Regulation of human polλ by ATM-mediated phosphorylation during non-homologous end joining

DNA Repair (Amst). 2017 Mar;51:31-45. doi: 10.1016/j.dnarep.2017.01.004. Epub 2017 Jan 17.

Abstract

DNA double strand breaks (DSBs) trigger a variety of cellular signaling processes, collectively termed the DNA-damage response (DDR), that are primarily regulated by protein kinase ataxia-telangiectasia mutated (ATM). Among DDR activated processes, the repair of DSBs by non-homologous end joining (NHEJ) is essential. The proper coordination of NHEJ factors is mainly achieved through phosphorylation by an ATM-related kinase, the DNA-dependent protein kinase catalytic subunit (DNA-PKcs), although the molecular basis for this regulation has yet to be fully elucidated. In this study we identify the major NHEJ DNA polymerase, DNA polymerase lambda (Polλ), as a target for both ATM and DNA-PKcs in human cells. We show that Polλ is efficiently phosphorylated by DNA-PKcs in vitro and predominantly by ATM after DSB induction with ionizing radiation (IR) in vivo. We identify threonine 204 (T204) as a main target for ATM/DNA-PKcs phosphorylation on human Polλ, and establish that its phosphorylation may facilitate the repair of a subset of IR-induced DSBs and the efficient Polλ-mediated gap-filling during NHEJ. Molecular evidence suggests that Polλ phosphorylation might favor Polλ interaction with the DNA-PK complex at DSBs. Altogether, our work provides the first demonstration of how Polλ is regulated by phosphorylation to connect with the NHEJ core machinery during DSB repair in human cells.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Ataxia Telangiectasia Mutated Proteins / metabolism*
  • DNA Breaks, Double-Stranded
  • DNA End-Joining Repair*
  • DNA Polymerase beta / chemistry
  • DNA Polymerase beta / metabolism*
  • DNA-Activated Protein Kinase / metabolism*
  • Enzyme Activation
  • Humans
  • Nuclear Proteins / metabolism*
  • Phosphorylation
  • Sequence Alignment

Substances

  • Nuclear Proteins
  • ATM protein, human
  • Ataxia Telangiectasia Mutated Proteins
  • DNA-Activated Protein Kinase
  • PRKDC protein, human
  • DNA Polymerase beta
  • DNA polymerase beta2