Caspases indirectly regulate cleavage of the mitochondrial fusion GTPase OPA1 in neurons undergoing apoptosis.
The critical processes of mitochondrial fission and fusion are regulated by members of the dynamin family of GTPases. Imbalances in mitochondrial fission and fusion contribute to neuronal cell death. For example, increased fission mediated by the dynamin-related GTPase, Drp1, or decreased fusion resulting from inactivating mutations in the OPA1 GTPase, causes neuronal apoptosis and/or neurodegeneration. Recent studies indicate that post-translational processing regulates OPA1 function in non-neuronal cells and moreover, aberrant processing of OPA1 is induced during apoptosis. To date, the post-translational processing of OPA1 during neuronal apoptosis has not been examined. Here, we show that cerebellar granule neurons (CGNs) or neuroblastoma cells exposed to pro-apoptotic stressors display a novel N-terminal cleavage of OPA1 which is blocked by either pan-caspase or caspase-8 selective inhibitors. OPA1 cleavage occurs concurrently with mitochondrial fragmentation and cytochrome c release in CGNs deprived of depolarizing potassium (5K condition). Although a caspase-8 selective inhibitor prevents both 5K-induced OPA1 cleavage and mitochondrial fragmentation, recombinant caspase-8 fails to cleave OPA1 in vitro. In marked contrast, either caspase-8 or caspase-3 stimulates OPA1 cleavage in digitonin-permeabilized rat brain mitochondria, suggesting that OPA1 is cleaved by an intermembrane space protease which is regulated by active caspases. Finally, the N-terminal truncation of OPA1 induced during neuronal apoptosis removes an essential residue (K301) within the GTPase domain. These data are the first to demonstrate OPA1 cleavage during neuronal apoptosis and they implicate caspases as indirect regulators of OPA1 processing in degenerating neurons.
Loucks
F Alexandra
FA
Research Service, Veterans Affairs Medical Center, Denver, Colorado, USA.
Schroeder
Emily K
EK
Zommer
Amelia E
AE
Hilger
Shea
S
Kelsey
Natalie A
NA
Bouchard
Ron J
RJ
Blackstone
Craig
C
Brewster
Jay L
JL
Linseman
Daniel A
DA
D000818:Animals; D017209:Apoptosis / Q000502:physiology*; D001921:Brain / Q000378:metabolism; D053148:Caspase 3 / Q000378:metabolism; D053181:Caspase 8 / Q000378:metabolism; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism*; D002478:Cells, Cultured; D002531:Cerebellum / Q000201:enzymology; D045304:Cytochromes c / Q000378:metabolism; D004072:Digitonin / Q000494:pharmacology; D020558:GTP Phosphohydrolases / Q000378:metabolism*; D006801:Humans; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism; D009447:Neuroblastoma / Q000201:enzymology; D009474:Neurons / Q000166:cytology* / Q000201:enzymology*; D017434:Protein Structure, Tertiary; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D019311:Staurosporine / Q000494:pharmacology
D061945:Caspase Inhibitors; D011500:Protein Synthesis Inhibitors; D045304:Cytochromes c; D053148:Caspase 3; D053181:Caspase 8; D020169:Caspases; D020558:GTP Phosphohydrolases; C415198:OPA1 protein, human; C498811:Opa1 protein, rat; D019311:Staurosporine; D004072:Digitonin