Efficient translesion replication in the absence of Escherichia coli Umu proteins and 3′-5′ exonuclease proofreading function

Translesion replication (TR) past a cyclobutane pyrimidine dimer in Escherichia coli normally requires the UmuD'C-2 complex, RecA protein, and DNA polymerase III holoenzyme (pol III). However, we find that efficient TR can occur in the absence of the Umu proteins if the 3'-5' exonuclease proofreading activity of the pol III epsilon-subunit also is disabled. TR was measured in isogenic uvrA6 Delta umuDC strains carrying the dominant negative dnaQ allele, mutD5, or Delta dnaQ spq-2 mutations by transfecting them with single-stranded M13-based vectors containing a specifically located cis-syn T-T dimer, As expected, little TR was observed in the Delta umuDC dnaQ(+) strain. Surprisingly, 26% TR occurred in UV-irradiated Delta umuDC mutD5 cells, one-half the frequency found in a uvrA6 umuDC(+)mutD5 strain, lexA3 (Ind(-)) derivatives of the strains showed that this TR was contingent on two inducible functions, one LexA-dependent, responsible for approximate to 70% of the TR, and another LexA-independent, responsible for the remaining approximate to 30%. Curiously, the Delta umuDC Delta dnaQ spq-2 strain exhibited only the LexA-independent level of TR, The cause of this result appears to be the spq-2 allele, a dnaE mutation required for viability in Delta dnaQ strains, since introduction of spq-2 into the Delta umuDC mutD5 strain also reduces the frequency of TR to the LexA-independent level, The molecular mechanism responsible for the LexA-independent TR is unknown but may be related to the UVM phenomenon [Palejwala, V. A., Wang, G. E., Murphy, H. S. & Humayun, M. Z. (1995) J. Bacteriol. 177, 6041-6048]. LexA-dependent TR does not result from the induction of pol II, since TR in the Delta umuDC mutD5 strain is unchanged by introduction of a Delta polB mutation.