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Hans Weiher - One of the best experts on this subject based on the ideXlab platform.
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A monoclonal antibody raised against bacterially expressed MPV17 sequences shows peroxisomal, endosomal and lysosomal localisation in U2OS cells
BMC research notes, 2016Co-Authors: Hans Weiher, Ralf M. Zwacka, Miia Vapola, J. Kalervo Hiltunen, Haymo Pircher, Pidder Jansen-dürr, Silke Hegenbarth, Percy A. Knolle, Silke Grunau, Elmon SchmelzerAbstract:Recessive mutations in the MPV17 gene cause mitochondrial DNA depletion syndrome, a fatal infantile genetic liver disease in humans. Loss of function in mice leads to glomerulosclerosis and sensineural deafness accompanied with mitochondrial DNA depletion. Mutations in the yeast homolog Sym1, and in the zebra fish homolog tra cause interesting, but not obviously related phenotypes, although the human gene can complement the yeast Sym1 mutation. The MPV17 protein is a hydrophobic membrane protein of 176 amino acids and unknown function. Initially localised in murine peroxisomes, it was later reported to be a mitochondrial inner membrane protein in humans and in yeast. To resolve this contradiction we tested two new mouse monoclonal antibodies directed against the human MPV17 protein in Western blots and immunohistochemistry on human U2OS cells. One of these monoclonal antibodies showed specific reactivity to a protein of 20 kD absent in MPV17 negative mouse cells. Immunofluorescence studies revealed colocalisation with peroxisomal, endosomal and lysosomal markers, but not with mitochondria. This data reveal a novel connection between a possible peroxisomal/endosomal/lysosomal function and mitochondrial DNA depletion.
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the human mitochondrial dna depletion syndrome gene MPV17 encodes a non selective channel that modulates membrane potential
Journal of Biological Chemistry, 2015Co-Authors: Vasily D. Antonenkov, Hans Weiher, Antti Isomursu, Daniela Mennerich, Miia Vapola, Thomas Kietzmann, Kalervo J HiltunenAbstract:The human MPV17-related mitochondrial DNA depletion syndrome is an inherited autosomal recessive disease caused by mutations in the inner mitochondrial membrane protein MPV17. Although more than 30 MPV17 gene mutations were shown to be associated with mitochondrial DNA depletion syndrome, the function of MPV17 is still unknown. Mice deficient in MPV17 show signs of premature aging. In the present study, we used electrophysiological measurements with recombinant MPV17 to reveal that this protein forms a non-selective channel with a pore diameter of 1.8 nm and located the channel's selectivity filter. The channel was weakly cation-selective and showed several subconductance states. Voltage-dependent gating of the channel was regulated by redox conditions and pH and was affected also in mutants mimicking a phosphorylated state. Likewise, the mitochondrial membrane potential (Δψm) and the cellular production of reactive oxygen species were higher in embryonic fibroblasts from MPV17−/− mice. However, despite the elevated Δψm, the MPV17-deficient mitochondria showed signs of accelerated fission. Together, these observations uncover the role of MPV17 as a Δψm-modulating channel that apparently contributes to mitochondrial homeostasis under different conditions. Background: MPV17 is a mitochondrial inner membrane protein with unknown function. Results: Recombinant human MPV17 shows highly regulated channel-forming activity; the mitochondrial membrane potential and the reactive oxygen species formation were elevated in embryonic fibroblasts from MPV17−/− mice. Conclusion: MPV17 functions as a non-selective channel modulating the membrane potential to preserve mitochondrial homeostasis. Significance: Our data are important for understanding the role of MPV17 protein under physiological and pathological conditions.
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The Human Mitochondrial DNA Depletion Syndrome Gene MPV17 Encodes a Non-selective Channel That Modulates Membrane Potential
The Journal of biological chemistry, 2015Co-Authors: Vasily D. Antonenkov, Hans Weiher, Antti Isomursu, Daniela Mennerich, Miia Vapola, Thomas Kietzmann, J. Kalervo HiltunenAbstract:The human MPV17-related mitochondrial DNA depletion syndrome is an inherited autosomal recessive disease caused by mutations in the inner mitochondrial membrane protein MPV17. Although more than 30 MPV17 gene mutations were shown to be associated with mitochondrial DNA depletion syndrome, the function of MPV17 is still unknown. Mice deficient in MPV17 show signs of premature aging. In the present study, we used electrophysiological measurements with recombinant MPV17 to reveal that this protein forms a non-selective channel with a pore diameter of 1.8 nm and located the channel's selectivity filter. The channel was weakly cation-selective and showed several subconductance states. Voltage-dependent gating of the channel was regulated by redox conditions and pH and was affected also in mutants mimicking a phosphorylated state. Likewise, the mitochondrial membrane potential (Δψm) and the cellular production of reactive oxygen species were higher in embryonic fibroblasts from MPV17(-/-) mice. However, despite the elevated Δψm, the MPV17-deficient mitochondria showed signs of accelerated fission. Together, these observations uncover the role of MPV17 as a Δψm-modulating channel that apparently contributes to mitochondrial homeostasis under different conditions.
