111 results on '"Tabak HF"'
Search Results
2. Peroxisomes: offshoots of the ER.
- Author
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van der Zand A and Tabak HF
- Subjects
- Animals, Endoplasmic Reticulum chemistry, Eukaryota metabolism, Membrane Proteins metabolism, Peroxisomes chemistry, Yeasts cytology, Yeasts metabolism, Endoplasmic Reticulum metabolism, Eukaryota cytology, Peroxisomes metabolism
- Abstract
Peroxisomes are part of the ubiquitous set of eukaryotic organelles. They are small, single membrane bounded vesicles, specialized in the degradation of very-long-chain fatty acids and in synthesis of myelin lipids. Once considered inconspicuous, recent new insights in the formation and function of peroxisomes have revealed a much more subtle interplay between organelles that warrant a re-evaluation of the historical assignment of peroxisomes as being either autonomous or ER-derived. Peroxisomes acquire their lipids and membrane proteins from the ER, whereas they import their matrix proteins directly from the cytosol. Remarkably, many of its metabolic enzymes and factors controlling peroxisome abundance (fission and inheritance) too are shared with other organelles, stressing interdependence among cellular compartments., (Copyright © 2013 Elsevier Ltd. All rights reserved.)
- Published
- 2013
- Full Text
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3. Peroxisome formation and maintenance are dependent on the endoplasmic reticulum.
- Author
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Tabak HF, Braakman I, and van der Zand A
- Subjects
- Animals, Endoplasmic Reticulum physiology, Humans, Peroxisomes physiology, Protein Transport, Endoplasmic Reticulum metabolism, Membrane Proteins metabolism, Peroxisomes metabolism
- Abstract
Looks can be deceiving. Although peroxisomes appear to be simple organelles, their formation and maintenance pose unique challenges for the cell. The birth of new peroxisomes starts at the endoplasmic reticulum (ER), which delivers lipids and membrane proteins. To form a new peroxisomal compartment, ER-derived preperoxisomal vesicles carrying different membrane proteins fuse, allowing the assembly of the peroxisomal translocon. To complete formation, peroxisomes import their soluble proteins directly from the cytosol using the newly assembled translocon. Together with the ER-derived biogenic route, peroxisomal fission and segregation subsequently maintain the cellular peroxisome population. In this review we highlight the latest insights on the life cycle of peroxisomes and show how the new cell biology concept of peroxisome formation affects our thinking about peroxisome-related diseases and their evolutionary past. The future challenge lies in the identification of all the proteins involved in this elaborate biogenic process and the dissection of their mechanism of action.
- Published
- 2013
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4. Biochemically distinct vesicles from the endoplasmic reticulum fuse to form peroxisomes.
- Author
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van der Zand A, Gent J, Braakman I, and Tabak HF
- Subjects
- Cytoplasmic Vesicles metabolism, Membrane Proteins metabolism, Models, Biological, Saccharomyces cerevisiae Proteins metabolism, Endoplasmic Reticulum metabolism, Peroxisomes metabolism, Saccharomyces cerevisiae cytology, Saccharomyces cerevisiae metabolism
- Abstract
As a rule, organelles in eukaryotic cells can derive only from pre-existing organelles. Peroxisomes are unique because they acquire their lipids and membrane proteins from the endoplasmic reticulum (ER), whereas they import their matrix proteins directly from the cytosol. We have discovered that peroxisomes are formed via heterotypic fusion of at least two biochemically distinct preperoxisomal vesicle pools that arise from the ER. These vesicles each carry half a peroxisomal translocon complex. Their fusion initiates assembly of the full peroxisomal translocon and subsequent uptake of enzymes from the cytosol. Our findings demonstrate a remarkable mechanism to maintain biochemical identity of organelles by transporting crucial components via different routes to their final destination., (Copyright © 2012 Elsevier Inc. All rights reserved.)
- Published
- 2012
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5. Peroxisomal membrane proteins insert into the endoplasmic reticulum.
