On the Molecular Mechanisms of Transposition (Transposable Elements/Rolling-Circle Replication/Replicon Fusion/Nonhomologous Recombination) DAVID J

Proc. Nati Acad. Sci. USA Vol. 78, No. 8, pp. 4858-4862, August 1981 Biochemistry

On the molecular mechanisms of transposition (transposable elements/rolling-circle replication/replicon fusion/nonhomologous recombination) DAVID J. GALAS AND MICHAEL CHANDLER D6partement de Biologie Mol6culaire, Universit6 de Genbve, Genbve, Switzerland Communicated by Martin D. Kamen, May 5, 1981 ABSTRACT We present a model for transposition that allows It is distinct both from the models ofShapiro (3) and Arthur and a choice between cointegrate formation (replicon fusion) and di- Sherratt (4) and from the previous model ofGrindley and Sher- rect transposition. We propose that initiation ofthe process occurs ratt (5). There are strong parallels between the model proposed by invasion of the target DNA by a single-stranded end of the here, the current model for the replication of bacteriophage transposable element. This leads to nicking of one of the DNA 4X174 (10), and ColEl transfer replication (11-13). strands of the target molecule and ligation of this strand to that Reconsideration of Molecular Models for Transposition. of the invading transposon. Transposition then occurs in a pro- The transposable element IS1 does not possess a site-specific cessive way by replication of the element from the invading end recombination system for internal resolution comparable to that into the target site in a looped rolling-circle mode similar to rep- the relative stability lication of phage 4X174 replicative form to viral strand. The ofthe Tn3 system. This is clearly implied by choice between cointegrate formation and direct transposition oc- ofplasmids such as R100 and others that contain more than one curs at the nick-ligation step, which terminates the process. We copy of IS1 in direct repeat (14-17). If IS1 has a site-specific suggest that the choice is determined by the topology ofthe trans- recombination system, it must be very weak or be repressed position enzymes and could be related to whether the element under normal conditions. generates five- or nine-base-pair repeats in the target DNA on Our study oftransposition and cointegrate formation induced insertion. by IS) and several ISJ-flanked transposons (to be published elsewhere) shows that these elements do induce the formation Transposable elements are discrete pieces of DNA that can in- ofcointegrates at a detectable rate, but we find that these coin- sert at many different, nonhomologous sites in the genomes of tegrates, once isolated, are very stable. In light ofthis stability, prokaryotic and eukaryotic cells. * In addition to the insertion the frequency ofIS)-induced cointegrate formation we observe event itself, they promote deletions and inversions ofDNA seg- is lower than would be required for their proposed role as es- ments and fusion ofreplicons (also called cointegrate formation). sential intermediates in transposition. For recent reviews see refs. 1 and 2. In experiments with the transposon Tn9O3, Grindley and As yet there is no in vitro system for the study of the non- Joyce (18) found stable cointegrates induced by IS903, the homologous recombination events of transposition. Models of flanking element of this transposon. They concluded, for this the molecular mechanisms must therefore rely on the inter- and other reasons, that there may be a basic difference between pretation of in vivo experiments and remain somewhat specu- the mechanisms for transposition ofTn3 and Tn9O3. For another lative. Several attempts have been made to explain the ex- transposon, Tn5, somewhat similar in structure to Tn903, we perimental observations with a systematic model (3-5). To date also find that the cointegrates are quite stable (unpublished re- the best-studied transposable element is the ampicillin-resis- sults). The above observations on IS) compound transposons, tance element Tn3 (6-8). This transposon has a site-specific on Tn903, and on Tn5 suggest that there is an alternative path- recombination system that induces recombination between two way for transposition, not involving cointegrate intermediates. Tn3s. Shapiro (3) and subsequently Arthur and Sherratt (4), These transposons appear to be related on the basis ofthe above adopting Tn3 as a general paradigm, have suggested that re- considerations, and to be different from Tn3. It is striking that combination between elements is essential to transposition (9). they all produce 9-base-pair repeats in the target DNA on in- These authors proposed very similar general models for trans- sertion, and that they are also similar in having independently position, in which a required intermediate step is the fusion of transposable elements at either end. In contrast, the Tn3-like the donor and recipient replicons. This composite structure elements (including Tn) and Tn2, closely related to Tn3, -yO, contains two directly repeated copies of the transposon joining Tn501, Tn551, and Tn) 721, all