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© 2015 The Authors. The Fanconi anemia (FA) pathway is critical for the cellular response to toxic DNA interstrand crosslinks (ICLs). Using a biochemical purification strategy, we identified UHRF1 as a protein that specifically interacts with ICLs invitro and invivo. Reduction of cellular levels of UHRF1 by RNAi attenuates the FA pathway and sensitizes cells to mitomycin C. Knockdown cells display a drastic reduction in FANCD2 foci formation. Using live-cell imaging, we observe that UHRF1 is rapidly recruited to chromatin in response to DNA crosslinking agents and that this recruitment both precedes and is required for the recruitment of FANCD2 to ICLs. Based on these results, we describe a mechanism of ICL sensing and propose that UHRF1 is a critical factor that binds to ICLs. In turn, this binding is necessary for the subsequent recruitment of FANCD2, which allows the DNA repair process to initiate.