Induction of SM-20 in PC12 cells leads to increased cytochrome c levels, accumulation of cytochrome c in the cytosol, and caspase-dependent cell death.
Sympathetic neurons deprived of nerve growth factor (NGF) release cytochrome c into the cytosol and undergo caspase-dependent cell death through a process that requires de novo gene expression. Expression of the SM-20 gene increases after NGF withdrawal, and ectopic SM-20 expression induces cell death in NGF-maintained neurons. To further evaluate the mechanism by which SM-20 promotes cell death, we developed a PC12-derived cell line in which SM-20 expression can be induced by addition of doxycycline to the culture medium. Induction of SM-20 in either undifferentiated or NGF-differentiated cells resulted in cell death. Cell death was accompanied by an increase in caspase activity and was inhibited by the caspase inhibitor zVAD-FMK. Analysis of cytochrome c in cytosolic and mitochondria-enriched subcellular fractions revealed that induction of SM-20 led to the accumulation of cytochrome c in the cytosol. Surprisingly, SM-20 expression also resulted in a selective increase in the total amount of cytochrome c protein. Thus, induction of SM-20 expression appears to affect both the amount and subcellular localization of cytochrome c in PC12 cells. These results suggest that SM-20 promotes caspase-dependent cell death through a mechanism involving cytochrome c.
Straub
Jennifer A
JA
Department of Pharmacology and Physiology, University of Rochester School of Medicine and Dentistry, Rochester, New York 14642, USA.
Lipscomb
Elizabeth A
EA
Yoshida
Eileen S
ES
Freeman
Robert S
RS
D000818:Animals; D017209:Apoptosis; D020169:Caspases / Q000378:metabolism*; D002452:Cell Count; D002454:Cell Differentiation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects / Q000235:genetics; D003574:Cytochrome c Group / Q000378:metabolism*; D003600:Cytosol / Q000378:metabolism*; D004268:DNA-Binding Proteins; D004318:Doxycycline / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects; D064799:Hypoxia-Inducible Factor-Proline Dioxygenases; D017874:Immediate-Early Proteins / Q000235:genetics / Q000378:metabolism*; D020932:Nerve Growth Factor / Q000494:pharmacology; D016716:PC12 Cells; D051381:Rats; D014161:Transduction, Genetic; D019076:Transgenes
D003574:Cytochrome c Group; D004268:DNA-Binding Proteins; D004791:Enzyme Inhibitors; D017874:Immediate-Early Proteins; D020932:Nerve Growth Factor; C087100:Egln3 protein, rat; D064799:Hypoxia-Inducible Factor-Proline Dioxygenases; D020169:Caspases; D004318:Doxycycline