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The role of the MPV17 protein mutations of which cause mitochondrial DNA depletion syndrome (MDDS): lessons from homologs in different species.
Biological chemistry, 2015Co-Authors: Stefanie Löllgen, Hans WeiherAbstract:Mitochondrial DNA depletion syndromes (MDDS) are severe pediatric diseases with diverse clinical manifestations. Gene mutations that underlie MDDS have been associated with alterations in the mitochondrial DNA (mtDNA) replication machinery or in mitochondrial deoxyribonucleoside triphosphate pools. However, the nuclear gene MPV17, whose mutated forms are associated with hepatocerebral MDDS in humans, plays a so-far unknown role in mtDNA maintenance. A high degree of conservation has been determined between MPV17 and its mouse (MPV17), zebrafish (tra) and yeast (SYM1) homologs, respectively, whereby mutants in these cause very different phenotypes. While dysfunction in this gene in humans causes fatal liver disease, kidney pathology is induced in mice. Moreover, in zebrafish inactivation of the MPV17 homolog was detected as a viable dyscolouration mutant. Knock out of the yeast ortholog results in a temperature-sensitive metabolic growth phenotype. Detailed analyses on common denominators between these different phenotypes strengthen the hypothesis that the MPV17 protein forms a channel in the inner mitochondrial membrane, allowing small molecules - in vertebrates probably nucleotides, and in yeast probably intermediates of the tricarboxylic acid cycle - to pass. Moreover, a function modifying the pathologic manifestations of MPV17-related disease in mice has been identified. This signaling pathway remarkably involves the non-mitochondrial catalytic subunit of DNA-dependent protein kinase (PRKDC), important in double-strand break repair resistance against reactive oxygen-induced genotoxic stress.
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MPV17 encodes an inner mitochondrial membrane protein and is mutated in infantile hepatic mitochondrial dna depletion
Nature Genetics, 2006Co-Authors: Antonella Spinazzola, Carlo Viscomi, Erika Fernandezvizarra, Franco Carrara, Pio Dadamo, Sarah E Calvo, Rene Massimiliano Marsano, Claudia Donnini, Hans WeiherAbstract:The mitochondrial (mt) DNA depletion syndromes (MDDS) are genetic disorders characterized by a severe, tissue-specific decrease of mtDNA copy number, leading to organ failure. There are two main clinical presentations: myopathic (OMIM 609560) and hepatocerebral1 (OMIM 251880). Known mutant genes, including TK2 (ref. 2), SUCLA2 (ref. 3), DGUOK (ref. 4) and POLG5,6, account for only a fraction of MDDS cases7. We found a new locus for hepatocerebral MDDS on chromosome 2p21-23 and prioritized the genes on this locus using a new integrative genomics strategy. One of the top-scoring candidates was the human ortholog of the mouse kidney disease gene MPV17 (ref. 8). We found disease-segregating mutations in three families with hepatocerebral MDDS and demonstrated that, contrary to the alleged peroxisomal localization of the MPV17 gene product9, MPV17 is a mitochondrial inner membrane protein, and its absence or malfunction causes oxidative phosphorylation (OXPHOS) failure and mtDNA depletion, not only in affected individuals but also in MPV17−/− mice.
Koichiro Kishi - One of the best experts on this subject based on the ideXlab platform.
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Human MPV17-like protein is localized in peroxisomes and regulates expression of antioxidant enzymes.
