Activation of caspase-2 mediates the apoptosis induced by GTP-depletion in insulin-secreting (HIT-T15) cells.
This study investigated the possible involvement of a specific caspase(s) (a family of aspartate-specific cysteine proteases) in programmed cell death of islet beta-cells due to sustained GTP depletion. Treatment (up to 48 h) with 3 microg/ml mycophenolic acid (MPA), which specifically depletes intracellular guanine nucleotides, reduced cell-cycle progression from G1 phase into S and G2/M phases (as assessed by flow cytometry) and, subsequently, induced apoptosis of HIT-15 cells (transformed pancreatic beta-cells). The latter was accompanied by a marked increase of caspase-2 activity (+343%) and moderate activation of caspase-9 (+150%) and caspase-3 (+145%). Importantly, only caspase-2 activation preceded induction of apoptosis. There was no change in activity of caspase-1, -4, -5, -6, and -8. Release of the mitochondrial protein cytochrome c into cytosol was also observed at a late stage. Cotreatment of cells with a permeable pan-caspase inhibitor (Z-VAD-FMK) blocked GTP depletion-induced cell death in a dose-dependent manner. A specific caspase-2 inhibitor (Z-VDVAD-FMK), but not a caspase-3 inhibitor (DEVD-CHO), was also capable of restoring cell viability. Interestingly, activation of caspase-2 leads to caspase-3 activation because the caspase-2 inhibitor abrogated caspase-3 activity. Our results indicate that, while activation of multiple caspases are involved in the execution phase of GTP depletion-induced apoptosis, caspase-2 appears to play the major role in the initiation of this program. This study revealed a novel, caspase-2 mediated form of apoptosis that may be consequent to impaired mitogenesis.
Huo
Jianxin
J
Cardiovascular Research Institute, National University Medical Institutes, National University of Singapore, 10 Medical Drive, Singapore 117597.
Luo
Rui-Hua
RH
Metz
Stewart A
SA
Li
Guodong
G
D000818:Animals; D000903:Antibiotics, Antineoplastic / Q000494:pharmacology; D017209:Apoptosis / Q000502:physiology*; D015153:Blotting, Western; D053143:Caspase 2; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism / Q000502:physiology*; D002460:Cell Line; D006224:Cricetinae; D003574:Cytochrome c Group / Q000378:metabolism; D003600:Cytosol / Q000187:drug effects / Q000378:metabolism; D053938:DNA Fragmentation; D004789:Enzyme Activation / Q000502:physiology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005434:Flow Cytometry; D006160:Guanosine Triphosphate / Q000502:physiology*; D007328:Insulin / Q000378:metabolism*; D000078790:Insulin Secretion; D007515:Islets of Langerhans / Q000187:drug effects / Q000201:enzymology / Q000502:physiology*; D007527:Isoenzymes / Q000037:antagonists & inhibitors / Q000378:metabolism; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism; D009173:Mycophenolic Acid / Q000494:pharmacology; D011233:Precipitin Tests
D000903:Antibiotics, Antineoplastic; D061945:Caspase Inhibitors; D003574:Cytochrome c Group; D004791:Enzyme Inhibitors; D007328:Insulin; D007527:Isoenzymes; D006160:Guanosine Triphosphate; D053143:Caspase 2; D020169:Caspases; D009173:Mycophenolic Acid