Journal
NUCLEIC ACIDS RESEARCH
Volume 37, Issue 2, Pages 482-492Publisher
OXFORD UNIV PRESS
DOI: 10.1093/nar/gkn957
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Funding
- Medical Research Council
- Association for International Cancer Research
- Department of Health, European Community Integrated project DNA repair [LSHG-CT-2005-512113]
- European Union RiscRad [FI6R-CT-2003-508842-]
- Alberta Heritage Foundation for Medical Research
- MRC [G0500897] Funding Source: UKRI
- Medical Research Council [G0300662B, G0500897] Funding Source: researchfish
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XLF-Cernunnos (XLF) is a component of the DNA ligase IVXRCC4 (LX) complex, which functions during DNA non-homologous end joining (NHEJ). Here, we use biochemical and cellular approaches to probe the impact of XLF on LX activities. We show that XLF stimulates adenylation of LX complexes de-adenylated by pyrophosphate or following LX decharging during ligation. XLF enhances LX ligation activity in an ATP-independent and dependent manner. ATP-independent stimulation can be attributed to enhanced end-bridging. Whilst ATP alone fails to stimulate LX ligation activity, addition of XLF and ATP promotes ligation in a manner consistent with XLF-stimulated readenylation linked to ligation. We show that XLF is a weakly bound partner of the tightly associated LX complex and, unlike XRCC4, is dispensable for LX stability. 2BN cells, which have little, if any, residual XLF activity, show a 3-fold decreased ability to repair DNA double strand breaks covering a range of complexity. These findings strongly suggest that XLF is not essential for NHEJ but promotes LX adenylation and hence ligation. We propose a model in which XLF, by in situ recharging DNA ligase IV after the first ligation event, promotes double stranded ligation by a single LX complex.
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