Biochemical and biophysical research communications, 2006Co-Authors: Reiko Iida, Toshihiro Yasuda, Etsuko Tsubota, Hisakazu Takatsuka, Takasumi Matsuki, Koichiro KishiAbstract:M-LP (MPV17-like protein) is a protein that was initially identified in mouse tissues and shows high sequence homology with MPV17 protein, a peroxisomal membrane protein involved in the development of early-onset glomerulosclerosis [R. Iida, T. Yasuda, E. Tsubota, H. Takatsuka, M. Masuyama, T. Matsuki, K. Kishi, M-LP, MPV17-like protein, has a peroxisomal membrane targeting signal comprising a transmembrane domain and a positively charged loop and up-regulates expression of the manganese superoxide dismutase gene, J. Biol. Chem. 278 (2003) 6301–6306]. Here we report the identification and characterization of a human homolog of the M-LP (M-LPH) gene. The M-LPH gene is composed of four exons, extends over 14 kb on chromosome 16p13.1, and is expressed as two alternatively spliced variants comprising four and three exons, respectively, which include open-reading frames encoding two distinct isoforms composed of 196 (M-LPH1) and 147 (M-LPH2) amino acids, respectively. These two variants were expressed ubiquitously in human tissues, however only M-LPH1 was detected at the protein level. Dual-color confocal analysis of COS-7 cells transfected with a green fluorescent protein-tagged M-LPH1 demonstrated that M-LPH1 is localized in peroxisomes. In order to elucidate the function of M-LPH1, we examined the mRNA levels of several enzymes involved in the metabolism of reactive oxygen species in COS-7 cells and found that transfection with M-LPH1 down-regulates expression of the plasma glutathione peroxidase and catalase genes. These results show the existence of the human homolog of M-LP and its participation in reactive oxygen species metabolism.
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A novel alternative spliced MPV17-like protein isoform localizes in cytosol and is expressed in a kidney- and adult-specific manner.
Experimental cell research, 2005Co-Authors: Reiko Iida, Toshihiro Yasuda, Etsuko Tsubota, Hisakazu Takatsuka, Mika Masuyama, Takasumi Matsuki, Koichiro KishiAbstract:Abstract MPV17-like protein (M-LP) has been identified as a new protein that shows high sequence homology with MPV17 protein, a peroxisomal membrane protein involved in the development of early onset glomerulosclerosis. We previously showed that the originally identified M-LP isoform, designated M-LP L , is, like MPV17, localized in peroxisomes, and that transfection with M-LP L up-regulates expression of the manganese superoxide dismutase (SOD2) gene [R. Iida, T. Yasuda, E. Tsubota, H. Takatsuka, M. Masuyama, T. Matsuki, K. Kishi, M-LP, MPV17-like protein, has a peroxisomal membrane targeting signal comprising a transmembrane domain and a positively charged loop and up-regulates expression of the manganese superoxide dismutase gene. J. Biol. Chem. 278 (2003) 6301–6306.]. We report here the identification of a novel alternative splicing product of the M-LP gene, designated M-LP S . A comparison of the genomic sequence with the cDNA sequences and an analysis of 5'-flanking regions revealed that the two isoforms are generated by alternative usage of two promoters. M-LP S consists of the C-terminal half of M-LP L (90 amino acids) and therefore lacks the peroxisome targeting signal of membrane protein that exists near the N-terminus of M-LP L. Expression of green fluorescent protein-tagged M-LP S in COS-7 cells demonstrated that M-LP S localizes in the cytosol. In mice, M-LP S is expressed exclusively in kidneys after the age of 6 weeks. Moreover, quantitative real-time PCR analysis revealed that transfection with M-LP S up-regulates expression of the SOD2 gene and down-regulates expression of the cellular glutathione peroxidase (Gpx1) and plasma glutathione peroxidase (Gpx3) genes. Taken together, these results suggest different functional attributes of the two M-LP isoforms during aging and development.
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m lp MPV17 like protein has a peroxisomal membrane targeting signal comprising a transmembrane domain and a positively charged loop and up regulates expression of the manganese superoxide dismutase gene
Journal of Biological Chemistry, 2003Co-Authors: Reiko Iida, Toshihiro Yasuda, Etsuko Tsubota, Hisakazu Takatsuka, Mika Masuyama, Takasumi Matsuki, Koichiro KishiAbstract:M-LP (MPV17-like protein) has been identified as a new protein that has high sequence homology with MPV17 protein, a peroxisomal membrane protein involved in the development of early onset glomerulosclerosis. In this study, we verified the peroxisomal localization of M-LP by performing dual-color confocal analysis of COS-7 cells cotransfected with green fluorescent protein-tagged M-LP and DsRED2-PTS1, a red fluorescent peroxisomal marker. To characterize the peroxisomal membrane targeting signal, we examined the intracellular localizations of several green fluorescent protein-tagged deletion mutants and demonstrated that, of the three transmembrane segments predicted, the first near the NH(2) terminus and NH(2)-terminal half of the following loop region, which is abundant in positively charged amino acids, were necessary and sufficient for peroxisomal targeting. To elucidate the function of M-LP, we examined the activities of several enzymes involved in reactive oxygen species metabolism in COS-7 cells and found that transfection with M-LP increased the superoxide dismutase activity significantly. Quantitative real-time PCR analysis revealed that the manganese SOD (SOD2) mRNA level of COS-7 cells transfected with M-LP was elevated. These results indicate that M-LP participates in reactive oxygen species metabolism.