- Author
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van der Zand A, Braakman I, and Tabak HF
- Subjects
- Intracellular Membranes metabolism, Membrane Proteins chemistry, Protein Transport, Saccharomyces cerevisiae cytology, Endoplasmic Reticulum metabolism, Membrane Proteins metabolism, Peroxisomes metabolism, Saccharomyces cerevisiae metabolism, Saccharomyces cerevisiae Proteins metabolism
- Abstract
We show that a comprehensive set of 16 peroxisomal membrane proteins (PMPs) encompassing all types of membrane topologies first target to the endoplasmic reticulum (ER) in Saccharomyces cerevisiae. These PMPs insert into the ER membrane via the protein import complexes Sec61p and Get3p (for tail-anchored proteins). This trafficking pathway is representative for multiplying wild-type cells in which the peroxisome population needs to be maintained, as well as for mutant cells lacking peroxisomes in which new peroxisomes form after complementation with the wild-type version of the mutant gene. PMPs leave the ER in a Pex3p-Pex19p-dependent manner to end up in metabolically active peroxisomes. These results further extend the new concept that peroxisomes derive their basic framework (membrane and membrane proteins) from the ER and imply a new functional role for Pex3p and Pex19p.
- Published
- 2010
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6. Peroxisomes: minted by the ER.
- Author
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Tabak HF, van der Zand A, and Braakman I
- Subjects
- Animals, Humans, Endoplasmic Reticulum physiology, Peroxisomes physiology
- Abstract
Peroxisomes are one of numerous organelles in a eukaryotic cell; they are small, single-membrane-bound vesicles involved in cellular metabolism, particularly fatty acid degradation. Transport of metabolites and co-factors in and across the membrane is taken care of by specific transporters. Peroxisome formation and maintenance has been debated for a long time: opinions swinging from autonomous to ER-derived organelles. Only recently it has been established firmly that the site of origin of peroxisomes is the ER. It implies that a new branch of the endomembrane system is open to further characterization.
- Published
- 2008
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7. Formation of peroxisomes: present and past.
- Author
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Tabak HF, Hoepfner D, Zand Av, Geuze HJ, Braakman I, and Huynen MA
- Subjects
- Animals, Biological Evolution, Endoplasmic Reticulum physiology, Humans, Peroxisomes physiology, Phylogeny
- Abstract
Eukaryotic cells contain functionally distinct, membrane enclosed compartments called organelles. Here we like to address two questions concerning this architectural lay out. How did this membrane complexity arise during evolution and how is this collection of organelles maintained in multiplying cells to ensure that new cells retain a complete set of them. We will try to address these questions with peroxisomes as a focal point of interest.
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- 2006
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8. The return of the peroxisome.
- Author
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van der Zand A, Braakman I, Geuze HJ, and Tabak HF
- Subjects
- Animals, Endoplasmic Reticulum physiology, Fatty Acids metabolism, Humans, Lipid Metabolism, Protein Transport, Peroxisomes physiology
- Abstract
Of the classical compartments of eukaryotic cells, peroxisomes were the last to be discovered. They are small, single-membrane-bound vesicles involved in cellular metabolism, most notably the beta-oxidation of fatty acids. Characterization of their properties and behavior has progressed rather slowly. However, during the past few years, peroxisomes have entered the limelight as a result of several breakthroughs. These include the observations that they are not autonomously multiplying organelles but are derived from the endoplasmic reticulum, and that partitioning of peroxisomes to progeny cells is an active and well-controlled process. In addition, we are discovering more and more proteins that are not only dedicated to peroxisomes but also serve other organelles.
- Published
- 2006
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9. Contribution of the endoplasmic reticulum to peroxisome formation.
