The catalytic subuntt of dna-dependent protein kinase is cleaved during fas-mediated apoptosis in jurkat cells

K. R. McConnell, J. A. Hardin

Research output: Contribution to journalArticle

Abstract

The DNA-dependent protein kinase (DNA-PK) is a serine/threonine kinase linked to DNA repair, cell cycle progression and V(D)J recombination. It is composed of an approximately 460 kDa catalytic subunit (DNA-PKcs) and the human autoantigen Ku (p70 and p86), which serves to target the catalytic subunit to the free ends of double-stranded DNA breaks. Since apoptosis is associated with formation of dsDNA breaks, we examined in Jurkat cells the behavior of the catalytic subunit during Fas (Apo-1, CD95) mediated apoptosis. Using sera from patients with connective tissue disease, polyclonal antibodies or a series of monoclonal antibodies directed toward DNA-PKcs, we found by immunoprecipitation of metabolically-labeled lysates that the catalytic subunit was cleaved into several smaller polypeptides at the onset of apoptosis. Some polypeptides appeared to remain associated with the Ku subunit after cleavage. Additional polypeptide fragments appeared as apoptosis progressed. Western blotting confirmed that the smaller polypeptides arose from DNA-PKcs and were not merely proteins which became associated with DNA-PKcs during apoptosis. In contrast to DNA-PKcs, the Ku subunits were neither cleaved nor decreased in amount during apoptosis, nor were other common autoantigens cleaved during the time required for DNA-PKcs cleavage. Finally, resting lymphocytes were more resistant than cycling lymphocytes to Fas-mediated apoptosis. We surmise that cleavage of DNA-PKcs may represent a mechanism for regulating the function of DNA-PK during programmed cell death. Whether the polypeptides which result from apoptosis-mediated cleavage of DNA-PKcS incite an autoimmune response remains to be determined.

Original languageEnglish (US)
JournalJournal of Investigative Medicine
Volume44
Issue number3
StatePublished - 1996
Externally publishedYes

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Jurkat Cells
Protein Kinases
Apoptosis
DNA
DNA Cleavage
Catalytic Domain
Peptides
DNA-Activated Protein Kinase
Lymphocytes
Autoantigens
Catalytic DNA
V(D)J Recombination
Connective Tissue Diseases
Double-Stranded DNA Breaks
Protein-Serine-Threonine Kinases
Autoimmunity
Immunoprecipitation
DNA Repair
Cell death
Cell Cycle

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)

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The catalytic subuntt of dna-dependent protein kinase is cleaved during fas-mediated apoptosis in jurkat cells. / McConnell, K. R.; Hardin, J. A.

In: Journal of Investigative Medicine, Vol. 44, No. 3, 1996.

Research output: Contribution to journalArticle

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