Caspase processing and nuclear export of CTP:phosphocholine cytidylyltransferase alpha during farnesol-induced apoptosis.
CTP:phosphocholine cytidylyltransferase alpha (CCT alpha) is a nuclear enzyme that catalyzes the rate-limiting step in the CDP-choline pathway, the primary route for synthesis of phosphatidylcholine (PtdCho) in eukaryotic cells. Induction of apoptosis by farnesol (FOH) and other cytotoxic drugs has been shown to alter PtdCho synthesis via the CDP-choline pathway. Here we report that FOH-induced apoptosis in CHO cells caused a dose-dependent activation of CCT alpha and inhibition of the final step in the pathway, resulting in a biphasic effect on PtdCho synthesis. Activation of CCT alpha was accompanied by enzyme translocation to the nuclear envelope within 30 min of FOH addition to cells. Following translocation to membranes, CCT alpha was exported from the nucleus and underwent caspase-mediated proteolysis that coincided with poly(ADP-ribose) polymerase cleavage. Site-directed mutagenesis and in vivo and in vitro expression studies mapped a caspase 6 and/or 8 cleavage site to TEED(28 downward arrow)G, the final residue in the CCT alpha nuclear localization signal. Nuclear export of CCT alpha appeared to be an active process in FOH-treated CHO cells that was independent of caspase removal of the nuclear localization signal. Caspase cleavage of CCT alpha occurred during UV or chelerythrine-induced apoptosis; however, nuclear membrane translocation and nuclear export were not evident under these conditions. Thus, caspase cleavage of CCT alpha was a late feature of several apoptotic programs that occurred in the nucleus or at the nuclear envelope. Activation and nuclear export of CCT alpha were early events in FOH-induced apoptosis that contributed to altered PtdCho synthesis and, in conjunction with caspase cleavage, excluded CCT alpha from the nucleus.
Lagace
Thomas A
TA
Atlantic Research Centre, Department of Pediatrics, Dalhousie University, Halifax, Nova Scotia, Canada B3H 4H7.
Miller
Jessica R
JR
Ridgway
Neale D
ND
D016475:3T3 Cells; D000470:Alkaloids; D000595:Amino Acid Sequence; D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D001224:Aspartic Acid / Q000378:metabolism; D053119:Benzophenanthridines; D019556:COS Cells; D020169:Caspases / Q000378:metabolism*; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism* / Q000528:radiation effects; D019916:Choline-Phosphate Cytidylyltransferase / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D006224:Cricetinae; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005204:Farnesol / Q000494:pharmacology*; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D016296:Mutagenesis; D019913:Nuclear Localization Signals / Q000378:metabolism; D010617:Phenanthridines / Q000494:pharmacology; D011499:Protein Processing, Post-Translational; D021381:Protein Transport; D014466:Ultraviolet Rays
D000470:Alkaloids; D053119:Benzophenanthridines; D004791:Enzyme Inhibitors; D019913:Nuclear Localization Signals; D010617:Phenanthridines; D001224:Aspartic Acid; D005204:Farnesol; C016299:chelerythrine; D019916:Choline-Phosphate Cytidylyltransferase; D020169:Caspases