ofwhich exhibit some similarity the original replicons and must be resolved by recombination in their DNA sequences) produce 5-base-pair repeats on inser- into two replicons, both now containing a copy of the transposon. tion and have been observed to transpose only as complete While it is clear that cointegrate formation and site-specific units. The model proposed here provides an explanation for the recombination do play important roles in the genetic rearrange- observations cited above and suggests the possibility that the ments induced by transposons, recent findings with the ele- dichotomy among transposons has its basis in the enzyme-spe- ments ISJ, Tn9O3, and Tn5 indicate that their transposition can- cific preference for one of two pathways. not be explained entirely by cointegrate formation followed by Description ofthe Model. In the present model transposition resolution (see next section). In this paper we present a simple proceeds in three basic steps: (i) initiation of transposition by molecular model that takes into account recent data and some a nick-ligation event (joining ofa DNA strand ofone end ofthe ofthe previous proposals. This model can accomodate all known insertion element with a target strand), (ii) semiconservative properties of the transposition process, including the apparent replication ofthe element itselfin a looped rolling-circle mode, dichotomy between the pathway for Tn3 and ISI-like elements. Abbreviations: IS, insertion sequence; Tn, transposon. The publication costs ofthis article were defrayed in part by page charge * These DNA units have been called insertion sequences (IS), trans- payment. This article must therefore be hereby marked "advertise- posable elements, and transposons (Tn). We will not make any dis- ment" in accordance with 18 U. S. C. §1734 solely to indicate this fact. tinction among these terms here. 4858 Downloaded by guest on October 2, 2021 Biochemistry: Galas and Chandler Proc. Natl. Acad. Sci. USA 78 (1981) 4859 and (iii) termination of replication accompanied by a second sition to the action of the 4X174 gene A protein has been nick-ligation event at the other end of the element. pointed out by Grindley and Sheratt (5), Shapiro (3), and Arthur An overall picture of the process is shown in Fig. 1. The im- and Sherratt (4). We suggest that the analogy may extend fur- portant features to note at this level ofdescription are: first, the ther than has been imagined and may include replication. process has two possible outcomes, the fusion of the replicons (b) Replication. The situation after the first double nick and or the direct transposition of the element from one replicon to ligation step is shown in Fig. 2a. We require that the target the other, and second, the replication fork of the transposon DNA, now with an appended strand, be bound by the protein remains anchored at the target site in the recipient replicon (or protein complex) and remain bound until the end of repli- throughout replication, resulting in a looping-out of one copy cation. The two strands of the transposon can now form the ofthe replicating transposon. Each ofthese steps could involve nucleus of a replication fork (Fig. 2b) onto which the protein several molecular events with several possible variations. We complex for DNA replication can condense. The simplest as- will discuss some of these possibilities below. First, however, sumption is that the primer for lead-strand synthesis on the we will describe the model in more detail. nascent replication fork is provided by the 3' end of the donor (a) Initiation. Initiation requires the nicking of both the end molecule originallyjoined to the end ofthe transposon (Fig. 2b). of the transposon and the target molecule, and the ligation of As replication proceeds the donor molecule must move with the two strands to produce a novel single-stranded joint (Figs. respect to the target site-protein complex, which remains la and 2a). This step is, of course, necessary in any model for bound to (or part of) the replication fork itself. transposition. The nicking and ligating reactions could be con- The replication fork can then proceed normally, with lag- certed or occur successively. Possible mechanisms for the se- strand synthesis occurring as in normal semiconservative rep- lection ofthe site in the target molecule are considered in a later lication. Fig. 2b shows the result of the constraints described section. above. One of the daughter molecules of the transposable ele- The possible similarity of this nick ligation step in transpo- ment, the one with an end joined to the target, must loop out progressively during its replication. The target-ligated end would be at one end of the loop, and the growing duplex end, the lag strand of the replication fork, at the other. We refer to

r this (Fig. lb) as a looped rolling-circle mode (19). We propose a~~~~~~~~~Eil that this target site-replication fork complex continues to func- I= =. tion in semiconservative replication until the other end of the transposable element enters the complex, (Fig. .1c and 2c).