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Cloning, mapping, genomic organization, and expression of mouse M-LP, a new member of the peroxisomal membrane protein MPV17 domain family.
Biochemical and biophysical research communications, 2001Co-Authors: Reiko Iida, Toshihiro Yasuda, Etsuko Tsubota, Takasumi Matsuki, Koichiro KishiAbstract:We have identified a mouse full-length cDNA and gene encoding a novel protein (M-LP), based on an expressed sequence tag (EST) sequence (GenBank Accession No. AI482564) obtained by differential display screening of age-dependently expressed genes in mouse kidney. The ML-P gene is composed of three exons, ranges over 5 kb on mouse chromosome 16B1-B2 and is expressed as two transcripts (1455 and 3058 bp), both of which include the same open-reading frame encoding 194 amino acids. M-LP is expressed mainly in kidney and spleen and shows age-dependent expression. M-LP has sequence homologies and membrane topologies very similar to the MPV17 protein, a peroxisomal protein involved in the development of early-onset glomerulosclerosis. Search of the protein domain family database (ProDom) revealed that M-LP is a new member of the MPV17 domain family (PD008400).
Ralf M. Zwacka - One of the best experts on this subject based on the ideXlab platform.
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A monoclonal antibody raised against bacterially expressed MPV17 sequences shows peroxisomal, endosomal and lysosomal localisation in U2OS cells
BMC research notes, 2016Co-Authors: Hans Weiher, Ralf M. Zwacka, Miia Vapola, J. Kalervo Hiltunen, Haymo Pircher, Pidder Jansen-dürr, Silke Hegenbarth, Percy A. Knolle, Silke Grunau, Elmon SchmelzerAbstract:Recessive mutations in the MPV17 gene cause mitochondrial DNA depletion syndrome, a fatal infantile genetic liver disease in humans. Loss of function in mice leads to glomerulosclerosis and sensineural deafness accompanied with mitochondrial DNA depletion. Mutations in the yeast homolog Sym1, and in the zebra fish homolog tra cause interesting, but not obviously related phenotypes, although the human gene can complement the yeast Sym1 mutation. The MPV17 protein is a hydrophobic membrane protein of 176 amino acids and unknown function. Initially localised in murine peroxisomes, it was later reported to be a mitochondrial inner membrane protein in humans and in yeast. To resolve this contradiction we tested two new mouse monoclonal antibodies directed against the human MPV17 protein in Western blots and immunohistochemistry on human U2OS cells. One of these monoclonal antibodies showed specific reactivity to a protein of 20 kD absent in MPV17 negative mouse cells. Immunofluorescence studies revealed colocalisation with peroxisomal, endosomal and lysosomal markers, but not with mitochondria. This data reveal a novel connection between a possible peroxisomal/endosomal/lysosomal function and mitochondrial DNA depletion.
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Age-dependent hypertension in MPV17-deficient mice, a transgenic model of glomerulosclerosis and inner ear disease.
Experimental gerontology, 1999Co-Authors: Martine Clozel, Ralf M. Zwacka, Alexander Reuter, Patrick Hess, Walter Fischli, Loeffler Bernd-michael, Hans WeiherAbstract:Abstract The mutant mouse strain MPV17−/−, carries a retroviral germline integration that inactivates the MPV17 gene. MPV17-deficient mice develop progressive glomerulosclerosis and sensineural deafness at early age. Characteristic basement membrane alterations are found in both sites of pathology. MPV17 is a peroxisomal protein involved in the metabolism of reactive oxygen species, yet its molecular function is unknown. Dysregulation of antioxidant enzymes and basal membrane components has been established in this model and successful therapeutic intervention with antioxidants prove the causal role of reactive oxygen species in the development of the disease phenotype. We here investigated if the MPV17−/ − mice might be hypertensive. Indeed, our study revealed that MPV17−/ − mice developed significant systemic hypertension and tachycardia between 4 weeks and 5 months of age, accompanied by polyuria and elevated natriuresis. Judging from serum and urine parameters, the hypertensive condition develops concommittantly with the renal disease. Biochemical and pharmacological studies that used the endothelin receptor antagonist bosentan and the angiotensin converting enzyme inhibitor cilazapril indicated no involvement of the endothelin and renin-angiotensin systems in this hypertension, suggesting a potential novel mechanism of blood pressure regulation in this new murine hypertension model. Thus, MPV17−/− mice unravel an intriguing new association between a defect in reactive oxygen metabolism and the age-dependent development of hypertension.
