Calpain 10: a mitochondrial calpain and its role in calcium-induced mitochondrial dysfunction.
Calpains, Ca(2+)-activated cysteine proteases, are cytosolic enzymes implicated in numerous cellular functions and pathologies. We identified a mitochondrial Ca(2+)-inducible protease that hydrolyzed a calpain substrate (SLLVY-AMC) and was inhibited by active site-directed calpain inhibitors as calpain 10, an atypical calpain lacking domain IV. Immunoblot analysis and activity assays revealed calpain 10 in the mitochondrial outer membrane, intermembrane space, inner membrane, and matrix fractions. Mitochondrial staining was observed when COOH-terminal green fluorescent protein-tagged calpain 10 was overexpressed in NIH-3T3 cells and the mitochondrial targeting sequence was localized to the NH(2)-terminal 15 amino acids. Overexpression of mitochondrial calpain 10 resulted in mitochondrial swelling and autophagy that was blocked by the mitochondrial permeability transition (MPT) inhibitor cyclosporine A. With the use of isolated mitochondria, Ca(2+)-induced MPT was partially decreased by calpain inhibitors. More importantly, Ca(2+)-induced inhibition of Complex I of the electron transport chain was blocked by calpain inhibitors and two Complex I proteins were identified as targets of mitochondrial calpain 10, NDUFV2, and ND6. In conclusion, calpain 10 is the first reported mitochondrially targeted calpain and is a mediator of mitochondrial dysfunction through the cleavage of Complex I subunits and activation of MPT.
Arrington
David D
DD
Department of Pharmaceutical Sciences, Medical University of South Carolina, Charleston, South Carolina 29425, USA.
Van Vleet
Terry R
TR
Schnellmann
Rick G
RG
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002154:Calpain / Q000235:genetics / Q000378:metabolism*; D016572:Cyclosporine / Q000378:metabolism; D004579:Electron Transport / Q000502:physiology; D042967:Electron Transport Complex I / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000378:metabolism; D005260:Female; D006801:Humans; D008297:Male; D051379:Mice; D008928:Mitochondria / Q000201:enzymology* / Q000648:ultrastructure; D028361:Mitochondrial Diseases / Q000378:metabolism*; D008969:Molecular Sequence Data; D009245:NADH Dehydrogenase / Q000378:metabolism; D021382:Protein Sorting Signals; D021122:Protein Subunits / Q000378:metabolism; D011817:Rabbits; D051381:Rats; D017207:Rats, Sprague-Dawley; D013367:Submitochondrial Particles / Q000201:enzymology
D004791:Enzyme Inhibitors; D021382:Protein Sorting Signals; D021122:Protein Subunits; D016572:Cyclosporine; D009245:NADH Dehydrogenase; D002154:Calpain; C415197:calpain 10; D042967:Electron Transport Complex I; D002118:Calcium