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X~~~~~ D FIG. 1. Interreplicon transposition pathways. An overall view of the model for transposition and cointegrate formation is shown in this COINTEGRATE FORMATION DIRECT TRANSPOSITION series of drawings. The initial transposon isrepresented asthe hatched section of the donor plasmid, with the ends labeled A and B. The target FIG. 2. Details of the transposition model. This figure shows the site is indicated in the first drawing as a small segment of the recipient steps of the model in more detail, illustrating the fate of each DNA plasmid flanked by labels C and D. The three basic steps are indicated strand. The strands of the transposon are indicated as in Fig. 1; the in the drawings labeled a, b, and c. Strands of DNA synthesized during donor plasmid strands are shaded in this figure for added clarity. In the process ofreplication are indicated as black strips. The two possible d and e the circled number indicates nicks to be made in the donor final states are shown infandg, labeled to correspond to the drawings strand and the heavy, curved arrow shows the ligation to be made. In of Fig. 2. The model steps are explained in the text. fand g the arrows point out this newly formed joint. Downloaded by guest on October 2, 2021 4860 Biochemistry: Galas and Chandler Proc. Nad'Acad. Sci.- USA 78 (1981) The initiation of replication we envision here is similar to 4X174 replicative to viral form replication (although replication of 4X174 involves no lagging strand synthesis). In our model, however, the protein complex is bound to another duplex DNA molecule (the target) as well'as to the replicating DNA strand, whereas for 4X174 only the single-stranded end remains bound to the gene A protein. For simplicity we have assumed that lead- strand synthesis occurs on the donor molecule, and lag-strand on the looped-out copy. Replication could occur, however, with the opposite polarity. Other important features of replication initiation remain unspecified: for example, whether there is a significant delay in lag-strand initiation, or whether single- strand-binding protein or RNA primers are involved. (c) Termination. In Fig. 2c we have depicted the situation as the trailing end of the transposon enters the replication complex. At this point three outcomes are possible. Termination could occur in one of two ways, leading to the configurations indicated at the bottom of Figs. 1 and 2, or termination could fail to occur and replication continue into the adjacent DNA. Termination requires a second nick-ligation reaction at the trailing end ofthe element. The target molecule must be nicked FIG. 3. Symmetry of the initiating (a) and terminating (b and c) on the strand opposite the initial nick'in the target site, at a ligation reactions. Here the strands of DNA are indicated by the con- distance from the first nick equal to the size ofthe repeat formed vention used in Fig. 2, and the hypothetical enzyme with two subunits on insertion (nine base pairs for ISI, Tn5, Tn903, TnlO, etc., is shown centered on the target site. The arrows in each ofthe drawings and five base pairs for Tn3, y8, Tn501, Tn551, etc . .. , see show the nicks required to make the ligations, which are indicated by ref. 2). A nick must also be made at the end of the insertion the broken lines. The symmetries are described in the text. element and the resulting ends must be ligated. In Fig. 2c we have indicated two possible ways that this nick could be made, DNA molecules in the figure. The product of the ligation cat- using one or the other of the branches of the replication fork. alyzed in this step is indicated by the broken lines. In the lower The corresponding alternative ligations are indicated in Fig. 2 two drawings (Fig. 3 b and c) we have illustrated the relative d and e. The position of the nick in each case is marked by the orientation of the signal sequence to the other subunit of the arrows, 1 and 2. Note that once the nicks are made the sub- protein, showing how it could be the same in the termination strates for ligation are essentially identical. The outcome of the complex as in Fig. 3a. In these two drawings the replication overall process is determined by which branch is chosen to par- forks are positioned so that the ligation to be performed by sub- ticipate in this reaction. If the branch marked 1 is chosen a re- unit 2 (indicated by broken lines), and the DNA molecules in- plicon fusion results (Fig. 2f), and if the branch marked 2 is volved, form the mirror image