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Perspectives Copyright Ó 2006 by the Genetics Society of America Perspectives Anecdotal, Historical and Critical Commentaries on Genetics Edited by James F. Crow and William F. Dove Chaos and Order in Spontaneous Mutation John W. Drake1 Laboratory of Molecular Genetics, National Institute of Environmental Health Sciences, Research Triangle Park, North Carolina 27709-2233 HIS story starts with one of those unanticipated vided by the NDV protocols. I quickly found that phages T turning points that mark the lives of most of us. produced interesting results at least an order of magni- Around 1960 I was working at the University of Illinois at tude more frequently than did NDV. Urbana using Newcastle disease virus (NDV), a tractable Phages and antimutators: There followed a burst of animal virus that infects chickens rather than humans. investigations into the mutation process in phage T4. A (Working previously with polioviruses, I had acquired key component of these investigations was reversion extraordinarily high serum titers against all three strains.) analysis of rII mutations, which tended to reveal that The NDV plaque assay required that chicken embryos of the mutants I was making—not only with UV, but also the correct age be harvested and dissociated into single arising spontaneously in free phages and induced by cells, which were then used to produce monolayers photosensitizers—contained several different kinds of that became confluent in a few days. Virus samples were mutations. Here, however, a barrier arose. Transitions adsorbed to the monolayers, which were then incubated of both types could be recognized, as well as many frame- for a few more days while plaques formed. The regimen shift mutations (which could be reverted by proflavin), was cumbersome and contained day-long gaps that, as it but no agent was known to induce only transversions. turned out, were incompletely filled with the planning The answer, I hoped, might lie in an altogether different and analysis of experiments, the preparation and de- kind of mutagen. Joe Speyer (1965) had just reported livery of lectures, and the miscellaneous duties of a that two temperature-sensitivity mutations in T4 gene 43 bottom-rung academic. (which encodes the viral DNA polymerase) were also As a graduate student I had followed the antics of the strong mutator mutations. Perhaps some such mutators Caltech phage group and had also become aware of the would make transversions specifically. To that end, I ac- mutagenic base analog 5-bromouracil. Over the next quired the entire Caltech collection of gene-43 mutants. few years I was increasingly impressed by the early work Elizabeth ‘‘B.J.’’Allen hunted among these for mutators on mutation by Ernst Freese and Seymour Benzer using and found many. Unexpectedly, she found others that phage T4. Ernst in particular was dissecting out muta- seemed to be antimutators.2 These were much more tional pathways with mutagens that seemed to be highly interesting, so we quickly changed course. specific in their actions, for instance, the base analogs Our first report on antimutators was at a Cold Spring 5-bromouracil and 2-aminopurine that induced transi- Harbor Symposium (Drake and Allen 1968) and con- tions in both directions and hydroxylamine that in- tained a formal description of the antimutators plus duced only GÁC / AÁT transitions. I often had occasion some general observations based on the literature. One to UV irradiate NDV, and I wondered if the mutagenic of the latter was that the phages T4 and l and the bac- specificity of UV irradiation could be ascertained by teria Escherichia coli and Salmonella typhimurium shared performing reversion tests on UV-induced mutations similar genomic mutation rates of roughly 0.2% per with base analogs, hydroxylamine, and proflavin, the chromosome replication. In a huge leap, I wrote that specificity of the last having been demonstrated by ‘‘This result suggests that mutation rates are usually Benzer and Sydney Brenner. Starting with a protocol developed by Raymond Latarjet for phage T4, I began to intercalate phage experiments into the free days pro- 2 I write ‘‘seemed to be’’ because most of the initial antimutator candidates were selected because of deleterious effects of the gene-43 mutations that rendered the mutantsthatAllenwasscoringdifficultto 1Author e-mail: [email protected] detect, a frequent artifact in the history of mutation research. Genetics 173: 1–8 (May 2006) 2 J. W. Drake about as large as can be tolerated’’ (p. 339). It was only pair, a lysogenic phage (whose rate, however, was based several years later that I discovered an important article on its lytic cycle), a bacterium, a yeast, and a filamentous by A. H. Sturtevant (1937) and contemporary (1960s) fungus. Why should such diversity not lead to different work by Motoo Kimura, addressing both the issue of optimal rates in such different microbes? I concluded downward