Mortalin/GRP75 Binds to Complement C9 and Plays a Role in Resistance to Complement-dependent Cytotoxicity

• Published in: The Journal of biological chemistry Vol. 289; no. 21; pp. 15014 - 15022
• Main Authors: Saar Ray, Moran, Moskovich, Oren, Iosefson, Ohad, Fishelson, Zvi
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 23-05-2014; American Society for Biochemistry and Molecular Biology
• Subjects: Adenosine Diphosphate - immunology; Adenosine Diphosphate - pharmacology; Adenosine Triphosphatases - immunology; Adenosine Triphosphatases - metabolism; Adenosine Triphosphate - immunology; Adenosine Triphosphate - pharmacology; Binding Sites - genetics; Binding Sites - immunology; Blotting, Western; C5b-9; Complement C9 - immunology; Complement C9 - metabolism; Complement Membrane Attack Complex - immunology; Complement Membrane Attack Complex - metabolism; Complement System; Complement System Proteins - immunology; Complement System Proteins - metabolism; Cytotoxicity, Immunologic - immunology; Green Fluorescent Proteins - genetics; Green Fluorescent Proteins - metabolism; GRP75; Heat Shock Protein; HSP70 Heat-Shock Proteins - genetics; HSP70 Heat-Shock Proteins - immunology; HSP70 Heat-Shock Proteins - metabolism; Humans; Immunology; K562 Cells; Microscopy, Confocal; Molecular Chaperone; Mortalin; Necrosis (Necrotic Death); Protein Binding - drug effects; Protein Binding - immunology; RNA Interference; Sodium Chloride - immunology; Sodium Chloride - pharmacology; Complement System; Immunology; Mortalin; C5b-9; Molecular Chaperone; Necrosis (Necrotic Death); GRP75; Heat Shock Protein
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M114.552406

Mortalin/GRP75, the mitochondrial heat shock protein 70, plays a role in cell protection from complement-dependent cytotoxicity (CDC). As shown here, interference with mortalin synthesis enhances sensitivity of K562 erythroleukemia cells to CDC, whereas overexpression of mortalin leads to their resistance to CDC. Quantification of the binding of the C5b-9 membrane attack complex to cells during complement activation shows an inverse correlation between C5b-9 deposition and the level of mortalin in the cell. Following transfection, mortalin-enhanced GFP (EGFP) is located primarily in mitochondria, whereas mortalinΔ51-EGFP lacking the mitochondrial targeting sequence is distributed throughout the cytoplasm. Overexpressed cytosolic mortalinΔ51-EGFP has a reduced protective capacity against CDC relative to mitochondrial mortalin-EGFP. Mortalin was previously shown by us to bind to components of the C5b-9 complex. Two functional domains of mortalin, the N-terminal ATPase domain and the C-terminal substrate-binding domain, were purified after expression in bacteria. Similar to intact mortalin, the ATPase domain, but not the substrate-binding domain, was found to bind to complement proteins C8 and C9 and to inhibit zinc-induced polymerization of C9. Binding of mortalin to complement C9 and C8 occurs through an ionic interaction that is nucleotide-sensitive. We suggest that to express its full protective effect from CDC, mortalin must first reach the mitochondria. In addition, mortalin can potentially target the C8 and C9 complement components through its ATPase domain and inhibit C5b-9 assembly and stability.
Mortalin was shown to contribute to removal of the complement membranolytic C5b-9 complex from the target cell surface.
Modulations of mortalin expression and activity affect deposition of C5b-9 and cell death.
Mortalin, through its ATPase domain, regulates the C5b-9 deposition and confers resistance to complement-dependent cytotoxicity.
Mortalin is a potential therapeutic target in autoimmune diseases and in cancer immunotherapy.