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Expression of the Recessive Glomerulosclerosis Gene MPV17 Regulates MMP-2 Expression in Fibroblasts, the Kidney, and the Inner Ear of Mice
Molecular biology of the cell, 1998Co-Authors: Alexander Reuter, Ralf M. Zwacka, Rüdiger Waldherr, Andrea Nestl, Jan P. Tuckermann, Eva Maria Wagner, Matti Höyhtyä, Angela M Meyer Zum Gottesberge, Peter Angel, Hans WeiherAbstract:The recessive mouse mutant MPV17 is characterized by the development of early-onset glomerulosclerosis, concomitant hypertension, and structural alterations of the inner ear. The primary cause of the disease is the loss of function of the MPV17 protein, a peroxisomal gene product involved in reactive oxygen metabolism. In our search of a common mediator exerting effects on several aspects of the phenotype, we discovered that the absence of the MPV17 gene product causes a strong increase in matrix metalloproteinase 2 (MMP-2) expression. This was seen in the kidney and cochlea of MPV17-negative mice as well as in tissue culture cells derived from these animals. When these cells were transfected with the human MPV17 homolog, an inverse causal relationship between MPV17 and MMP-2 expression was established. These results indicate that the MPV17 protein plays a crucial role in the regulation of MMP-2 and suggest that enhanced MMP-2 expression might mediate the mechanisms leading to glomerulosclerosis, inner ear disease, and hypertension in this model.
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Loss of auditory function in transgenic MPV17-deficient mice.
Hearing research, 1997Co-Authors: M Müller, Ralf M. Zwacka, A. Reuter, Hans Weiher, J W Smolders, A M Meyer Zum Gottesberge, R KlinkeAbstract:The transgenic mouse strain MPV17 develops severe morphological degeneration of the inner ear and nephrotic syndrome at a young age (Meyer zum Gottesberge et al., 1996; Weiher et al., 1990). The audiograms (1-32 kHz) of MPV17-negative mice were determined from auditory brain stem responses in young (2 months) and old (7 months) animals. Audiograms of age-matched wild-type mice with the same genetic background, but wild-type at the MPV17 locus, were also determined. Furthermore, young MPV17-negative mice that carried a human MPV17 homologue gene were studied. NMRI mice served as a reference for normal hearing. MPV17-negative mice suffer from severe sensorineural hearing loss as early as 2 months after birth. In the old MPV17-negative mice no responses could be elicited at all. The 2 month old wild-type mice had normal audiograms, at 7 months only high threshold responses were seen. The poor audiograms of the MPV17-negative mice are assumed to be the functional correlate of the morphological degeneration of the cochlea described earlier (Meyer zum Gottesberge et al., 1996). The finding that 2 out of 4 MPV17-negative mice with the human MPV17 gene had normal audiograms, shows that the gene inactivation can be functionally compensated by the human MPV17 gene product.
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Functional rescue of the glomerulosclerosis phenotype in MPV17 mice by transgenesis with the human MPV17 homologue
Kidney international, 1995Co-Authors: Johannes Schenkel, Ralf M. Zwacka, Christiane Rutenberg, Alexander Reuter, Rüdiger Waldherr, Hans WeiherAbstract:Functional rescue of the glomerulosclerosis phenotype in MPV17 mice by transgenesis with the human MPV17 homologue. The germ line insertion of a defective retrovirus into the MPV17 gene of mice is associated with a recessive phenotype. Mice homozygous for the integration develop glomerulosclerosis at a young age. The phenotype resembles human glomerulosclerosis in its physiological parameters as well as in histology. A human homologue of the MPV17 gene has been identified, isolated and analyzed. We here show that this gene, which has a role in the production of reactive oxygen species, can rescue the phenotype of MPV17 deficient mice when introduced by transgenesis. This provides formal proof for the hypothesis that the phenotype is caused by the loss of function of the MPV17 gene. It also provides evidence for the functional conservation of the MPV17 gene in mammals and points to a potential role of this gene in human kidney disease.
Claudia Donnini - One of the best experts on this subject based on the ideXlab platform.