of the ligation by-subunit 1 in chosen a direct transposition of the element from one replicon Fig. 3a. The point of this exercise is to illustrate the symmetry to the other is accomplished (Fig. 2g). The preference for one between the events initiating and terminating transposition. choice or the other is probably built into the enzymes of the Thus it is quite plausible that the same protein, capable ofrecog- transposition apparatus and is therefore specific for each nizing the sequence of the ends of the element, catalyzes both element. nick-ligation reactions and remains bound to the target DNA The final structures shown at the bottom of Fig. 2 (Fig. 2f throughout the entire process. and g) require filling in ofthe gaps and ligation to complete the The symmetry between the initial and final nick-ligation also process. These steps could be accomplished by the host cell's extends to the two branches of the replication fork: either one repair system. of them may be involved in the final nick-ligation. The simi- A key element of the termination process is the DNA se- larities between the ends ofthe transposons on each branch are quence signal provided by the end ofthe transposon as it enters evident on comparing Fig. 3b with Fig. 3c (also see Fig. 2c). the replication complex. Because the two ends of most known The single difference between the two branches is that, in one transposons (and flanking insertion sequences) are essentially case, the strand to be ligated has a free 5' end before the nick identical to each other, in inverse orientation (2), the signal se- is made, and in the other case the free end must be generated quence is the same as that recognized and nicked in the initial by the nick. While the nick ligation could, in principle, be car- step. It is a simple matter to picture a dimeric protein (for ex- ried out on either branch of the fork, the two could be distin- ample), with subunits arranged face to face as shown in Fig. 3, guished by an enzyme so that one or the other is likely to be in such a way that the same protein that catalyzed the initial preferred by the transposition apparatus. We suggest that it is nick-ligation catalyzes the nick-ligation that terminates the pro- just this distinction between the participation of the two cess, and in response to the same signal sequence in the op- branches that is responsible for the dichotomy between classes posite orientation. of transposons: those that carry out the final step as shown in Although the specific proposal set forth in Fig. 3 is not es- Fig. 3b (the leftward pathway in Fig. 2) would produce a coin- sential to the model, we think it is useful in making the model tegrate, and those that carry out the final step as in Fig. 3c as concrete as possible. In Fig. 3a we have represented the ini- (rightward pathway in Fig. 2) would produce a direct tial nick-ligation event, showing the orientation of the end se- transposition. quence of the element (this is marked SIGNAL in reversed let- Once a cointegrate is formed it may be resolved by a recom- ters) and a hypothetical dimer of a transposition protein. Bear bination event between two copies of the transposon (Fig. if). in mind that the parallel, linear representation of the DNA This was pointed out by Gill et aL (20), Shapiro (3), and Arthur strands in this figure is for clarity only, and that there is no to- and Sherratt (4) and was proposed as an essential step in trans- pological significance to the "upper" and "lower" strands of the position. Indeed, Tn3 has a site-specific recombination system Downloaded by guest on October 2, 2021 IBiochemistry: Galas and Chandler Proc. Natl. Acad. Sci. USA 78 (1981) 4861

that serves the purpose of resolving cointegrates. If the transposition process were to generate cointegrates as a major product, such a recombination system would be essential to complete transposition. We propose therefore that transposons of the class that have site-specific recombination systems favor the branch choice leading to cointegrate formation. The specificity in the molecular event determiningthe choice ofpathway (Fig. 2c) may be related to the distance between the staggered nicks at the target site. For the five-base-pair repeating elements the nicks are five base pairs apart (on opposite strands) and are therefore on the same side ofthe double helix. For the nine-base-pair repeating elements, on the other hand, RECOMPLETED ' the nicks must be on opposite sides of the helix. This approxi- REPLI CATION mate relation holds true for all known