XLF Interacts with the XRCC4-DNA Ligase IV Complex to Promote DNA Nonhomologous End-Joining

Abstract

DNA nonhomologous end-joining (NHEJ) is a predominant pathway of DNA double-strand break repair in mammalian cells, and defects in it cause radiosensitivity at the cellular and whole-organism levels. Central to NHEJ is the protein complex containing DNA Ligase IV and XRCC4. By searching for additional XRCC4-interacting factors, we identified a previously uncharacterized 33 kDa protein, XRCC4-like factor (XLF, also named Cernunnos), that has weak sequence homology with XRCC4 and is predicted to display structural similarity to XRCC4. We show that XLF directly interacts with the XRCC4-Ligase IV complex in vitro and in vivo and that siRNA-mediated downregulation of XLF in human cell lines leads to radiosensitivity and impaired NHEJ. Furthermore, we establish that NHEJ-deficient 2BN cells derived from a radiosensitive and immune-deficient patient lack XLF due to an inactivating frameshift mutation in its gene, and that reintroduction of wild-type XLF into such cells corrects their radiosensitivity and NHEJ defects. XLF thus constitutes a novel core component of the mammalian NHEJ apparatus.

Programmed DNA breaks in lymphoid cells: repair mechanisms and consequences in human disease

“…The XRCC4‐like factor (XLF, also called Cernunnos) shares sequence homology and structure similarity with XRCC4 77. Indeed, cells depleted for XLF display increased radiosensitivity and a defect in NHEJ 77.…”

“…Indeed, cells depleted for XLF display increased radiosensitivity and a defect in NHEJ 77. In addition, XLF was suggested to have overlapping functions with H2AX and 53BP1 in the assembly of DSB response factors on chromatin during V(D)J recombination 54…”

DNA double-strand break repair and development

“…Ligation of the DNA ends occurs via Lig4 in conjunction with its binding partner Xrcc4. An additional factor, Cernunnos/ XLF, has been identified as a binding partner of the Lig4-Xrcc4 complex and appears to be necessary for efficient ligation via NHEJ (Ahnesorg et al, 2006;Buck et al, 2006a). Subsets of NHEJ may involve other factors such as Artemis (Moshous et al, 2001;Jeggo and O'Neill, 2002).…”

Non-homologous end-joining, a sticky affair

“…Compatible DNA ends can now be joined directly by the ligase IV/XRCC4 complex. This reaction is stimulated by the recently discovered XLF/Cernunnos protein, which interacts with XRCC4 (Ahnesorg et al, 2006;Buck et al, 2006). However, in many circumstances the DNA ends are not compatible.…”