DNA Repair 2 (2003) 1361–1369 Altered translesion synthesis in E. coli Pol V mutants selected for increased recombination inhibition Suzanne Sommer a,∗ , Olivier J. Becherel b,1 , Geneviève Coste a , Adriana Bailone a , Robert P.P. Fuchs b a Institut de Génétique et Microbiologie, Bˆ at. 409, Université Paris-Sud, F-91405 Orsay, France b Cancérogénèse et Mutagenèse Moléculaire et Structurale, UPR 9003 CNRS, UPR Conventionnée avec l’Université Louis Pasteur de Strasbourg , ESBS, Bld Sébastien Brant, F-67400 Illkirch, France Received 25 June 2003; accepted 7 August 2003 Abstract Replication of damaged DNA, also termed as translesion synthesis (TLS), involves specialized DNA polymerases that bypass DNA lesions. In Escherichia coli, although TLS can involve one or a combination of DNA polymerases depending on the nature of the lesion, it generally requires the Pol V DNA polymerase (formed by two SOS proteins, UmuD ′ and UmuC) and the RecA protein. In addition to being an essential component of translesion DNA synthesis, Pol V is also an antagonist of RecA-mediated recombination. We have recently isolated umuD ′ and umuC mutants on the basis of their increased capacity to inhibit homologous recombination. Despite the capacity of these mutants to form a Pol V complex and to interact with the RecA polymer, most of them exhibit a defect in TLS. Here, we further characterize the TLS activity of these Pol V mutants in vivo by measuring the extent of error-free and mutagenic bypass at a single (6-4)TT lesion located in double stranded plasmid DNA. TLS is markedly decreased in most Pol V mutants that we analyzed (8/9) with the exception of one UmuC mutant (F287L) that exhibits wild-type bypass activity. Somewhat unexpectedly, Pol V mutants that are partially deficient in TLS are more severely affected in mutagenic bypass compared to error-free synthesis. The defect in bypass activity of the Pol V mutant polymerases is discussed in light of the location of the respective mutations in the 3D structure of UmuD ′ and the DinB/UmuC homologous protein Dpo4 of Sulfolobus solfataricus. © 2003 Elsevier B.V. All rights reserved. Keywords: SOS mutagenesis; E. coli Pol V DNA polymerase; Translesion synthesis; UV-induced base substitution mutagenesis; Bypass polymerase ∗ Corresponding author. Tel.: +33-1-69-15-46-14; fax: +33-1-69-15-78-08. E-mail address: suzanne.sommer@igmors.u-psud.fr (S. Sommer). 1 Present address: Radiation Biology and Oncology Laboratory, Queensland Institute of Medical Research (QIMR), The Ban- croft Centre, P.O. Royal Brisbane Hospital, Brisbane 4029, Qld, Australia. 1. Introduction Despite multiple and efficient DNA repair pro- cesses that have evolved to cope with a large variety of DNA damage events, some lesions escape repair and, as a consequence, represent a possible threat for the DNA replication process. Replication of damaged DNA molecules is achieved via two major strategies: (i) an error-free process called damage avoidance (DA), that involves homologous recombination using 1568-7864/$ – see front matter © 2003 Elsevier B.V. All rights reserved. doi:10.1016/j.dnarep.2003.08.008