Proteolysis of the human DNA polymerase epsilon catalytic subunit by caspase-3 and calpain specifically during apoptosis.
Human DNA polymerase epsilon (pol epsilon) normally contains a 261-kDa catalytic subunit (p261), but from some sources it is isolated as a 140-kDa catalytic core of p261. This shortened form possesses normal or somewhat enhanced polymerase activity and its significance is unknown. We report here that caspase-3 and calpain can form p140 from p261 in vitro and in vivo and that during early stages of apoptosis induced in Jurkat cells by staurosporine or anti-Fas-activating antibody, p261 is cleaved into p140 by caspase-3. At later stages, activated calpain might also contribute to this conversion. The sites of cleavage by caspase-3 have been identified, and mutations at these 'DEAD boxes' resulted in cleavage-resistant enzyme. Cleavage at these sites separates the 'N-terminal catalytic core' from the 'C-terminal' regions described for p261. Cleavage does not occur during necrosis or following exposure to H(2)O(2) or methanesulfonic acid methyl ester. p140 is unlikely to be able to functionally replace p261 in vivo, since it does not bind to PCNA or the other pol epsilon subunits.
Liu
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W
Division of Biochemistry and Molecular Biology, 229 Stanley Hall, University of California, Berkeley, CA 94720-3206, USA.
Linn
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S
D000595:Amino Acid Sequence; D017209:Apoptosis; D001665:Binding Sites; D015153:Blotting, Western; D002154:Calpain / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism*; D020134:Catalytic Domain; D002453:Cell Cycle; D004249:DNA Damage / Q000187:drug effects / Q000235:genetics; D053938:DNA Fragmentation; D004257:DNA Polymerase II / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology; D019169:Jurkat Cells; D007700:Kinetics; D008741:Methyl Methanesulfonate / Q000494:pharmacology; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009153:Mutagens / Q000494:pharmacology; D009154:Mutation / Q000235:genetics; D009336:Necrosis; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D011485:Protein Binding; D011499:Protein Processing, Post-Translational; D012045:Regulatory Sequences, Nucleic Acid / Q000235:genetics; D013379:Substrate Specificity; D014162:Transfection; D020798:Two-Hybrid System Techniques
D009153:Mutagens; D010446:Peptide Fragments; D008741:Methyl Methanesulfonate; D006861:Hydrogen Peroxide; D011065:Poly(ADP-ribose) Polymerases; D004257:DNA Polymerase II; C505407:CASP3 protein, human; D002154:Calpain; D053148:Caspase 3; D020169:Caspases