- Author
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Hoepfner D, Schildknegt D, Braakman I, Philippsen P, and Tabak HF
- Subjects
- Bacterial Proteins analysis, Bacterial Proteins genetics, Bacterial Proteins metabolism, Endoplasmic Reticulum ultrastructure, Luminescent Proteins analysis, Luminescent Proteins genetics, Luminescent Proteins metabolism, Membrane Proteins genetics, Membrane Proteins metabolism, Microscopy, Fluorescence, Peroxins, Peroxisomes ultrastructure, Recombinant Fusion Proteins analysis, Recombinant Fusion Proteins genetics, Recombinant Fusion Proteins metabolism, Saccharomyces cerevisiae genetics, Saccharomyces cerevisiae metabolism, Saccharomyces cerevisiae Proteins genetics, Saccharomyces cerevisiae Proteins metabolism, Endoplasmic Reticulum metabolism, Peroxisomes metabolism
- Abstract
How peroxisomes are formed in eukaryotic cells is unknown but important for insight into a variety of diseases. Both human and yeast cells lacking peroxisomes due to mutations in PEX3 or PEX19 genes regenerate the organelles upon reintroduction of the corresponding wild-type version. To evaluate how and from where new peroxisomes are formed, we followed the trafficking route of newly made YFP-tagged Pex3 and Pex19 proteins by real-time fluorescence microscopy in Saccharomyces cerevisiae. Remarkably, Pex3 (an integral membrane protein) could first be observed in the endoplasmic reticulum (ER), where it concentrates in foci that then bud off in a Pex19-dependent manner and mature into fully functional peroxisomes. Pex19 (a farnesylated, mostly cytosolic protein) enriches first at the Pex3 foci on the ER and then on the maturing peroxisomes. This trafficking route of Pex3-YFP is the same in wild-type cells. These results demonstrate that peroxisomes are generated from domains in the ER.
- Published
- 2005
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10. Peroxisomes start their life in the endoplasmic reticulum.
- Author
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Tabak HF, Murk JL, Braakman I, and Geuze HJ
- Subjects
- Animals, Dendritic Cells physiology, Dendritic Cells ultrastructure, Endoplasmic Reticulum ultrastructure, Mice, Microscopy, Electron, Peroxisomes ultrastructure, Endoplasmic Reticulum physiology, Peroxisomes physiology
- Abstract
Peroxisomes belong to the ubiquitous organelle repertoire of eukaryotic cells. They contribute to cellular metabolism in various ways depending on species, but a consistent feature is the presence of enzymes to degrade fatty acids. Due to the pioneering work of DeDuve and coworkers, peroxisomes were in the limelight of cell biology in the sixties with a focus on their metabolic role. During the last decade, interest in peroxisomes has been growing again, this time with focus on their origin and maintenance. This has resulted in our understanding how peroxisomal proteins are targeted to the organelle and imported into the organellar matrix or recruited into the single membrane surrounding it. With respect to the formation of peroxisomes, the field is divided. The long-held view formulated in 1985 by Lazarow and Fujiki (Lazarow PB, Fujiki Y. Biogenesis of peroxisomes. Annu Rev Cell Biol 1985; 1: 489-530) is that we are dealing with autonomous organelles multiplying by growth and division. This view is being challenged by various observations that call attention to a more active contribution of the ER to peroxisome formation. Our contribution to this debate consists of recent observations using immuno-electronmicroscopy and electron tomography in mouse dendritic cells that show the peroxisomal membrane to be derived from the ER.
- Published
- 2003
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11. Involvement of the endoplasmic reticulum in peroxisome formation.
- Author
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Geuze HJ, Murk JL, Stroobants AK, Griffith JM, Kleijmeer MJ, Koster AJ, Verkleij AJ, Distel B, and Tabak HF
- Subjects
- ATP-Binding Cassette Transporters physiology, Animals, Cells, Cultured, Dendritic Cells physiology, Dendritic Cells ultrastructure, Endoplasmic Reticulum physiology, Image Processing, Computer-Assisted, Membrane Proteins physiology, Mice, Mice, Inbred C57BL, Microscopy, Immunoelectron, Peroxisomes physiology, Endoplasmic Reticulum ultrastructure, Peroxisomes ultrastructure
- Abstract
The traditional view holds that peroxisomes are autonomous organelles multiplying by growth and division. More recently, new observations have challenged this concept. Herein, we present evidence supporting the involvement of the endoplasmic reticulum (ER) in peroxisome formation by electron microscopy, immunocytochemistry and three-dimensional image reconstruction of peroxisomes and associated compartments in mouse dendritic cells. We found the peroxisomal membrane protein Pex13p and the ATP-binding cassette transporter protein PMP70 present in specialized subdomains of the ER that were continuous with a peroxisomal reticulum from which mature peroxisomes arose. The matrix proteins catalase and thiolase were only detectable in the reticula and peroxisomes. Our results suggest the existence of a maturation pathway from the ER to peroxisomes and implicate the ER as a major source from which the peroxisomal membrane is derived.
