title: The deinococcal DdrB protein is involved in an early step of DNA double strand break repair and in plasmid transformation through its single-strand annealing activity.
abstract: The Deinococcus radiodurans bacterium exhibits an extreme resistance to ionizing radiation. Here, we investigated the in vivo role of DdrB, a radiation-induced Deinococcus specific protein that was previously shown to exhibit some in vitro properties akin to those of SSB protein from Escherichia coli but also to promote annealing of single stranded DNA. First we report that the deletion of the C-terminal motif of the DdrB protein, which is similar to the SSB C-terminal motif involved in recruitment to DNA of repair proteins, did neither affect cell radioresistance nor DNA binding properties of purified DdrB protein. We show that, in spite of their different quaternary structure, DdrB and SSB occlude the same amount of ssDNA in vitro. We also show that DdrB is recruited early and transiently after irradiation into the nucleoid to form discrete foci. Absence of DdrB increased the lag phase of the extended synthesis-dependent strand annealing (ESDSA) process, affecting neither the rate of DNA synthesis nor the efficiency of fragment reassembly, as indicated by monitoring DNA synthesis and genome reconstitution in cells exposed to a sub-lethal ionizing radiation dose. Moreover, cells devoid of DdrB were affected in the establishment of plasmid DNA during natural transformation, a process that requires pairing of internalized plasmid single stranded DNA fragments, whereas they were proficient in transformation by a chromosomal DNA marker that integrates into the host chromosome through homologous recombination. Our data are consistent with a model in which DdrB participates in an early step of DNA double strand break repair in cells exposed to very high radiation doses. DdrB might facilitate the accurate assembly of the myriad of small fragments generated by extreme radiation exposure through a single strand annealing (SSA) process to generate suitable substrates for subsequent ESDSA-promoted genome reconstitution.
authors.affiliation: Univ. Paris-Sud 11, CNRS UMR 8621, LRC CEA 42V, Institut de Génétique et Microbiologie, Bâtiment 409, Université Paris-Sud, 91405 Orsay, France.
authors.forename: Claire
authors.initials: C
authors.lastname: Bouthier de la Tour
authors.forename: Stéphanie
authors.initials: S
authors.lastname: Boisnard
authors.forename: Cédric
authors.initials: C
authors.lastname: Norais
authors.forename: Magali
authors.initials: M
authors.lastname: Toueille
authors.forename: Esma
authors.initials: E
authors.lastname: Bentchikou
authors.forename: Françoise
authors.initials: F
authors.lastname: Vannier
authors.forename: Michael M
authors.initials: MM
authors.lastname: Cox
authors.forename: Suzanne
authors.initials: S
authors.lastname: Sommer
authors.forename: Pascale
authors.initials: P
authors.lastname: Servant
mesh_terms: D021581:Active Transport, Cell Nucleus / Q000528:radiation effects; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002467:Cell Nucleus / Q000378:metabolism / Q000528:radiation effects; D053903:DNA Breaks, Double-Stranded / Q000528:radiation effects; D053938:DNA Fragmentation / Q000528:radiation effects; D004260:DNA Repair / Q000235:genetics* / Q000528:radiation effects; D004269:DNA, Bacterial / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism; D004277:DNA, Single-Stranded / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D034301:Deinococcus / Q000235:genetics / Q000378:metabolism* / Q000528:radiation effects; D009154:Mutation; D010957:Plasmids / Q000235:genetics*; D017434:Protein Structure, Tertiary; D011836:Radiation Tolerance / Q000235:genetics; D013997:Time Factors; D014169:Transformation, Bacterial / Q000528:radiation effects
chemical_list: D001426:Bacterial Proteins; D004269:DNA, Bacterial; D004277:DNA, Single-Stranded