Activity of DNA ligase IV stimulated by complex formation with XRCC4 protein in mammalian cells

Mutation of the XRCC4 gene in mammalian cells1,2 prevents the formation of the signal and coding joints in the V(D)J recombination reaction3, which is necessary for production of a functional immunoglobulin gene, and renders the cells highly sensitive to ionizing radiation4. However, XRCC4 shares no sequence homology with other proteins, nor does it have a biochemical activity to indicate what its function might be2. Here we show that DNA ligase IV (ref. 5) co-immunoprecipitates with XRCC4 and that these two proteins specifically interact with one another in a yeast two-hybrid system. Ligation of DNA double-strand breaks in a cell-free system by DNA ligase IV is increased fivefold by purified XRCC4 and seven- to eightfold when XRCC4 is co-expressed with DNA ligase IV. We conclude that the biological consequences of mutating XRCC4 are primarily due to the loss of its stimulatory effect on DNA ligase IV: the function of the XRCC4–DNA ligase IV complex may be to carry out the final steps of V(D)J recombination and joining of DNA ends.