title: Ubiquitination of specific mitochondrial matrix proteins.
abstract: Several protein quality control systems in bacteria and/or mitochondrial matrix from lower eukaryotes are absent in higher eukaryotes. These are transfer-messenger RNA (tmRNA), The N-end rule ATP-dependent protease ClpAP, and two more ATP-dependent proteases, HslUV and ClpXP (in yeast). The lost proteases resemble the 26S proteasome and the role of tmRNA and the N-end rule in eukaryotic cytosol is performed by the ubiquitin proteasome system (UPS). Therefore, we hypothesized that the UPS might have substituted these systems - at least partially - in the mitochondrial matrix of higher eukaryotes. Using three independent experimental approaches, we demonstrated the presence of ubiquitinated proteins in the matrix of isolated yeast mitochondria. First, we show that isolated mitochondria contain ubiquitin (Ub) conjugates, which remained intact after trypsin digestion. Second, we demonstrate that the mitochondrial soluble fraction contains Ub-conjugates, several of which were identified by mass spectrometry and are localized to the matrix. Third, using immunoaffinity enrichment by specific antibodies recognizing digested ubiquitinated peptides, we identified a group of Ub-modified matrix proteins. The modification was further substantiated by separation on SDS-PAGE and immunoblots. Last, we attempted to identify the ubiquitin ligase(s) involved, and identified Dma1p as a trypsin-resistant protein in our mitochondrial preparations. Taken together, these data suggest a yet undefined role for the UPS in regulation of the mitochondrial matrix proteins.
authors.affiliation: The Janet and David Polak Tumor and Vascular Biology Research Center and the Technion Integrated Cancer Center (TICC), The Rappaport Faculty of Medicine and Research Institute, Haifa, 31096, Israel.
authors.forename: Gilad
authors.initials: G
authors.lastname: Lehmann
authors.affiliation: The Smoler Proteomics Center, Faculty of Biology - Technion-Israel Institute of Technology, Haifa, 32000, Israel.
authors.forename: Tamar
authors.initials: T
authors.lastname: Ziv
authors.affiliation: The Janet and David Polak Tumor and Vascular Biology Research Center and the Technion Integrated Cancer Center (TICC), The Rappaport Faculty of Medicine and Research Institute, Haifa, 31096, Israel.
authors.forename: Ori
authors.initials: O
authors.lastname: Braten
authors.affiliation: The Smoler Proteomics Center, Faculty of Biology - Technion-Israel Institute of Technology, Haifa, 32000, Israel.
authors.forename: Arie
authors.initials: A
authors.lastname: Admon
authors.affiliation: The Janet and David Polak Tumor and Vascular Biology Research Center and the Technion Integrated Cancer Center (TICC), The Rappaport Faculty of Medicine and Research Institute, Haifa, 31096, Israel.
authors.forename: Ronald G
authors.initials: RG
authors.lastname: Udasin
authors.affiliation: The Janet and David Polak Tumor and Vascular Biology Research Center and the Technion Integrated Cancer Center (TICC), The Rappaport Faculty of Medicine and Research Institute, Haifa, 31096, Israel. Electronic address: aaroncie@tx.technion.ac.il.
authors.forename: Aaron
authors.initials: A
authors.lastname: Ciechanover
mesh_terms: D000595:Amino Acid Sequence; D005656:Fungal Proteins / Q000737:chemistry / Q000378:metabolism*; D024101:Mitochondrial Proteins / Q000737:chemistry / Q000378:metabolism*; D046988:Proteasome Endopeptidase Complex / Q000737:chemistry / Q000378:metabolism; D059748:Proteolysis; D025801:Ubiquitin / Q000378:metabolism; D054875:Ubiquitination; D015003:Yeasts / Q000737:chemistry / Q000166:cytology / Q000378:metabolism*
chemical_list: D005656:Fungal Proteins; D024101:Mitochondrial Proteins; D025801:Ubiquitin; D046988:Proteasome Endopeptidase Complex
keywords: Matrix; Mitochondria; Proteasome; Protein degradation; Ubiquitin