- Published
- 2003
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12. Pex15p of Saccharomyces cerevisiae provides a molecular basis for recruitment of the AAA peroxin Pex6p to peroxisomal membranes.
- Author
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Birschmann I, Stroobants AK, van den Berg M, Schäfer A, Rosenkranz K, Kunau WH, and Tabak HF
- Subjects
- ATPases Associated with Diverse Cellular Activities, Microscopy, Electron, Peroxisomes ultrastructure, Protein Structure, Tertiary, Saccharomyces cerevisiae ultrastructure, Adenosine Triphosphatases metabolism, Intracellular Membranes metabolism, Membrane Proteins metabolism, Peroxisomes metabolism, Phosphoproteins metabolism, Saccharomyces cerevisiae metabolism, Saccharomyces cerevisiae Proteins
- Abstract
The gene products (peroxins) of at least 29 PEX genes are known to be necessary for peroxisome biogenesis but for most of them their precise function remains to be established. Here we show that Pex15p, an integral peroxisomal membrane protein, in vivo and in vitro binds the AAA peroxin Pex6p. This interaction functionally interconnects these two hitherto unrelated peroxins. Pex15p provides the mechanistic basis for the reversible targeting of Pex6p to peroxisomal membranes. We could demonstrate that the N-terminal part of Pex6p contains the binding site for Pex15p and that the two AAA cassettes D1 and D2 of Pex6p have opposite effects on this interaction. A point mutation in the Walker A motif of D1 (K489A) decreased the binding of Pex6p to Pex15p indicating that the interaction of Pex6p with Pex15p required binding of ATP. Mutations in Walker A (K778A) and B (D831Q) motifs of D2 abolished growth on oleate and led to a considerable larger fraction of peroxisome bound Pex6p. The nature of these mutations suggested that ATP-hydrolysis is required to disconnect Pex6p from Pex15p. On the basis of these results, we propose that Pex6p exerts at least part of its function by an ATP-dependent cycle of recruitment and release to and from Pex15p.
- Published
- 2003
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13. Dissection of transient oxidative stress response in Saccharomyces cerevisiae by using DNA microarrays.
- Author
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Koerkamp MG, Rep M, Bussemaker HJ, Hardy GP, Mul A, Piekarska K, Szigyarto CA, De Mattos JM, and Tabak HF
- Subjects
- Active Transport, Cell Nucleus, Algorithms, DNA-Binding Proteins genetics, DNA-Binding Proteins metabolism, Genes, Fungal, Oleic Acid metabolism, Oxidation-Reduction, Peroxisomes metabolism, RNA, Messenger genetics, RNA, Messenger metabolism, Reactive Oxygen Species metabolism, Recombinant Fusion Proteins genetics, Recombinant Fusion Proteins metabolism, Saccharomyces cerevisiae genetics, Saccharomyces cerevisiae Proteins genetics, Saccharomyces cerevisiae Proteins metabolism, Time Factors, Transcription Factors genetics, Transcription Factors metabolism, Transcription, Genetic, Gene Expression Profiling, Gene Expression Regulation, Fungal, Oligonucleotide Array Sequence Analysis, Oxidative Stress, Saccharomyces cerevisiae physiology
- Abstract
Yeast cells were grown in glucose-limited chemostat cultures and forced to switch to a new carbon source, the fatty acid oleate. Alterations in gene expression were monitored using DNA microarrays combined with bioinformatics tools, among which was included the recently developed algorithm REDUCE. Immediately after the switch to oleate, a transient and very specific stress response was observed, followed by the up-regulation of genes encoding peroxisomal enzymes required for fatty acid metabolism. The stress response included up-regulation of genes coding for enzymes to keep thioredoxin and glutathione reduced, as well as enzymes required for the detoxification of reactive oxygen species. Among the genes coding for various isoenzymes involved in these processes, only a specific subset was expressed. Not the general stress transcription factors Msn2 and Msn4, but rather the specific factor Yap1p seemed to be the main regulator of the stress response. We ascribe the initiation of the oxidative stress response to a combination of poor redox flux and fatty acid-induced uncoupling of the respiratory chain during the metabolic reprogramming phase.
