Zinc is a potent inhibitor of the apoptotic protease, caspase-3. A novel target for zinc in the inhibition of apoptosis.
The prevention of apoptosis by Zn2+ has generally been attributed to its inhibition of an endonuclease acting in the late phase of apoptosis. In this study we investigated the effect of Zn2+ on an earlier event in the apoptotic process, the proteolysis of the "death substrate" poly(ADP-ribose) polymerase (PARP). Pretreatment of intact Molt4 leukemia cells with micromolar concentrations of Zn2+ caused an inhibition of PARP proteolysis induced by the chemotherapeutic agent etoposide. Using a cell-free system consisting of purified bovine PARP as a substrate and an apoptotic extract or recombinant caspase-3 as the PARP protease, Zn2+ inhibited PARP proteolysis in the low micromolar range. To rule out an effect of Zn2+ on PARP, a protein with two zinc finger domains, we used recombinant caspase-3 and a chromogenic tetrapeptide substrate containing the caspase-3 cleavage site. In this system, Zn2+ inhibited caspase-3 with an IC50 of 0.1 microM. These results identify caspase-3 as a novel target of Zn2+ inhibition in apoptosis and suggest a regulatory role for Zn2+ in modulating the upstream apoptotic machinery.
Perry
D K
DK
Department of Medicine, Duke University Medical Center, Education and Clinical Center (GRECC) Veterans Affairs Medical Center, Durham, North Carolina 27710, USA.
Smyth
M J
MJ
Stennicke
H R
HR
Salvesen
G S
GS
Duriez
P
P
Poirier
G G
GG
Hannun
Y A
YA
D000818:Animals; D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects*; D053148:Caspase 3; D020169:Caspases; D002384:Catalysis; D002417:Cattle; D002474:Cell-Free System; D003546:Cysteine Endopeptidases / Q000378:metabolism*; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology*; D004792:Enzyme Precursors / Q000037:antagonists & inhibitors; D005047:Etoposide / Q000494:pharmacology; D006801:Humans; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D014407:Tumor Cells, Cultured; D015032:Zinc / Q000494:pharmacology*
D000972:Antineoplastic Agents, Phytogenic; D015853:Cysteine Proteinase Inhibitors; D004792:Enzyme Precursors; D005047:Etoposide; D011065:Poly(ADP-ribose) Polymerases; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases; D003546:Cysteine Endopeptidases; D015032:Zinc