title: Rabies virus phosphoprotein interacts with mitochondrial Complex I and induces mitochondrial dysfunction and oxidative stress.
abstract: Our previous studies in an experimental model of rabies showed neuronal process degeneration in association with severe clinical disease. Cultured adult rodent dorsal root ganglion neurons infected with challenge virus standard (CVS)-11 strain of rabies virus (RABV) showed axonal swellings and reduced axonal growth with evidence of oxidative stress. We have shown that CVS infection alters a variety of mitochondrial parameters and increases reactive oxygen species (ROS) production and mitochondrial Complex I activity vs. mock infection. We have hypothesized that a RABV protein targets mitochondria and triggers dysfunction. Mitochondrial extracts of mouse neuroblastoma cells were analyzed with a proteomics approach. We have identified peptides belonging to the RABV nucleocapsid protein (N), phosphoprotein (P), and glycoprotein (G), and our data indicate that the extract was most highly enriched with P. P was also detected by immunoblotting in RABV-infected purified mitochondrial extracts and also in Complex I immunoprecipitates from the extracts but not in mock-infected extracts. A plasmid expressing P in cells increased Complex I activity and increased ROS generation, whereas expression of other RABV proteins did not. We have analyzed recombinant plasmids encoding various P gene segments. Expression of a peptide from amino acid 139-172 increased Complex I activity and ROS generation similar to expression of the entire P protein, whereas peptides that did not contain this region did not increase Complex I activity or induce ROS generation. These results indicate that a region of the RABV P interacts with Complex I in mitochondria causing mitochondrial dysfunction, increased generation of ROS, and oxidative stress.
authors.affiliation: Department of Internal Medicine (Neurology), University of Manitoba, Winnipeg, MB, Canada.
authors.forename: Wafa
authors.initials: W
authors.lastname: Kammouni
authors.forename: Heidi
authors.initials: H
authors.lastname: Wood
authors.forename: Ali
authors.initials: A
authors.lastname: Saleh
authors.forename: Camila M
authors.initials: CM
authors.lastname: Appolinario
authors.forename: Paul
authors.initials: P
authors.lastname: Fernyhough
authors.forename: Alan C
authors.initials: AC
authors.lastname: Jackson
mesh_terms: D000818:Animals; D015153:Blotting, Western; D045744:Cell Line, Tumor; D042967:Electron Transport Complex I / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D007150:Immunohistochemistry; D047468:Immunoprecipitation; D051379:Mice; D008928:Mitochondria; D016297:Mutagenesis, Site-Directed; D018384:Oxidative Stress; D010750:Phosphoproteins / Q000378:metabolism*; D040901:Proteomics; D011818:Rabies / Q000378:metabolism / Q000821:virology*; D011820:Rabies virus / Q000502:physiology*; D014162:Transfection; D014764:Viral Proteins / Q000378:metabolism*
chemical_list: D010750:Phosphoproteins; D014764:Viral Proteins; D042967:Electron Transport Complex I