Lamin B phosphorylation by protein kinase calpha and proteolysis during apoptosis in human leukemia HL60 cells.
Protein phosphorylation plays an important role in signal transduction, but its involvement in apoptosis still remains unclear. In this report, the p53-null human leukemia HL60 cells were used to investigate phosphorylation and degradation of lamin B during apoptosis. We found that lamin B was phosphorylated within 1 h after addition of the DNA topoisomerase I inhibitor, camptothecin, and that lamin B phosphorylation preceded lamin B degradation and DNA fragmentation. Using a cell-free system we also found that cytosol from camptothecin-treated cells induced lamin B phosphorylation and degradation in isolated nuclei from untreated HL60 cells. Lamin B phosphorylation was prevented by the protein kinase C (PKC) inhibitor 7-hydroxystaurosporine (UCN-01) but not by the Cdc2 inhibitor, flavopiridol. Phosphorylation of lamin B was inhibited by immunodepletion of PKCalpha from activated cytosol and was restored by addition of purified PKCalpha. PKCalpha activity also increased rapidly as lamin B was phosphorylated after initiation of the apoptotic response in HL60 cells. These data suggest that lamin B is phosphorylated by PKCalpha and proteolyzed before DNA fragmentation in HL60 cells undergoing apoptosis.
Shimizu
T
T
Laboratory of Molecular Pharmacology, Division of Basic Sciences, NCI, National Institutes of Health, Bethesda, Maryland 20892-4255, USA.
Cao
C X
CX
Shao
R G
RG
Pommier
Y
Y
D000595:Amino Acid Sequence; D017209:Apoptosis / Q000187:drug effects; D002166:Camptothecin / Q000494:pharmacology; D002467:Cell Nucleus / Q000378:metabolism; D003374:Coumarins / Q000494:pharmacology; D003600:Cytosol / Q000378:metabolism; D053938:DNA Fragmentation; D018922:HL-60 Cells / Q000166:cytology / Q000378:metabolism*; D006801:Humans; D049934:Isocoumarins; D007527:Isoenzymes / Q000378:metabolism*; D007700:Kinetics; D034921:Lamin Type B; D034882:Lamins; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000737:chemistry / Q000378:metabolism*; D010446:Peptide Fragments / Q000737:chemistry; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism*; D051571:Protein Kinase C-alpha; D017433:Protein Structure, Secondary; D015842:Serine Proteinase Inhibitors / Q000494:pharmacology; D013997:Time Factors; D059004:Topoisomerase I Inhibitors; D016159:Tumor Suppressor Protein p53 / Q000172:deficiency
D003374:Coumarins; D049934:Isocoumarins; D007527:Isoenzymes; D034921:Lamin Type B; D034882:Lamins; D009687:Nuclear Proteins; D010446:Peptide Fragments; D015842:Serine Proteinase Inhibitors; D059004:Topoisomerase I Inhibitors; D016159:Tumor Suppressor Protein p53; C046235:3,4-dichloroisocoumarin; C496628:PRKCA protein, human; D011493:Protein Kinase C; D051571:Protein Kinase C-alpha; D002166:Camptothecin