pressures on mutation rates and the limits of that the balancing forces must be very deep, but they such reductions. remain mysterious to this day. When the formal antimutator story was published soon In an effort to expand the range of DNA microbes thereafter, it was divided (by the journal editor) into two exhibiting this standard rate, more examples were slowly parts: one an article about the T4 work (Drake et al. added to the table. The most recent was for a DNA virus 1969) and the other a short note (Drake 1969) on that is also a human pathogen, herpes simplex virus comparative genomic mutation rates in diverse organ- (Drake and Hwang 2005). The most bizarre was for isms. The note presented the phage/bacteria rate con- an archaeon adapted to growth in hot acid, Sulfolobus cordances but also cited a rate about tenfold lower for acidocaldarius (Grogan et al. 2001). Both organisms Neurospora crassa (which eventually turned out to be an display typical genomic rates, but the Sulfolobus rate of underestimate) and a value for Drosophila melanogaster 0.0018 may have been slightly lower than usual for that must have confused many readers because it was a reasons to be discussed later. rate (of about 0.8) per sexual generation rather than per Rates per gene and per base pair: Half a century of germ-line cell division. The same table and its legend work in many laboratories has revealed such a plethora allowed astute readers to estimate that the rate per germ- of mechanisms that it is now abundantly clear that line cell division was about 1.4%. Although this number mutagenesis reflects the sum of many ways in which a was based on several components now known to be complex consortium of fidelity processes can go wrong. inaccurate, the result was close to a later calculation This being the case, it is unlikely that any single mu- based on somewhat better information. Not yet having tation could lower the genomic mutation rate very discovered Kimura’s articles on the subject, I wrote that much. I argued thusly against the existence of general ‘‘Mutation rates may be adjusted by natural selection to antimutator mutations of any considerable strength and achieve a balance among the mostly deleterious effects of free of strongly deleterious pleiotropic effects (Drake mutation, the need for variation, and the cost of sup- 1993a) and supported the argument with the experi- pressing mutation’’ (p. 1132). Well, two out of three was mental demonstration that all of the nine antimutators perhaps not too bad for a beginner; Kimura (1960, 1967) in phage T4 gene 43 and one in gene 45 exhibit muta- had already pointed out that selection for downmodifiers tion rates the same as or slightly greater than that of the of mutation rates would be balanced by the cost of wild type when assayed using reporter genes containing further reducing rates and that selection for upmodifiers roughly 3600 bp; clearly, while mutation rates at some of mutation rates hitchhiking with rare advantageous sites were decreased in these antimutators, rates at other mutations was rapidly obviated by recombination. sites were increased. This result had been modestly DNA-based microbes: I soon became attached to the anticipated by some of the values posted by Drake et al. possibility of a universal genomic mutation rate. As (1969) and strongly anticipated by Lynn Ripley when reports appeared offering numerical orts that could she developed the first T4 system for scoring trans- reasonably be converted into genomic mutation rates, versions (Ripley and Drake 1972; Ripley 1975). In fact, my table was adjusted and expanded and published the T4 antimutators all seem to do the same thing: serially in several chapters and reviews in the 1970s reducing the rate of AÁT / GÁC transitions while mod- and 1980s. The data of best quality were those for DNA estly increasing rates of transversions and small indels.3 viruses, bacteria, and fungi, and by the early 1990s I In an extensive series of investigations, Roel Schaaper was able to produce a compilation that showed that all has had a similar experience: antimutators of modest the DNA-based microbes for which data were available effect can be selected in E. coli but they are again rather shared a genomic rate of spontaneous mutation of close pathway specific. The only exception that one would to 0.003 (mean 0.0033; range 0.0019–0.0046, excluding expect would be an organism that already harbors a two formally defined outliers) (Drake 1991). This result mutator mutation, perhaps not known to the investiga- fascinated me because it implied that the action of tor; reversion or suppression of that specific defect powerful evolutionary forces was capable of finely tun- could then produce a strong antimutator effect, but ing the rate of spontaneous mutation. Both Kimura and would simply lower the rate toward, but not below, the I had imagined a mutation rate balanced between the standard value.
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