Cleavage at the carboxyl-terminus of Ku80 during apoptosis in human Jurkat T cells.
We have previously reported that the amount of Apg-2, an Hsp110 family protein, decreases during apoptosis in Jurkat T cells. Since we hypothesized that Apg-2 would be cleaved by caspase-3 during apoptosis, a cleavage-site-directed antibody was raised against the carboxyl-terminus of the Apg-2 fragment that appears after the cleavage by caspase-3. Although this antibody could not detect the Apg-2 fragment in apoptotic cells, three additional fragments were unexpectedly detected. Based on the results of microsequencing, one of these fragments was identified as Ku80. Ku80 is a nuclear protein and a component of DNA-dependent protein kinase (DNA-PK). In this study, we observed that Ku80 is cleaved at Asp-730 residue during apoptosis, and this cleavage occurs in the nucleus in the early apoptotic phase. Furthermore, Ku80 is distributed in the cytoplasm of nuclear fragmented apoptotic cells, although the cleaved fragment contains the nuclear-localization signal (NLS). Our study clearly shows that Ku80 is cleaved in the nucleus, and distributes in the cytoplasm during apoptosis.
Department of Basic Medical Sciences, Institute of Medical Science, University of Tokyo, Minato-ku, Tokyo 108-8639, Japan.
Masahiko
M
Kato
Takashi
T
Nonaka
Shinobu
S
Imajoh-Ohmi
D000595:Amino Acid Sequence; D000906:Antibodies; D017209:Apoptosis; D020169:Caspases / Q000378:metabolism; D002467:Cell Nucleus / Q000378:metabolism; D003593:Cytoplasm / Q000378:metabolism; D051747:DNA-Activated Protein Kinase; D004268:DNA-Binding Proteins / Q000378:metabolism; D050957:HSP110 Heat-Shock Proteins; D006367:HeLa Cells; D006360:Heat-Shock Proteins / Q000276:immunology / Q000378:metabolism*; D006801:Humans; D019169:Jurkat Cells; D018613:Microscopy, Confocal; D008969:Molecular Sequence Data; D009687:Nuclear Proteins; D010446:Peptide Fragments / Q000032:analysis; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D014162:Transfection
D000906:Antibodies; D004268:DNA-Binding Proteins; D050957:HSP110 Heat-Shock Proteins; C121993:HSPA4 protein, human; D006360:Heat-Shock Proteins; D009687:Nuclear Proteins; D010446:Peptide Fragments; D051747:DNA-Activated Protein Kinase; C497263:PRKDC protein, human; D017346:Protein Serine-Threonine Kinases; D020169:Caspases