title: Exposure to arginine analog canavanine induces aberrant mitochondrial translation products, mitoribosome stalling, and instability of the mitochondrial proteome.
abstract: Impairment of mitochondrial protein homeostasis disrupts mitochondrial function and causes human diseases and aging, but the molecular mechanisms of protein synthesis and quality control in mammalian mitochondria are not fully understood. Here we demonstrate in human cells that misincorporation of an arginine analog, canavanine, during mitochondrial protein synthesis, induced aberrant translation products and destabilized the mtDNA-encoded proteome, leading to loss of mitochondrial respiratory chain complexes. Furthermore, in the presence of a high concentration of canavanine, mitoribosome stalling could be demonstrated. The stalling did not, however, occur at arginine codons, but downstream of those codons. In particular, two adjacent arginines induced the most prominent downstream stalling effect, with the distance between the arginine codons and the stalling peak corresponding roughly to the length of the ribosomal exit tunnel. These results suggest that misincorporated canavanine disrupted the proper folding of the hydrophobic nascent polypeptides within the exit tunnel or while being inserted into the inner mitochondrial membrane. The canavanine treatment provides a model system for studying the consequences of mitoribosome stalling and the responses to misfolded proteins exiting the mitochondrial ribosome.
authors.affiliation: Research Programs Unit, Molecular Neurology, University of Helsinki, Helsinki, Finland.
authors.forename: Svetlana
authors.initials: S
authors.lastname: Konovalova
authors.affiliation: Research Programs Unit, Molecular Neurology, University of Helsinki, Helsinki, Finland.
authors.forename: Taru
authors.initials: T
authors.lastname: Hilander
authors.affiliation: Division of Biological Stress Response, The Netherlands Cancer Institute, Amsterdam, The Netherlands.
authors.forename: Fabricio
authors.initials: F
authors.lastname: Loayza-Puch
authors.affiliation: Division of Biological Stress Response, The Netherlands Cancer Institute, Amsterdam, The Netherlands.
authors.forename: Koos
authors.initials: K
authors.lastname: Rooijers
authors.affiliation: Division of Biological Stress Response, The Netherlands Cancer Institute, Amsterdam, The Netherlands.
authors.forename: Reuven
authors.initials: R
authors.lastname: Agami
authors.affiliation: Research Programs Unit, Molecular Neurology, University of Helsinki, Helsinki, Finland. Electronic address: henna.tyynismaa@helsinki.fi.
authors.forename: Henna
authors.initials: H
authors.lastname: Tyynismaa
mesh_terms: D001120:Arginine / Q000378:metabolism*; D002172:Canavanine / Q000494:pharmacology*; D045744:Cell Line, Tumor; D006801:Humans; D008928:Mitochondria / Q000187:drug effects* / Q000378:metabolism*; D024101:Mitochondrial Proteins / Q000378:metabolism*; D014176:Protein Biosynthesis; D020543:Proteome / Q000378:metabolism*; D012270:Ribosomes / Q000378:metabolism*
chemical_list: D024101:Mitochondrial Proteins; D020543:Proteome; D002172:Canavanine; D001120:Arginine
keywords: Canavanine; Mitochondria; Protein misfolding; Protein synthesis; Ribosome stalling