abundance of opportunities for hitch-hiking is the high frequency of mutators found in cystic fibrosis lungs (Oliver et al., 2000
).
Hitch-hiking of mutator alleles with beneficial mutations depends on the physical linkage between modifier (mutator allele) and effect (beneficial mutation). If modifier and effect are separated regularly, as in sexual populations, mutators are not expected to profit from their indirect beneficial effect (Tenaillon et al., 2000 ). As a consequence, mutators are not expected to be as frequent in sexual as in asexual populations, a premise that needs to be verified empirically. However, other forms of constitutive mutators can evolve in sexual populations. One example are transposable elements, such as transposons and insertion sequences, which can be considered mutator genes due to their mutagenic properties (Chao et al., 1983
). The absolute linkage of transposable elements to their effect allows them to hitch-hike to high frequency even in sexual populations. Another form of mutator that may evolve in sexual populations are so-called contingency genes with increased mutation rate (Moxon et al., 1994
; Metzgar & Wills, 2000
). Here, again, modifier and effect are tightly linked. The presence of short-sequence DNA repeats in the promoter or coding region of a gene, or local cassette-like mechanisms to activate alternative alleles of a gene are among the mechanisms that can increase the mutation rate at specific loci only. This way, only genes involved in adaptation to unpredictable aspects of the environment might profit from an increased production of beneficial mutations, while the costs of producing a high load of deleterious mutations is very much limited. Examples of such local mutators include virulence genes of pathogenic bacteria (Moxon et al., 1994
; Metzgar & Wills, 2000
).
Some mutator alleles (e.g. with deficient mismatch-repair genes mutS and mutL) not only increase the mutation rate, but also tend to have a hyper-recombination phenotype (Rayssiguier et al., 1989 ). On the one hand this additional effect offers further opportunities for genetic hitch-hiking, i.e. along with beneficial recombinants. A recent study showed that selection for recombinants of a mating between Escherichia coli and Salmonella enterica indirectly selected for mismatch-repair-deficient mutators (Funchain et al., 2001
). On the other hand, the hyper-recombination phenotype of mutators might frustrate the process of hitch-hiking by rapidly separating modifier and effect. However, the hyper-recombination phenotype of such mutators particularly involves recombination between unrelated DNA molecules (Vulic et al., 1997
), and thus will not interfere with the process of hitch-hiking within a clonal population.
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Long-term fate |
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Because a mutator population has limited possibilities to re-establish a low mutation rate, its fate largely depends on the outcome of competition with non-mutator populations. Competition experiments between mutator and wild-type populations have revealed that mutator populations can be competitively superior under certain conditions (Cox & Gibson, 1974 ; Chao & Cox, 1983
; Tröbner & Piechocki, 1984
; Giraud et al., 2001a
). What then determines the competitive ability of a mutator population? Any benefit of a mutator population must rely on its greater adaptability fuelled by a higher production of beneficial mutations. However, faster adaptation of a mutator population is only possible if further adaptation is (i) possible at all, and (ii) limited by the populations supply rate of mutations. As I will show, these premises largely depend on the populations size and adaptedness.
Extinction by mutation accumulation
For the first prerequisite to be met, the population needs to be not well adapted (to allow beneficial mutations), as well as sufficiently large (to produce these rare mutations and to allow efficient selection against deleterious mutations). Once a population becomes too small, it risks the irreversible accumulation of deleterious mutations by genetic drift, a process called Mullers ratchet (Muller, 1964 ; Haigh, 1978
). This risk must be higher for a mutator population. Mullers ratchet depends on the interplay between mutation rate and population size. For example, when mutator (mutS) E. coli populations were passaged through single-cell bottlenecks, Funchain et al. (2000)
observed the extinction of 4% of the lineages after 1000 generations. Similarly, mutator yeast populations (msh2, i.e. defective in a mismatch-repair function, resulting in
30-fold increased mutation rate) which were regularly passaged through bottlenecks of a single cell accumulated deleterious mutations at an increased rate relative to non-mutator populations (Zeyl & de Visser, 2001
; Wloch et al., 2001
). In another study, similar msh2 and wild-type yeast populations were grown in liquid medium by the daily transfer of a constant small volume, containing
25 cells, to fresh medium (Zeyl et al., 2001
). By generation 2900 (after 175 bottlenecks), two of the 12 mutator populations had gone extinct due to the accumulation of deleterious mutations in a mutational meltdown process. A mutational meltdown results from the accelerated accumulation of deleterious mutations caused by their effect of reducing population size (Lynch & Gabriel, 1990
). In contrast, all 12 wild-type populations had retained their initial fitness. These studies affirm that the costs of an elevated mutation rate strongly depend on population size.
Clonal interference
Provided that adaptive evolution is possible (i.e. adaptation is not maximal and the population is sufficiently large), when is adaptation limited by the populations supply of beneficial mutations? The populations supply rate of beneficial mutations (Beneficial Mutation Supply Rate, BMSR) is the product of (i) its supply rate of all mutations (Mutation Supply Rate, MSR; de Visser et al., 1999 ), and (ii) the fraction of mutations that are beneficial. The MSR is simply the product of mutation rate and population size, while the fraction of beneficial mutations depends on the populations level of adaptedness. Thus a mutator population will have a high BMSR if the population is large and not well adapted, and a low BMSR if the population is small and already well adapted. At high BMSR, several beneficial mutants are present in the population simultaneously, all in different genetic backgrounds. Due to the clonal structure of most microbial populations, these contending mutants (Gerrish, 2001
) will then compete with each other, a process that has been dubbed clonal interference (Gerrish & Lenski, 1998
). Clonal interference on the one hand increases the predictability of adaptive evolution by causing: (i) the fixation of beneficial mutations in decreasing order of their fitness effect (i.e. large ones first; Gerrish & Lenski, 1998
; Miralles et al., 1999
), and (ii) a low temporal variation between, or strong rhythm of, fixations (Gerrish, 2001
). On the other hand, it causes the rate of adaptation to increase in a decelerating way with increasing BMSR, leading to an effective speed limit on the rate of adaptive evolution in asexual populations (Gerrish & Lenski, 1998
; de Visser et al., 1999
; Miralles et al., 1999
, 2000
).
