title: Homologous recombination mediated by the mycobacterial AdnAB helicase without end resection by the AdnAB nucleases.
abstract: Current models of bacterial homologous recombination (HR) posit that extensive resection of a DNA double-strand break (DSB) by a multisubunit helicase-nuclease machine (e.g. RecBCD, AddAB or AdnAB) generates the requisite 3' single-strand DNA substrate for RecA-mediated strand invasion. AdnAB, the helicase-nuclease implicated in mycobacterial HR, consists of two subunits, AdnA and AdnB, each composed of an N-terminal ATPase domain and a C-terminal nuclease domain. DSB unwinding by AdnAB in vitro is stringently dependent on the ATPase activity of the 'lead' AdnB motor translocating on the 3' ssDNA strand, but not on the putative 'lagging' AdnA ATPase. Here, we queried genetically which activities of AdnAB are pertinent to its role in HR and DNA damage repair in vivo by inactivating each of the four catalytic domains. Complete nuclease-dead AdnAB enzyme can sustain recombination in vivo, as long as its AdnB motor is intact and RecO and RecR are available. We conclude that AdnAB's processive DSB unwinding activity suffices for AdnAB function in HR. Albeit not excluding the agency of a backup nuclease, our findings suggest that mycobacterial HR can proceed via DSB unwinding and protein capture of the displaced 3'-OH single strand, without a need for extensive end-resection.
authors.affiliation: Immunology Program, Memorial Sloan Kettering Cancer Center, 1275 York Avenue, New York, NY 10065, USA.
authors.forename: Richa
authors.initials: R
authors.lastname: Gupta
authors.affiliation: Molecular Biology Program, Memorial Sloan Kettering Cancer Center, 1275 York Avenue, New York, NY 10065, USA.
authors.forename: Mihaela-Carmen
authors.initials: MC
authors.lastname: Unciuleac
authors.affiliation: Molecular Biology Program, Memorial Sloan Kettering Cancer Center, 1275 York Avenue, New York, NY 10065, USA s-shuman@ski.mskcc.org.
authors.forename: Stewart
authors.initials: S
authors.lastname: Shuman
authors.affiliation: Immunology Program, Memorial Sloan Kettering Cancer Center, 1275 York Avenue, New York, NY 10065, USA glickmam@mskcc.org.|Division of Infectious Diseases, Memorial Sloan Kettering Cancer Center, 1275 York Avenue, New York, NY 10065, USA.
authors.forename: Michael S
authors.initials: MS
authors.lastname: Glickman
mesh_terms: D000251:Adenosine Triphosphatases / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D001426:Bacterial Proteins / Q000378:metabolism; D053903:DNA Breaks, Double-Stranded; D004265:DNA Helicases / Q000235:genetics / Q000378:metabolism*; D004260:DNA Repair; D004720:Endonucleases / Q000378:metabolism; D059765:Homologous Recombination; D006868:Hydrolysis; D018625:Microscopy, Atomic Force; D009154:Mutation; D009161:Mycobacterium / Q000201:enzymology*; D011485:Protein Binding; D011836:Radiation Tolerance / Q000235:genetics
chemical_list: D001426:Bacterial Proteins; D000255:Adenosine Triphosphate; D004720:Endonucleases; D000251:Adenosine Triphosphatases; D004265:DNA Helicases