A mechanism for DNA-PK activation requiring unique contributions from each strand of a DNA terminus and implications for microhomology-mediated nonhomologous DNA end joining

Katherine S. Pawelczak, John J. Turchi

Research output: Contribution to journalArticle

21 Scopus citations

Abstract

DNA-dependent protein kinase (DNA-PK) is an essential component of the nonhomologous end joining pathway (NHEJ), responsible for the repair of DNA double-strand breaks. Ku binds a DSB and recruits the catalytic subunit, DNA-PKcs, where it is activated once the kinase is bound to the DSB. The precise mechanism by which DNA activates DNA-PK remains unknown. We have investigated the effect of DNA structure on DNA-PK activation and results demonstrate that in Ku-dependent DNA-PKcs reactions, DNA-PK activation with DNA effectors containing two unannealed ends was identical to activation observed with fully duplex DNA effectors of the same length. The presence of a 6-base single-stranded extension resulted in decreased activation compared to the fully duplex DNA. DNA-PK activation using DNA effectors with compatible termini displayed increased activity compared to effectors with noncompatible termini. A strand orientation preference was observed in these reactions and suggests a model where the 3′ strand of the terminus is responsible for annealing and the 5′ strand is involved in activation of DNA-PK. These results demonstrate the influence of DNA structure and orientation on DNA-PK activation and provide a molecular mechanism of activation resulting from compatible termini, an essential step in microhomology-mediated NHEJ.

Original languageEnglish (US)
Pages (from-to)4022-4031
Number of pages10
JournalNucleic acids research
Volume36
Issue number12
DOIs
StatePublished - Jul 1 2008

ASJC Scopus subject areas

  • Genetics

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