DNA duplex forms-i.e., OF TnX B form, A, C, Z, and Z' (21-23). The relative spatial position of the nick sites would also determine the initial position ofthe CONTINUES single-stranded end of the target DNA that participates in the final ligation. Because the relative position ofthe reactants must strongly influence the efficacy of the reaction, it is quite possible that the choice of the transposon strand to be ligated to FIG. 4. Model pathways for a compound transposon (a section of the strand is related to the This DNAflankedbytwo transposable elements), showing the effectsof the target intimately repeat length. branching between termination and continued replication and be- effect might explain the fact that. the class of transposons that tween cointegrate formation and direct transposition. See text for de- form and resolve cointegrates rapidly also makes 5-base-pair tails. The possibility that a cointegrate can be resolved by recombi- repeats on insertion. Similar preferences for one branch over nation is shown in parentheses to indicate the observed stability of the other may also be expected to exist for the 4- and 11-base- these structures. In the case of resolution of a cointegrate formed by pair repeating elements, IS3 and IS4, respectively (2). duplication ofTnX, there are several possible ways thatresolution can Site Selection. For many elements, transposition is charac- occur. These pathways are not shown. terized by a low level of target specificity. There are, however, wide variations from element to element in the nature and de- viously suggested a processive mechanism for transposition on gree of this specificity. Although the model proposed here is the basis of the observation that transposition frequencies of a not intended to explain site selection, there are several plausible series of IS1-flanked compound transposons are inversely re- mechanisms that are fully compatible with the model. If the lated to their size (33). transposon is nicked in an energy-conserving reaction by a pro- If normal termination occurs at the end of the second ele- tein that remains bound to the end of the strand of transposon ment, the effect would be to transpose all the intervening DNA, DNA, this end becomes a reactive intermediate for ligation to together with its flanking elements. The behavior of a com- another single-stranded end. The gene A protein of 4X174 (10) pound transposon would then involve two separate branching and a relaxation protein of the ColEl plasmid (11-13) are events: the choice between transposition of a flanking element thought to act in just this way. The protein-terminated DNA and the entire compound transposon, and the choice between strands may form a complex with the target DNA molecule and cointegrate formation and direct transposition. In Fig. 4 we unwind the duplex in a reaction analogous to that catalyzed by have diagrammed this doubly branched pathway. Replication the recA protein (24-26). The feature of the target DNA that is represented in the vertical sequence on the left with the favors or even triggers the actual nicking ofthe target strand and branching events leading to the final products on the right. An its ligation to the transposon end may be a partial homology with immediate consequence ofthis view ofa compound transposon the ends ofthe element (27). Another possibility is that a partial is that, as long as the termination event is equally likely at either homology may stabilize the preligation complex, favoring suc- end of an element in the proper orientation, the flanking ele- cessful ligation at sites where such stabilization is possible (27). ments of a compound transposon should transpose more fre- The fact that some transposons, notably Tn9 and Tn5, appear quently than the entire structure. We have some preliminary to favor easily denaturable regions of DNA (27-31) may reflect data indicating that this prediction is fulfilled by several ISI- the ease with which the target duplex is unwound and invaded flanked transposons (unpublished results). Under selection by the transposon strand. It is probably significant that the nick- pressure that favors transposition of the compound transposon ing action ofthe 4X174 gene A protein is also enhanced strongly as a unit, the inside ends of the flanking elements (adjacent to by the presence of an A+T-rich region adjacent to the nick site the intervening DNA segment) might evolve into less effective (32). termination signals than the outer ends and cause the flanking Compound Transposons. As we have