Interleukin-1 beta-converting enzyme-like protease cleaves DNA-dependent protein kinase in cytotoxic T cell killing.

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Song, Q 
Burrows, SR 
Smith, G 
Lees-Miller, SP 
Kumar, S 

Cytotoxic T cells (CTL) represent the major defense mechanism against the spread of virus infection. It is believed that the pore-forming protein, perforin, facilitates the entry of a series of serine proteases (particularly granzyme B) into the target cell which ultimately leads to DNA fragmentation and apoptosis. We demonstrate here that during CTL-mediated cytolysis the catalytic subunit of DNA-dependent protein kinase (DNA-PKcs), an enzyme implicated in the repair of double strand breaks in DNA, is specifically cleaved by an interleukin (IL)-1 beta-converting enzyme (ICE)-like protease. A serine protease inhibitor, 3,4-dichloroisocoumarin (DCl), which is known to block granzyme B activity, inhibited CTL-induced apoptosis and prevented the degradation of DNA-PKcs in cells but failed to prevent the degradation of purified DNA-PKcs by CTL extracts. However, Tyr-Val-Ala-Asp-CH2Cl (YVAD-CMK) and other cysteine protease inhibitors prevented the degradation of purified DNA-PKcs by CTL extracts. Furthermore, incubation of DNA-PKcs with granzyme B did not produce the same cleavage pattern observed in cells undergoing apoptosis and when this substrate was incubated with either CTL extracts or the ICE-like protease, CPP32. Sequence analysis revealed that the cleavage site in DNA-PKcs during CTL killing was the same as that when this substrate was exposed to CPP32. This study demonstrates for the first time that the cleavage of DNA-PKcs in this intact cell system is exclusively due to an ICE-like protease.

Amino Acid Sequence, Apoptosis, Caspase 3, Caspases, Cells, Cultured, Cysteine Endopeptidases, Cytotoxicity, Immunologic, DNA-Activated Protein Kinase, DNA-Binding Proteins, Granzymes, Humans, Hydrolysis, Metalloendopeptidases, Molecular Sequence Data, Nuclear Proteins, Peptide Fragments, Peptides, Protein Serine-Threonine Kinases, Serine Endopeptidases, Signal Transduction, T-Lymphocytes, Cytotoxic, Time Factors
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J Exp Med
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Rockefeller University Press