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Perspectives Copyright Ó 2008 by the Genetics Society of America Perspectives Anecdotal, Historical and Critical Commentaries on Genetics Edited by James F. Crow and William F. Dove The Phage Mating Theory, With Lessons for Yeast Geneticists Frank Stahl1 Institute of Molecular Biology, University of Oregon, Eugene, Oregon 97403-1229 HEN physicist Max Delbru¨ck undertook the study sistance to the possibility of genetic recombination with W of phage growth (Ellis and Delbru¨ck 1939), he its implications of sexual reproduction and the variety of anticipated that phage would be the best model for highly evolved stuff that so often goes with it (Delbru¨ck elucidating biological reproduction and mutation, un- and Bailey 1947). complicated by sex (Delbru¨ck 1970). This Perspectives However, Max’s hopes for simplicity were soon chal- traces Max’s attempt to come to grips with realities that lenged again by the results and interpretation of experi- threatened that view, and it considers present-day rel- ments conducted with UV-inactivated phages (Luria evanceforyeastgeneticistsoftwolessonsthatremainfrom 1947; Luria and Dulbecco 1949). The UV experiments his heroic effort. showed that phage particles killed by irradiation could Readers should understand (or recall) that in 1939 cooperate to produce live phage, a trick that was labeled essentially nothing of what we now know about the ‘‘multiplicity reactivation’’ (MR) because this cooper- chemistry of either reproduction or mutation was even ation required that a bacterial cell be infected with two imagined—for nucleic acids, it was ‘‘known’’ only that or more phage particles. Luria and Dulbecco (1949) most of the DNA is in the nucleus and most of the RNA is collected MR data for a range of UV doses and a variety of in the cytoplasm and, for proteins, only that some were multiplicities and found that the data could be fitted to a enzymes and that they were probably the stuff that genes mathematically expressed theory. The infected cells pro- are made of. Furthermore, Max seemed to have little duced live phage as if the only requirement for success hope that the biochemistry of the day would ask the right was that each of n identically sensitive, but functionally questions (Delbru¨ck 1949). (Overheard, one graduate distinct, subunits be represented in the infecting pop- of Max’s phage course to another: ‘‘Would you want your ulation by at least one ‘‘unhit’’ subunit of each type daughter to marry a biochemist?’’) I think Max hoped to (Luria 1947). For T2, n ¼ 25; for T4, n ¼ 15. As long as solve the secret of life using physics and algebra along each type of subunit had a surviving member, reassort- with open communication subject to tough criticism. ment of subunits following their multiplication guaran- teed the production of live phages. The assumption that live, unirradiated phage repro- duced in the same manner was the troublesome feature SOME REALITIES OF PHAGE BIOLOGY of the MR theory. Hence, when Dulbecco (1952), using Max ignored the first sign of sexual complexities in T2 phage, demonstrated that the MR theory as mathe- phage reproduction when he found that cells co- matically formulated was inadequate, Max asserted with isconti elbru¨ck infected with the related phages T2 and T4 yielded evident relief (V and D 1953) that the tahl some phage particles that had inherited characteristics theory was baseless (but see Perspectives by S 1995). from both parents. He titled his article ‘‘Induced mu- On the other hand, the more conventional evidence for tations in bacterial viruses,’’ evidencing stubborn re- genetic recombination was not so easily put down. Hershey and Rotman (1948, 1949), who infected cells jointly with two or more multiply marked strains of T2 (a phage ‘‘cross’’), had earlier demonstrated that phages This article is dedicated to the memory of Charley Steinberg.2 1Author e-mail: [email protected] do, indeed, swap genetic information with each other in 2For a fuller appreciation of Charley Steinberg, see the Perspectives by a more-or-less conventional fashion as shown by some Wu and Lindahl (2001). (very roughly) additive recombinant frequencies. On Genetics 180: 1–6 (September 2008) 2 F. Stahl that occasion, Max wrote, ‘‘This is news that is exciting bacterial cycle of phage growth. The interpretation of principally by the blow it deals to our fond hope of mixed infections thus becomes a problem in population analyzing a simple situation’’ (Delbru¨ck 1949, reprin- genetics.’’ Max proposed that (i) phages mate pairwise, ted in Cairns et al. 1992, p. 14). repeatedly, and at random with respect to genotype; (ii) Additional studies (Doermann and Hill 1953) each mating allows for several break-join exchanges strengthened the view that genetic markers in T-even along linear linkage structures, as in meiosis; and (iii) phage could be assigned map positions on the basis of unlinked markers assort at random in individual mating their linkage relations with each other. It began to look acts. He then incorporated these assumptions into an as though conventional meiotic concepts of crossing over algebraically formulated model that related frequencies of linked markers, perhaps concurrent with random of recombinants in the phage population to the events assortment of a modest number of chromosomes, might in the hypothesized individual matings. Because phages be more-or-less applicable to phage crosses. However, are ‘‘haploid’’ and because Max restricted his algebra to the data from T-even phage crosses (Hershey and crosses with no more than three factors, the derivation, Rotman 1948, 1949; Doermann and Hill 1953) in- though ponderous, was simpler than the earlier treat- dicated that, by several criteria, a phage cross was not the ment by Geiringer (1944), which he used as a guide. simple equivalent of an act of meiosis. Data collected by The model would be deemed successful if it proved to be Visconti, then working in Hershey’s lab at Cold Spring compatible with features 1–5, above. Success would then Harbor, supported that view: allow Max to assume that reproduction, although annoy- ingly contemporaneous with recombination, was not 1. Recombinant frequencies among mature phage demonstrably dependent on it, so that one could ignore particles increased with time of sampling of the recombination when investigating reproduction. population (by ‘‘premature,’’ normal, or ‘‘delayed’’ lysis of the infected culture). The genetic exchanges implied by these observations occurred during a FORMULATING AND TESTING THE MODEL noninfectious ‘‘vegetative’’state, in which the phage Max’s first equations for recombinant frequencies multiplied prior to becoming mature virions. Mat- were for populations in which the matings were syn- uration began midway through the infection cycle chronized. Algebraic legerdemain then yielded equa- and continued until the host lysed. tions for a more realistic population in which matings 2. When the infection involved unequal numbers of two were desynchronized and random in time. For a two- parental types, the frequency of recombinants in the factor cross (considered here for simplicity) the ele- progeny of the cross was sometimes greater than the mentary, random-in-time equation for the frequency of frequency of the minority parent genotype. recombinants was 3. The markers available fell into three linkage groups. By two criteria, markers on the three groups appeared R ¼ 2ð f Þð1 À f Þð1 À eÀmpÞ; to be unlinked: (i) crosses between any two markers where f is the fraction of one of the two parental types from different groups gave about the same frequency in the infecting phage mixture, p is the probability per of recombinants, and (ii) in three-factor crosses, a mating that the two factors will recombine, and m is the marker in one group appeared to assort randomly average number of matings per phage lineage. In a among progeny preselected for being recombinant further development, Max averaged R (from m to m ) within another linkage group. However, crosses be- 1 2 to account for the view that some of the phages in the tween any two of these ‘‘unlinked’’ markers gave ‘‘vegetative pool’’ (known to us now as DNA) become ,50% recombinants, violating the simple, meiotic recombinationally inert by being packaged into heads expectation for unlinked markers. when m ¼ m , while the last to be packaged have enjoyed 4. In crosses involving three linked factors, double cross- 1 an average of m rounds of mating. A full-fledged equa- overs were found in excess of the ‘‘expectation’’; i.e., 2 tion contains, in addition, a factor accounting for cell- coefficients of coincidence were greater than unity to-cell variation in f (Lennox et al. 1953). (‘‘negative interference’’). Testing the theory required that m and p be separately 5. Following infection by three distinguishable geno- estimated. This was done by setting p ¼ 1 for putatively types, phage appeared that had inherited markers 2 unlinked markers; m could then be calculated from the from all three of the parents. observed value of R for those markers. With m in hand, Max recognized that these distinctive features of a values of R for pairs of linked markers could be trans- phage cross were properties of a population that was pro- formed to p-values. When these p-values, presumably ceeding toward linkage equilibrium. He formalized his characteristic of individual matings, were tested, they views in a Genetics article that included Visconti’s data were found to be free of negative interference. The (Visconti and Delbru¨ck 1953). He wrote (p. 6) ‘‘... negative interference that characterized R was thereby one may attempt to explain the genetic findings ... by attributable to the heterogeneity in pairwise mating the idea that mating occurs repeatedly during every intra- experiences embodied in i–iii, above. The theory Perspectives 3 worked! In fact, it accounted for all the features 1–5 reciprocal.
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