Nitration of the mitochondrial complex I subunit NDUFB8 elicits RIP1- and RIP3-mediated necrosis.
Nitric oxide (NO) and other reactive nitrogen species target multiple sites in the mitochondria to influence cellular bioenergetics and survival. Kinetic imaging studies revealed that NO from either activated macrophages or donor compounds rapidly diffuses to the mitochondria, causing a dose-dependent progressive increase in NO-dependent DAF fluorescence, which corresponded to mitochondrial membrane potential loss and initiated alterations in cellular bioenergetics that ultimately led to necrotic cell death. Cellular dysfunction is mediated by an elevated 3-nitrotyrosine signature of the mitochondrial complex I subunit NDUFB8, which is vital for normal mitochondrial function as evidenced by selective knockdown via siRNA. Overexpression of mitochondrial superoxide dismutase substantially decreased NDUFB8 nitration and restored mitochondrial homeostasis. Further, treatment of cells with either necrostatin-1 or siRNA knockdown of RIP1 and RIP3 prevented NO-mediated necrosis. This work demonstrates that the interaction between NO and mitochondrially derived superoxide alters mitochondrial bioenergetics and cell function, thus providing a molecular mechanism for reactive oxygen and nitrogen species-mediated alterations in mitochondrial homeostasis.
Davis
Christiana W
CW
Institute for Environmental Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA.
Hawkins
Brian J
BJ
Ramasamy
Subbiah
S
Irrinki
Krishna M
KM
Cameron
Bruce A
BA
Islam
Khalid
K
Daswani
Varsha P
VP
Doonan
Patrick J
PJ
Manevich
Yefim
Y
Madesh
Muniswamy
M
D000818:Animals; D002460:Cell Line; D042967:Electron Transport Complex I / Q000235:genetics / Q000378:metabolism*; D004730:Endothelium, Vascular / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D007211:Indoles / Q000494:pharmacology; D008264:Macrophages / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D053078:Membrane Potential, Mitochondrial; D051379:Mice; D008856:Microscopy, Fluorescence; D008928:Mitochondria / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D009336:Necrosis / Q000235:genetics; D009569:Nitric Oxide / Q000378:metabolism; D028861:Nuclear Pore Complex Proteins / Q000235:genetics / Q000378:metabolism; D021122:Protein Subunits / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics; D016601:RNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D051381:Rats; D053422:Receptor-Interacting Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism
C119724:AGFG1 protein, human; D007093:Imidazoles; D007211:Indoles; C000618340:NDUFB8 protein, human; D028861:Nuclear Pore Complex Proteins; D021122:Protein Subunits; D034741:RNA, Small Interfering; D016601:RNA-Binding Proteins; C507699:necrostatin-1; D009569:Nitric Oxide; C119705:RIPK3 protein, human; D053422:Receptor-Interacting Protein Serine-Threonine Kinases; D042967:Electron Transport Complex I