indicated above, be- elements to lose some oftheir independence to the larger struc- cause the terminal insertion sequences of the 9-base-pair re- ture (perhaps internal ends could be dispensed with completely peating transposons (Tn9, Tn9O3, Tn5, and TnlO) have been by deletion). New transposons could evolve from smaller ele- shown to behave as transposable elements themselves, these ments by such a process. elements can be considered compound transposons. If the ter- Discussion. The observations referred to in the previous sec- mination events fail to occur at the distal end of the first trans- tions indicate that the original models proposed by Shapiro (3) posable element, the replication fork may continue past the end and Arthur and Sherratt (4) are inadequate to explain the be- ofthe element into the adjoining DNA. Because a second trans- havior of some transposons. These models require both ends posable element lies farther along, termination gets a second ofthe element to come together and be ligated at the target site. chance in response to the signal provided by the distal end of Two potential replication forks would thereby be created. Rep- *this second element (the proximal end would be in the wrong lication would then proceed in either direction, and the product orientation to act as a signal). This readthrough replication could of the completed reaction would be a cointegrate between the explain the properties ofcompound transposons. We have pre- donor and recipient replicons. [Shapiro has recently modified Downloaded by guest on October 2, 2021 4862 Biochemistry: Galas and Chandler Proc. Natl. Acad. Sci. USA 78 (1981)

his model (34) by suggesting that if-recombination between the Clerget, R. Deonier, R. Epstein, B. Hirschel, H. Krisch, J. Miller, P. elements takes place during separation of the replication prod- Prentki, and A. Schmitz. This work was supported by Grant 3.591.79 ucts, mature cointegrate molecules need not be formed.] In to L. Caro and Grant 3.493.79 to J. Miller from the Swiss National Sci- contrast, the model originally proposed by Grindley and Sher- ence Foundation. ratt (5) calls for one end of the transposon to be nicked and ligated to the target, followed by two rounds of displacement 1. Starlinger, P. (1980) Plasmid 3, 241-259. synthesis: up and down the transposon. According to the Grin- 2. Calos, M. P. & Miller, J. H. (1980) Cell 20, 579-595. dley-Sherratt model the transposable element undergoes direct 3. Shapiro, J. A. (1979) Proc. Nati. Acad. Sci. USA 76, 1933-1937. 4. Arthur, A. & Sherratt, D. J. (1979) Mol Gen. Genet. 175, 267- transposition (unless strand switching occurs during displace- 274. ment synthesis). 5. Grindleyv N. D. F. & Sherratt, D. J. (1979) Cold Spring Harbor The model described here differs in several respects from the Symp. Quant. Biol. 43, 1257-1261. foregoing models. The proposed branching pathway can lead 6. Heffron, F., McCarthy, B. J., Ohtsubo, H. & Ohtsubo, E. (1979) either to cointegrate formation or direct transposition. Several Cell 18, 1153-1163. features of the previous models [the Grindley-Sherratt model 7. Ohtsubo, H., Ohmori, H. & Ohtsubo, E. (1979) Cold Spring (5), in particular] have been incorporated into ours, and some Harbor Symp. Quant. Biol. 43, 1269-1277. 8. Heffron, F., Bedinger, P., Champoux, J. J. & Falkow, S. (1977) features suggested by 4X174 replication and transferreplication Proc. Natl. Acad. Sci. USA 74, 702-706. of ColEl. 9. Sherratt, D. J., Arthur, A. & Burke, M. (1981) Cold Spring Har- To explain their electron microscopy data, Harshey and Buk- bor Symp. Quant. Biol 45, 275-281. hari (35) have proposed a roll-in replication model, which has 10. Eisenberg, S. & Denhardt, D. T. (1974) Proc. Natl. Acad. Sci. several features in common with our model. In particular, pro- USA 71, 984-988. 11. Warren, G. J., Twigg, A. J. & Sherratt, D. J. (1978) Nature (Lon- gressive replication from one end followed by strand ligation don) 274, 259-261. could produce either cointegrates or direct transpositions, as 12. Warren, G. J. & Clark, A. J. (1980) Proc. Nati Acad. Sci. USA 77, in our model. Our proposal differs from theirs notably by re- 6724-6728. quiring that the replication fork complex of the transposon re- 13. Broome-Smith, J. (1980) Plasmid 4, 51-63. main anchored at the target site, resulting in the looping-out 14. Ptashne, K. & Cohen, S. N. (1975)J. Bacteriol. 122, 776-781. of one copy of the transposon during replication. This allows a 15. Hu, S., Ohtsubo, E., Davidson, N. & Saedler, H. (1975)J. Bac- second, symmetric, nick-ligation event to occur at the termi- teriol. 