Abstract. All recB-and recC-mutants of E. coli carry out significant residual genetic recombination, whereas all recA -mutants form no recombinants. This observation suggests that an alternative minor pathway of recombination, independent of recB + and recC + products, may be operative in Escherichia coli. Rec+ revertants of recB-recC+, recB+ recC-, and recB-recC-strains of E. coli have been isolated and are shown to fall into at least two major genotypic classes. One class carries revertant mutations which map in or very near the recB and recC genes. In this class an ATP-dependent DNase characteristic of wild type E. coli is restored. The reversions in this class are probably back-mutations or intragenic suppressor mutations. A second class carries revertant mutations which are located far from the recB and recC genes. In this class there is a high level of DNase activity which does hot require ATP and is inactive on T4 DNA. Indirect and not informational suppression appears to be responsible for the second class of revertants. The suggestion is made that restoration of recombination by indirect suppression involves an activation or derepression of one or a series of enzymes, which participate in a pathway of recombination, alternative to the recB and recC pathway, but normally of minor importance. The ATPindependent DNase may be one of these enzymes.Mutations affectingg recombination proficiency in E. coli K12 have been found to occur in at least three genes: recA, recB, and recC.1'2 An ATP-dependent DNase activity has been found in lysates of wild-type strains but is missing in lysates of recB or recC mutant strains.3-5 Tests of a series of merodiploid strains heterozygous or homozygous for recB21 and recC22 indicate that these mutations are responsible for the loss of the ATP-dependent nuclease activity.6 Revertant studies are also desirable to establish this point. Oishi4 has examined one ultraviolet resistant (UVR) revertant which has regained nuclease activity.Our studies of Rec+ revertants are reported here. A preliminary report of our work has been presented.6Two types of reversion are possible: back-mutation at the original site and suppression. One possibility for suppression is suggested by the occurrence of 128

Abstract. An adenosine triphosphate-dependent deoxyribonuclease activity has been detected in lysates of recB+ recC+ strains. Mutations in recB or recC lead to loss of this activity, suggesting that these two genes determine the nuclease activity. The over-all reaction in crude lysates digests native DNA to nucleoside monophosphates. Complementation between recB21 and recC22 in vivo leads to normal levels of ATP-dependent nuclease activity. No complementation in vitro has been detected. Mutations in a third recombination gene (recA) do not alter significantly the wild-type levels of this nuclease activity.Introduction. Recombination-deficient (Rec-) mutants of E. coli have been isolated in several laboratories.1 These mutants are defective in their ability to produce genetic recombinants by conjugation or transduction. A complementation analysis of the recessive rec mutations reveals the existence of three genes or cistrons: recA, recB, and recC. recB and recC lie between argA and thyA2 on the standard genetic map of E. coli.3 A mutation in either of these two rec genes leads to a similar mutant phenotype; namely, reduced but detectable recombination,4 increased sensitivity to ultraviolet (UY) light, and reduced breakdown of DNA following UV irradiation.' recA mutations, on the other hand, are located between pheA and cysC6 on the E. coli map. These mutations result in a drastic reduction in recombination, high sensitivity to UV irradiation, and an increased amount of breakdown following exposure to UV. It appears from these results both recB+ and recC+ might determine a nuclease activity responsible for much

We have studied the growth properties of 17 isogenic strains of Escherichia coli K-12 differing only in the recA, recB, recC, and sbcA alleles. We have observed the following. (i) All recombination deficient strains have decreased growth rates and decreased viabilities compared with recombination proficient strains. The large populations of nonviable cells in Reccultures may arise by spontaneous lethal sectoring (9). (ii) A recA mutant strain which is entirely recombination deficient and which shows high ultraviolet sensitivity and "reckless" deoxyribonucleic acid (DNA) breakdown has approximately the same growth rate and twice the viability as recB and recC mutant strains which have residual recombination proficiency, moderate ultraviolet sensitivity, and "cautious" DNA breakdown. (iii) Indirectly suppressed (sbcA -) recombination proficient (Rec+) revertants of recB and recC mutant strains have approximately normal growth rates and are three times as viable as their Recancestors (but not as viable as rec+ cells). We suggest the following hypothesis to account for the low viability of Rec-E. coli. Single-strand breaks in the DNA duplex, necessary for normal bacterial growth, may be repaired in a Rec+ cell. Failure of Reccells to repair this normal DNA damage may lead to the observed loss of viability. 204 on August 9, 2020 by guest

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