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Pathological alleles of MPV17 modeled in the yeast Saccharomyces cerevisiae orthologous gene SYM1 reveal their inability to take part in a high molecular weight complex.
PloS one, 2018Co-Authors: Micol Gilberti, Claudia Donnini, Enrico Baruffini, Cristina DallabonaAbstract:Mitochondrial DNA depletion syndromes (MDDS) are a genetically and clinically heterogeneous group of human diseases caused by mutations in nuclear genes and characterized by a severe reduction in mitochondrial DNA (mtDNA) copy number leading to impaired energy production in affected tissues and organs. Mutations in the MPV17 gene, whose role is still elusive, were described as cause of the hepatocerebral form of MDDS and Navajo neuro-hepathopathy. The high degree of conservation observed between MPV17 and its yeast homolog SYM1 made the latter a good model for the study of the pathology. Here, we used Saccharomyces cerevisiae to elucidate the molecular consequences of seven MPV17 missense mutations identified in patients and localized in different protein domains. The phenotypic analysis of the appropriate sym1 mutant strains created demonstrated deleterious effect for all mutations regarding OXPHOS metabolism and mtDNA stability. We deepened the pathogenic effect of the mutations by investigating whether they prevented the correct protein localization into the mitochondria or affected the stability of the proteins. All the Sym1 mutant proteins correctly localized into the mitochondria and only one mutation predominantly affects protein stability. All the other mutations compromised the formation of the high molecular weight complex of unknown composition, previously identified both in yeast, cell cultures and mouse tissues, as demonstrated by the consistent fraction of the Sym1 mutant proteins found free or in not fully assembled complex, strengthening its role as protein forming part of a high molecular weight complex.
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Sym1, the yeast ortholog of the MPV17 human disease protein, is a stress-induced bioenergetic and morphogenetic mitochondrial modulator
Human molecular genetics, 2009Co-Authors: Cristina Dallabona, Massimo Zeviani, Rene Massimiliano Marsano, Paola Arzuffi, Daniele Ghezzi, Patrizia Mancini, Iliana Ferrero, Claudia DonniniAbstract:A peculiar form of hepatocerebral mtDNA depletion syndrome is caused by mutations in the MPV17 gene, which encodes a small hydrophobic protein of unknown function located in the mitochondrial inner membrane. In order to define the molecular basis of MPV17 variants associated with the human disorder, we have previously taken advantage of S. cerevisiae as a model system thanks to the presence of an MPV17 ortholog gene, SYM1. We demonstrate here that the SYM1 gene product is essential to maintain OXPHOS, glycogen storage, mitochondrial morphology and mtDNA stability in stressing conditions such as high temperature and ethanol-dependent growth. To gain insight into the molecular basis of the Sym1-less phenotype, we identified and characterized multicopy suppressor genes and metabolic suppressor compounds. Our results suggest that (i) metabolic impairment and mtDNA instability occur independently from each other as a consequence of SYM1 ablation; (ii) ablation of Sym1 causes depletion of glycogen storage, possibly due to defective anaplerotic flux of tricarboxylic acid (TCA) cycle intermediates to the cytosol; (iii) flattening of mitochondrial cristae in Sym1-defective organelles suggests a role for Sym1 in the structural preservation of the inner mitochondrial membrane, which could in turn control mtDNA maintenance and stability.
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MPV17 encodes an inner mitochondrial membrane protein and is mutated in infantile hepatic mitochondrial dna depletion
Nature Genetics, 2006Co-Authors: Antonella Spinazzola, Carlo Viscomi, Erika Fernandezvizarra, Franco Carrara, Pio Dadamo, Sarah E Calvo, Rene Massimiliano Marsano, Claudia Donnini, Hans WeiherAbstract:The mitochondrial (mt) DNA depletion syndromes (MDDS) are genetic disorders characterized by a severe, tissue-specific decrease of mtDNA copy number, leading to organ failure. There are two main clinical presentations: myopathic (OMIM 609560) and hepatocerebral1 (OMIM 251880). Known mutant genes, including TK2 (ref. 2), SUCLA2 (ref. 3), DGUOK (ref. 4) and POLG5,6, account for only a fraction of MDDS cases7. We found a new locus for hepatocerebral MDDS on chromosome 2p21-23 and prioritized the genes on this locus using a new integrative genomics strategy. One of the top-scoring candidates was the human ortholog of the mouse kidney disease gene MPV17 (ref. 8). We found disease-segregating mutations in three families with hepatocerebral MDDS and demonstrated that, contrary to the alleged peroxisomal localization of the MPV17 gene product9, MPV17 is a mitochondrial inner membrane protein, and its absence or malfunction causes oxidative phosphorylation (OXPHOS) failure and mtDNA depletion, not only in affected individuals but also in MPV17−/− mice.
