The genetic requirements for adaptive mutation in Escherichia coli parallel those for homologous recombination in the RecBCD pathway. Recombination-deficient recA and recB null mutant strains are deficient in adaptive reversion. A hyper-recombinagenic recD strain is hypermutable, and its hypermutation depends on functional recA and recB genes. Genes of subsidiary recombination systems are not required. These results indicate that the molecular mechanism by which adaptive mutation occurs includes recombination. No such association is seen for spontaneous mutation in growing cells.

Adaptive reversion of a +1 frameshift mutation in Escherichia coli, which requires homologous recombination functions, is shown here to occur by -1 deletions in regions of small mononucleotide repeats. This pattern makes improbable recombinational mechanisms for adaptive mutation in which blocks of sequences are transferred into the mutating gene, and it supports mechanisms that use DNA polymerase errors. The pattern appears similar to that of mutations found in yeast cells and in hereditary colon cancer cells that are deficient in mismatch repair. These results suggest a recombinational mechanism for adaptive mutation that functions through polymerase errors that persist as a result of a deficiency in post-synthesis mismatch repair.

Postsynthesis mismatch repair is an important contributor to mutation avoidance and genomic stability in bacteria, yeast, and humans. Regulation of its activity would allow organisms to regulate their ability to evolve. That mismatch repair might be down-regulated in stationary-phase Escherichia coli was suggested by the sequence spectrum of some stationary-phase (''adaptive'') mutations and by the observations that MutS and MutH levels decline during stationary phase. We report that overproduction of MutL inhibits mutation in stationary phase but not during growth. MutS overproduction has no such effect, and MutL overproduction does not prevent stationary-phase decline of either MutS or MutH. These results imply that MutS and MutH decline to levels appropriate for the decreased DNA synthesis in stationary phase, whereas functional MutL is limiting for mismatch repair specifically during stationary phase. Modulation of mutation rate and genetic stability in response to environmental or developmental cues, such as stationary phase and stress, could be important in evolution, development, microbial pathogenicity, and the origins of cancer.[Key Words: Mismatch repair; MutL; mutation; adaptive mutation; genetic instability; evolution; stationary phase]Received April 4, 1997; revised version accepted July 18, 1997.In Escherichia coli mismatch repair is the single largest contributor to avoidance of mutations due to DNA polymerase errors in replication (Radman 1988;Modrich 1991). Mismatch repair also promotes genetic stability by editing the fidelity of genetic recombination and transposon excision, and by the involvement of its component proteins in transcription-coupled DNA repair and very-short-patch repair (Radman 1988;Modrich 1991;Lieb and Shehnaz 1995;Mellon and Champe 1996). The mismatch repair proteins are highly conserved throughout evolution and appear to play roles in simple and complex eukaryotes similar to those that they play in bacteria (Reenan and Kolodner 1992;Modrich 1994;Baker et al. 1995Baker et al. , 1996de Wind et al. 1995;Datta et al. 1996;Hunter et al. 1996;Kolodner 1996). These proteins act on incorrectly paired and unpaired bases in DNA that arise via DNA synthesis errors, recombination of diverged sequences, and DNA damage. In all of these circumstances, mismatch repair enforces genetic stability. The consequences of failing to maintain this enforcement are profound for speciation (Rayssiguier et al. 1989;Radman and Wagner 1993;Matic et al. 1995;Zahrt and Maloy 1997) and for formation of cancers (Modrich 1994(Modrich , 1995Kolodner 1996).Four proteins play critical roles in mismatch repair in E. coli (for review, see Modrich 1991). MutS binds to DNA base mismatches, to insertion/deletion singlestrand loops of four or fewer nucleotides, which are intermediates in frameshift mutation, and probably also to sites of DNA damage (Mello et al. 1996). MutL interacts with MutS after mismatch binding and is thought to coordinate MutS with MutH. MutH endonuclease then nicks the unmethylated (new) DNA s...

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The frequency of somatic hypermutations of an Ig kappa transgene with an artificial test insert, RS, is at least 4-fold higher than that of three related transgenes. The four transgenes differ only in the sequence of a 96 bp insert within the variable region. RS is hypermutable over the total 625 nucleotides of the variable/joining region. The RS insert contains two CAGGTG sequences, potential binding sites for basic helix-loop-helix proteins. Changing CAGGTG to AAGGTG reduces the mutability to that of the non-RS transgenes without altering the mutation pattern. The CAGGTG motif enhances somatic hypermutation without enhancing transcription. A DNA probe containing the two CAGGTG sites, but not AAGGTG, binds E47 and gives rise to two specific EMSA bands with nuclear extracts from mutating cells. Possible actions of this enhancer of somatic hypermutation are discussed.