122, 764-775. 16. lida, S. & Arber, W. (1980) Mol. Gen. Genet. 177, 261-270. nation ofreplication, in contrast to the Harshey-Bukhari model, 17. Ohtsubo, E., Zenilman, M. & Ohtsubo, H. (1980) Proc. Natl. in which a concerted double-strand cleavage of the target DNA Acad. Sci. USA 77, 750-754. initiates the process. 18. Grindley, N. D. F. & Joyce, C. M. (1981) Cold Spring Harbor The implications of the present model for the induction of Symp. Quant. Biol. 45, 125-133. deletions and inversions by transposons are straightforward. As 19. Kornberg, A. (1980) DNA Replication (Freeman, San Francisco), pointed out by Shapiro (3) and Arthur and Sherratt (4), if the p. 510. 20. Gill, R., Heffron, F., Dougan, G. & Falkow, S. (1978) J..Bacte- target is within the same replicon as the donor transposon, both rioi 136, 742-756. deletion and inversion can be explained by the same processes 21. Drew, H., Takano, T., Tanaka, S., Itakura, K. & Dickerson, R. leading to transposition. In our model the cointegrate-forming E. (1980) Nature (London) 286, 567-573. pathway can induce these rearrangements in just the manner 22. Arnott, S., Chandrasekaran, R., Birdsall, D. L., Leslie, G. W. described by these authors. The other pathway, which leads to & Ratliff, R. L. (1980) Nature (London) 283, 743-745. direct transposition, can also induce rearrangements when the 23. Rhodes, D. & Klug, A. (1980) Nature (London) 286, 573-578. 24. Shibata, T., Das Gupta, C., Cunningham, R. P. & Radding, C. target is in the same replicon as the donor, but only after a re- M. (1980) Proc. Natl. Acad. Sci. USA 77, 2606-2611. combination event between the two copies of the transposon. 25. McEntee, K., Weinstock, G. M. & Lehman, I. R. (1980)-Proc. When the recombination is between transposons in direct re- Natl. Acad. Sci. USA 77, 857-861. peat, a deletion results (7); when they are in inverted repeat, 26. Wilson, J. H. (1979) Proc. Natl. Acad. Sci. USA 76, 3641-3644. an inversion results. 27. Galas, D. J., Calos, M. P. & Miller, J. H. (1980)J. Mol. Biol. 144, The model has suggested a classification of known transpo- 19-41. 28. Federoff, N. V. (1979) Cold Spring Harbor Symp. Quant. Biol. sons into two principal groups, the transposons that favor one 43, 1287-1292. pathway or the other, preferentially favoring cointegrates or 29. Meyer, J., Iida, S. & Arber, W. (1980) Mol. Gen. Genet. 178, 471- direct transpositions. It might be that this criterion is too sim- 473. ple, and that under different conditions (various temperatures, 30. Miller, J. H., Calos, M. P., Galas, D. J., Hofer, M., Buchel, D. for example) the preferences could be altered. With this qual- E. & Muller-Hill, B. (1980)J. Mol. Biol 1-18. 31. Devos, R., Contreras, R., Van Emmelo, J. & Fiers, W. (1979)J. ification, we can summarize our suggestions as follows. The Mol. Biol. 128, 621-632. transposons of the class we have called Tn3-like favor the coin- 32. Van Mansfeld, A. D. M., Langveld, S. A., Baas, P. D., Jansz, H. tegrate-forming pathway and have a recombination system for S., Van der Marel, G. A., Veeneman, G. H. & Van Boom (1980) resolution of cointegrates. Transposons of the class we have Nature (London) 288, 561-566. called IS-like appear to favor the direct transposition pathway, 33. Chandler, M., Clerget, M. & Caro, L. (1981) Cold Spring Har- at least under some conditions, and form stable cointegrates as bor Symp. Quant. Biol 45, 157-165. well. This dichotomy may have its basis in the transposition 34. Shapiro, J. A. (1980) in Plasmids and Transposons, eds. Stod- dard, C. & Rossi, K. (Academic, New York), pp. 229-247. enzymes themselves and be reflected in the five-base-pair or 35. Harshey, R. M. & Bukhari, A. I. (1981) Proc. Natl Acad. Sci. USA nine-base-pair repeat characteristic of each class. In addition, 78, 1090-1094; some transposons may even be facultative in this respect, ca- 36. Coelho, A., Leach, D., Maynard-Smith, S. & Symonds, N. pable of favoring one pathway or the other in response to ex- (1981) Cold Spring Harbor Symp. Quant. Biol 45, 323-328. ternal conditions [phage Mu may be an example (36-38)]. 37. Kamp, D. & Kahmann, R. (1981) Cold Spring Harbor Symp. Quant. Biol 45, 329-336. We thank the following people for useful discussions and criticisms 38. Chaconas, G., Harshey, R. M., Sarvetnick, N. & Bukhari, A. I. ofthe manuscript: M. Calos, L. Caro, G. Churchward, S. Clarkson, M. (1981) Cold Spring Harbor Symp. Quant. BioL 45, 311-322. Downloaded by guest on October 2, 2021