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Top of Form Nature ? 2001 Nature Publishing Group Volume 412(6842)? ? ? ? ? ? ?5 July 2001? ? ? ? ? ? ?pp 95-99 Endonuclease G is an apoptotic DNase when released from mitochondria [Letters to Nature] Li, Lily Y.; Luo, Xu; Wang, Xiaodong Howard Hughes Medical Institute Department of Biochemistry, University of Texas Southwestern Medical Center at Dallas, Dallas, Texas 75390, USA Received 20 February; accepted 23 May 2001. Supplementary information is available on Natures World-Wide Web site ( ) or as paper copy from the London editorial office of Nature. Correspondence and requests for materials should be addressed to X.W. (e-mail: xwang@). Outline Methods In vitro assay for DNase activity Purification of the supernatant DNase Protein identification Production of recombinant proteins Immunodepletion, blotting and immunostaining Acknowledgements Graphics Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Nucleosomal fragmentation of DNA is a hallmark of apoptosis (programmed cell death)1, and results from the activation of nucleases in cells undergoing apoptosis. One such nuclease, DNA fragmentation factor (DFF, a caspase-activated deoxyribo-nuclease (CAD) and its inhibitor (ICAD)), is capable of inducing DNA fragmentation and chromatin condensation after cleavage by caspase-3 (refs 2-4). However, although transgenic mice lacking DFF45 or its caspase cleavage site have significantly reduced DNA fragmentation5,6, these mice still show residual DNA fragmentation and are phenotypically normal5-7. Here we report the identification and characterization of another nuclease that is specifically activated by apoptotic stimuli and is able to induce nucleosomal fragmentation of DNA in fibroblast cells from embryonic mice lacking DFF. This nuclease is endonuclease G (endoG), a mitochondrion-specific nuclease that translocates to the nucleus during apoptosis. Once released from mitochondria, endoG cleaves chromatin DNA into nucleosomal fragments independently of caspases. T