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Nucleic Acids Research, 2002, Vol. 30, No. 14 3045-3051
© 2002 Oxford University Press

The effect of ICAD-S on the formation and intracellular distribution of a nucleolytically active caspase-activated DNase

Sebastian Richard Scholz, Christian Korn, Oleg Gimadutdinow1, Michael Knoblauch2, Alfred Pingoud and Gregor Meiss*

Institut für Biochemie (FB 08), Justus-Liebig-Universität, Heinrich-Buff-Ring 58, D-35392 Giessen, Germany, 1 Department of Genetics, Kazan State University, Kremlevskaja 18, 420008 Kazan, Russian Federation and 2 Institut für Allgemeine Botanik und Pflanzenphysiologie, Justus-Liebig-Universität, Senckenbergstrasse 17 and 25, D-35390 Giessen, Germany

*To whom correspondence should be addressed. Tel: +49 641 99 35404; Fax: +49 641 99 35409; Email: gregor.meiss{at}chemie.bio.uni-giessen.de

We show here that co-expression of murine CAD with either ICAD-L or ICAD-S in Escherichia coli as well as mammalian cells leads to a functional DFF complex, which after caspase-3 activation releases a nucleolytically active DNase. The chaperone activity of ICAD-S is between one and two orders of magnitude less effective than that of ICAD-L, as deduced from cleavage experiments with different activated recombinant DFF complexes produced in E.coli. With nucleolytically active EGFP fusion proteins of CAD it is demonstrated that co-expression of ICAD-S, which lacks the C-terminal domain of ICAD-L, including the NLS, leads to a homogeneous intracellular distribution of the DNase in transfected cells, whereas co-expression of human or murine ICAD-L variants lacking the NLS leads to exclusion of EGFP–CAD from the nuclei in ~50% of cells. These results attribute a particular importance of the NLS in the long isoform of the inhibitor of CAD for nuclear accumulation of the DFF complex in living cells. It is concluded that ICAD-L and ICAD-S in vivo might function as tissue-specific modulators in the regulation of apoptotic DNA degradation by controlling not only the enzymatic activity but also the amount of CAD available in the nuclei of mammalian cells.


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