Role of XRCC4 and XLF in NHEJ Resolving how XRCC4 and XLF interact with DNA and what functions

Non-Homologous End Joining (NHEJ) is an efficient mechanism to repair DNA double-strand breaks. XRCC4 and XLF are two structurally-similar core NHEJ proteins. They can directly interact at the protein-protein level and engage DNA by an unknown mechanism. Here, we use optical tweezers and fluorescence microscopy to visualize XRCC4XLF complexes on DNA in real time. We find that the behavior of XRCC4-XLF on DNA is multifaceted: complexes rearrange continuously and show periods of rapid diffusion along the DNA, interspersed by stationary binding. By manipulating two DNA molecules independently we study bridge formation and demonstrate that XRCC4-XLF complexes can directly bridge two independent DNA molecules. Our approach reveals the transient and dynamic aspects of the interaction of XRCC4-XLF with DNA providing new insights on how they could function during DNA double-strand break repair by NHEJ.