Apoptosis induced by gliotoxin is preceded by phosphorylation of histone H3 and enhanced sensitivity of chromatin to nuclease digestion.
The fungal toxin gliotoxin induces apoptotic cell death in a variety of cells. Apoptosis induced in thymocytes by gliotoxin is rapid, and DNA fragmentation is observable within 4 h treatment. Apoptosis induced by gliotoxin is calcium-independent and unaffected by protein synthesis inhibitors. We have previously shown that gliotoxin results in phosphorylation of a 16.3-kDa protein within 10 min treatment of thymocytes. Here we show that this protein is histone H3 and phosphorylation occurs on Ser-10. Cyclic AMP levels and activity of protein kinase A (PKA) are raised in cells treated with gliotoxin. Apoptosis is inhibited by genistein which also inhibits PKA and histone H3 phosphorylation. Apoptosis is also inhibited by a number of specific inhibitors of PKA suggesting apoptosis induced by gliotoxin is modulated by this kinase. The agents forskolin and cholera toxin do not induce rapid phosphorylation of H3 although some increase in phosphorylation of H3 does occur after 8 h with these agents. Forskolin and cholera toxin also induce apoptosis but over a longer time course than gliotoxin. In all cases levels of apoptosis correlate with degree of H3 phosphorylation. Cells treated with gliotoxin show an early sensitivity to micrococcal nuclease and DNase I digestion indicating a functional relationship between DNA fragmentation and H3 phosphorylation.
Waring
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P
Division of Immunology and Cell Biology, John Curtin School of Medical Research, Australian National University, P.O. Box 334, Canberra City, Australian Capital Territory 2601, Australia. Paul.Waring@anu.edu.au
Khan
T
T
Sjaarda
A
A
D000595:Amino Acid Sequence; D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D016203:CDC2 Protein Kinase / Q000378:metabolism; D002478:Cells, Cultured; D002843:Chromatin / Q000187:drug effects / Q000502:physiology*; D000242:Cyclic AMP / Q000378:metabolism / Q000494:pharmacology; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D053938:DNA Fragmentation; D003850:Deoxyribonuclease I / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D019833:Genistein; D005912:Gliotoxin / Q000494:pharmacology*; D006657:Histones / Q000737:chemistry / Q000378:metabolism*; D007529:Isoflavones / Q000494:pharmacology; D051379:Mice; D008807:Mice, Inbred BALB C; D008836:Micrococcal Nuclease / Q000494:pharmacology; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry / Q000302:isolation & purification / Q000494:pharmacology; D010766:Phosphorylation; D047428:Protein Kinase Inhibitors; D013601:T-Lymphocytes / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D013997:Time Factors
D002843:Chromatin; D004791:Enzyme Inhibitors; D006657:Histones; D007529:Isoflavones; D010446:Peptide Fragments; D047428:Protein Kinase Inhibitors; C518603:protein kinase inhibitor peptide (5-24); D005912:Gliotoxin; D019833:Genistein; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D016203:CDC2 Protein Kinase; D003850:Deoxyribonuclease I; D008836:Micrococcal Nuclease