Thus large or not well-adapted mutator populations may not adapt faster than similar populations with low mutation rate due to the hampering effect of clonal interference. Only when BMSR is intermediate (e.g. due to relatively small population size) may mutator populations profit from their higher BMSR by adapting faster. Under these conditions, beneficial mutations are extremely rare and mutator populations shorten the waiting time between mutations. For example, pathogens might profit from a mutator phenotype by allowing faster adaptation, because they experience severe bottlenecks during the initial stages of infection (Moxon & Murphy, 1978 ). The above predictions were tested in a laboratory experiment with evolving E. coli populations that differed in mutation rate, population size and initial adaptedness (de Visser et al., 1999
). Consistent with predictions, the results showed diminishing returns from the MSR on the rate of adaptation if the populations founders were not well adapted, and hence BMSR was high. If, however, the founding strains were already well adapted, adaptation appeared indeed limited by the MSR, so that mutator populations would adapt faster. The experiment also showed that well-adapted populations may benefit from an increased mutation rate, but that this benefit may be small. The reasons are that (i) adaptive events have become exceedingly rare and, moreover, (ii) clonal interference causes these later beneficial mutations to be small in effect, because the big-effect mutations will then have been used up (Gerrish & Lenski, 1998
).
Loss of functions needed in future environments
There are yet more limitations to the competitive ability of mutator populations. Their long-term competitive ability may be hindered by the rapid loss of functions, not essential in the present, but important in future environments, if environmental conditions change frequently (Funchain et al., 2000 ; Cooper & Lenski, 2000
; Giraud et al., 2001a
). Many mutations which improve functions needed at present have negative effects on functions that are essential elsewhere, a principle referred to as antagonistic pleiotropy (Cooper & Lenski, 2000
). Other mutations are neutral in the selective environment but show negative effects in secondary environments, and may accumulate by genetic drift (Funchain et al., 2000
). Since mutator populations are thought to accumulate both classes of mutations at increased rate, they should rapidly specialize in exploiting the present environment at the expense of functions acquired previously, a process that has been dubbed genetic amnesia (Giraud et al., 2001b
). Thus rapidly changing environments provide a selective advantage to mutator mutants within populations in the short term, while they may deprive mutator populations of adaptive flexibility in the long run. However, these studies have not been able to attribute the poor long-term performance of mutators to any particular mechanism, i.e. antagonistic pleiotropy, mutation accumulation, or just low BMSR.
Adaptive landscape
A final peculiarity of mutator populations that may affect their long-term fitness has recently been observed in digital organisms evolving in computers (Wilke et al., 2001 ). In this study, pairs of digital organisms with the same ancestor but a fourfold difference in mutation rate were allowed to evolve for 1000 generations by mutation and selection processes similar to those of biological organisms. Several organisms that evolved with a high mutation rate appeared to evolve lower fitness (measured as rate of replication) than their low-mutation-rate equivalent if both populations had a low mutation rate during competition. However, if both populations had a high mutation rate during competition, the organisms that had also evolved with a high mutation rate would win. Apparently, these mutator digital organisms had adapted to lower but flatter fitness peaks in the adaptive landscape (Wright, 1931
), making them more robust to the effects of deleterious mutations at the expense of a lower replication rate (Wilke et al., 2001
). Hence mutator populations not only rapidly adapt to the latest resources they encounter, but they even adapt to the effects of their increased mutation rate itself. Their lower replication rates, however, make them inferior competitors in direct competition with organisms that evolved with lower mutation rates. To what extent these results can be expected in real microbes is an empirical question that begs experimental testing.
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Conclusions |
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Studies addressing the exact short- and long-term costs and benefits of mutators are needed for a more conclusive view on the evolutionary causes and consequences of mutators. What in particular should be done? Little is known about the mutational spectra of general mutators. A comparison between the distribution of mutational fitness effects especially the ratio of beneficial to deleterious mutation numbers of mutators and DNA-repair-proficient strains is fundamental for understanding the basic costs and benefits involved. Recently, protocols have become available that use microbes for this purpose (Imhof & Schloetterer, 2001 ; D. E. Rozen, J. A. G. M. de Visser & P. J. Gerrish, unpublished results). Emphasis should also be given to the long-term evolutionary persistence of mutators. To study their long-term fate, competition experiments over many generations between mutators and wild-type strains should be performed under conditions applied to test the present hypotheses. This way, the long-term fate of mutators in fluctuating or heterogeneous environments versus constant and homogeneous environments can be empirically tested. Finally, by varying the population size in competition experiments, the balance between adaptive and maladaptive evolution can be explored, as well as its impact on the long-term evolutionary fate of mutators.
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ACKNOWLEDGEMENTS |
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