Reiko Iida - One of the best experts on this subject based on the ideXlab platform.
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Human MPV17-like protein with a mitigating effect on mtDNA damage is involved in cAMP/PKA signaling in the mitochondrial matrix.
Biochimica et biophysica acta. Molecular cell research, 2020Co-Authors: Reiko Iida, Misuzu Ueki, Toshihiro YasudaAbstract:Human MPV17-like protein (M-LPH/MPV17L) is thought to play a role in minimizing mitochondrial dysfunction caused by mitochondrial DNA (mtDNA) damage. We have recently demonstrated that, in addition to an increase of mtDNA damage, M-LPH-knockout (M-LPH-KO) in HepG2 cells causes a significant reduction of mitochondrial transcription factor A (TFAM) protein, an essential factor for mtDNA maintenance, along with an increase in its phosphorylation. These intracellular changes suggested an association of M-LPH with the cAMP/PKA signaling pathway, as selective degradation of TFAM by mitochondrial protease is driven by protein kinase A (PKA)-dependent phosphorylation. In the present study, we observed that M-LPH-KO in HepG2 cells caused an increase in the level of mitochondrial cAMP and a reduction of total cellular cyclic nucleotide phosphodiesterase (PDE) activity. In vitro-synthesized M-LPH showed PDE activity, which was inhibited by IBMX, a non-selective inhibitor of PDE. Furthermore, M-LPH-KO promoted PKA-dependent phosphorylation of some mitochondrial proteins. Taken together, the present findings suggest that M-LPH, which has structural features atypical of PDE family members, might be a novel human PDE involved in cAMP/PKA signaling in the mitochondrial matrix.
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human MPV17 like protein with a mitigating effect on mtdna damage is involved in camp pka signaling in the mitochondrial matrix
Biochimica et Biophysica Acta, 2020Co-Authors: Reiko Iida, Misuzu Ueki, Toshihiro YasudaAbstract:Human MPV17-like protein (M-LPH/MPV17L) is thought to play a role in minimizing mitochondrial dysfunction caused by mitochondrial DNA (mtDNA) damage. We have recently demonstrated that, in addition to an increase of mtDNA damage, M-LPH-knockout (M-LPH-KO) in HepG2 cells causes a significant reduction of mitochondrial transcription factor A (TFAM) protein, an essential factor for mtDNA maintenance, along with an increase in its phosphorylation. These intracellular changes suggested an association of M-LPH with the cAMP/PKA signaling pathway, as selective degradation of TFAM by mitochondrial protease is driven by protein kinase A (PKA)-dependent phosphorylation. In the present study, we observed that M-LPH-KO in HepG2 cells caused an increase in the level of mitochondrial cAMP and a reduction of total cellular cyclic nucleotide phosphodiesterase (PDE) activity. In vitro-synthesized M-LPH showed PDE activity, which was inhibited by IBMX, a non-selective inhibitor of PDE. Furthermore, M-LPH-KO promoted PKA-dependent phosphorylation of some mitochondrial proteins. Taken together, the present findings suggest that M-LPH, which has structural features atypical of PDE family members, might be a novel human PDE involved in cAMP/PKA signaling in the mitochondrial matrix.
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Human MPV17-like protein is localized in peroxisomes and regulates expression of antioxidant enzymes.
Biochemical and biophysical research communications, 2006Co-Authors: Reiko Iida, Toshihiro Yasuda, Etsuko Tsubota, Hisakazu Takatsuka, Takasumi Matsuki, Koichiro KishiAbstract:M-LP (MPV17-like protein) is a protein that was initially identified in mouse tissues and shows high sequence homology with MPV17 protein, a peroxisomal membrane protein involved in the development of early-onset glomerulosclerosis [R. Iida, T. Yasuda, E. Tsubota, H. Takatsuka, M. Masuyama, T. Matsuki, K. Kishi, M-LP, MPV17-like protein, has a peroxisomal membrane targeting signal comprising a transmembrane domain and a positively charged loop and up-regulates expression of the manganese superoxide dismutase gene, J. Biol. Chem. 278 (2003) 6301–6306]. Here we report the identification and characterization of a human homolog of the M-LP (M-LPH) gene. The M-LPH gene is composed of four exons, extends over 14 kb on chromosome 16p13.1, and is expressed as two alternatively spliced variants comprising four and three exons, respectively, which include open-reading frames encoding two distinct isoforms composed of 196 (M-LPH1) and 147 (M-LPH2) amino acids, respectively. These two variants were expressed ubiquitously in human tissues, however only M-LPH1 was detected at the protein level. Dual-color confocal analysis of COS-7 cells transfected with a green fluorescent protein-tagged M-LPH1 demonstrated that M-LPH1 is localized in peroxisomes. In order to elucidate the function of M-LPH1, we examined the mRNA levels of several enzymes involved in the metabolism of reactive oxygen species in COS-7 cells and found that transfection with M-LPH1 down-regulates expression of the plasma glutathione peroxidase and catalase genes. These results show the existence of the human homolog of M-LP and its participation in reactive oxygen species metabolism.
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A novel alternative spliced MPV17-like protein isoform localizes in cytosol and is expressed in a kidney- and adult-specific manner.
Experimental cell research, 2005Co-Authors: Reiko Iida, Toshihiro Yasuda, Etsuko Tsubota, Hisakazu Takatsuka, Mika Masuyama, Takasumi Matsuki, Koichiro KishiAbstract:Abstract MPV17-like protein (M-LP) has been identified as a new protein that shows high sequence homology with MPV17 protein, a peroxisomal membrane protein involved in the development of early onset glomerulosclerosis. We previously showed that the originally identified M-LP isoform, designated M-LP L , is, like MPV17, localized in peroxisomes, and that transfection with M-LP L up-regulates expression of the manganese superoxide dismutase (SOD2) gene [R. Iida, T. Yasuda, E. Tsubota, H. Takatsuka, M. Masuyama, T. Matsuki, K. Kishi, M-LP, MPV17-like protein, has a peroxisomal membrane targeting signal comprising a transmembrane domain and a positively charged loop and up-regulates expression of the manganese superoxide dismutase gene. J. Biol. Chem. 278 (2003) 6301–6306.]. We report here the identification of a novel alternative splicing product of the M-LP gene, designated M-LP S . A comparison of the genomic sequence with the cDNA sequences and an analysis of 5'-flanking regions revealed that the two isoforms are generated by alternative usage of two promoters. M-LP S consists of the C-terminal half of M-LP L (90 amino acids) and therefore lacks the peroxisome targeting signal of membrane protein that exists near the N-terminus of M-LP L. Expression of green fluorescent protein-tagged M-LP S in COS-7 cells demonstrated that M-LP S localizes in the cytosol. In mice, M-LP S is expressed exclusively in kidneys after the age of 6 weeks. Moreover, quantitative real-time PCR analysis revealed that transfection with M-LP S up-regulates expression of the SOD2 gene and down-regulates expression of the cellular glutathione peroxidase (Gpx1) and plasma glutathione peroxidase (Gpx3) genes. Taken together, these results suggest different functional attributes of the two M-LP isoforms during aging and development.
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m lp MPV17 like protein has a peroxisomal membrane targeting signal comprising a transmembrane domain and a positively charged loop and up regulates expression of the manganese superoxide dismutase gene
Journal of Biological Chemistry, 2003Co-Authors: Reiko Iida, Toshihiro Yasuda, Etsuko Tsubota, Hisakazu Takatsuka, Mika Masuyama, Takasumi Matsuki, Koichiro KishiAbstract:M-LP (MPV17-like protein) has been identified as a new protein that has high sequence homology with MPV17 protein, a peroxisomal membrane protein involved in the development of early onset glomerulosclerosis. In this study, we verified the peroxisomal localization of M-LP by performing dual-color confocal analysis of COS-7 cells cotransfected with green fluorescent protein-tagged M-LP and DsRED2-PTS1, a red fluorescent peroxisomal marker. To characterize the peroxisomal membrane targeting signal, we examined the intracellular localizations of several green fluorescent protein-tagged deletion mutants and demonstrated that, of the three transmembrane segments predicted, the first near the NH(2) terminus and NH(2)-terminal half of the following loop region, which is abundant in positively charged amino acids, were necessary and sufficient for peroxisomal targeting. To elucidate the function of M-LP, we examined the activities of several enzymes involved in reactive oxygen species metabolism in COS-7 cells and found that transfection with M-LP increased the superoxide dismutase activity significantly. Quantitative real-time PCR analysis revealed that the manganese SOD (SOD2) mRNA level of COS-7 cells transfected with M-LP was elevated. These results indicate that M-LP participates in reactive oxygen species metabolism.