- Published
- 2002
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14. Saccharomyces cerevisiae acyl-CoA oxidase follows a novel, non-PTS1, import pathway into peroxisomes that is dependent on Pex5p.
- Author
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Klein AT, van den Berg M, Bottger G, Tabak HF, and Distel B
- Subjects
- Acyl-CoA Oxidase, Amino Acid Sequence, Base Sequence, Binding Sites, DNA Primers, Molecular Sequence Data, Peroxisome-Targeting Signal 1 Receptor, Protein Binding, Protein Transport, Receptors, Cytoplasmic and Nuclear chemistry, Sequence Homology, Amino Acid, Oxidoreductases metabolism, Peroxisomes metabolism, Receptors, Cytoplasmic and Nuclear metabolism, Saccharomyces cerevisiae enzymology, Saccharomyces cerevisiae Proteins
- Abstract
The peroxisomal protein acyl-CoA oxidase (Pox1p) of Saccharomyces cerevisiae lacks either of the two well characterized peroxisomal targeting sequences known as PTS1 and PTS2. Here we demonstrate that peroxisomal import of Pox1p is nevertheless dependent on binding to Pex5p, the PTS1 import receptor. The interaction between Pex5p and Pox1p, however, involves novel contact sites in both proteins. The interaction region in Pex5p is located in a defined area of the amino-terminal part of the protein outside of the tetratricopeptide repeat domain involved in PTS1 recognition; the interaction site in Pox1p is located internally and not at the carboxyl terminus where a PTS1 is normally found. By making use of pex5 mutants that are either specifically disturbed in binding of PTS1 proteins or in binding of Pox1p, we demonstrate the existence of two independent, Pex5p-mediated import pathways into peroxisomes in yeast as follows: a classical PTS1 pathway and a novel, non-PTS1 pathway for Pox1p.
- Published
- 2002
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15. Mutational spectrum in the PEX7 gene and functional analysis of mutant alleles in 78 patients with rhizomelic chondrodysplasia punctata type 1.
- Author
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Motley AM, Brites P, Gerez L, Hogenhout E, Haasjes J, Benne R, Tabak HF, Wanders RJ, and Waterham HR
- Subjects
- Amino Acid Sequence, Animals, COS Cells, Chondrodysplasia Punctata, Rhizomelic classification, Chondrodysplasia Punctata, Rhizomelic enzymology, Chondrodysplasia Punctata, Rhizomelic pathology, Codon genetics, DNA Mutational Analysis, Fibroblasts, Frameshift Mutation genetics, Genes, Recessive genetics, Genes, Reporter genetics, Genetic Complementation Test, Homozygote, Humans, Luciferases genetics, Luciferases metabolism, Molecular Sequence Data, Open Reading Frames genetics, Peroxisomal Targeting Signal 2 Receptor, Phenotype, Protein Folding, Protein Structure, Secondary, Receptors, Cytoplasmic and Nuclear chemistry, Repetitive Sequences, Amino Acid genetics, Sequence Alignment, Structure-Activity Relationship, Alleles, Chondrodysplasia Punctata, Rhizomelic genetics, Mutation genetics, Receptors, Cytoplasmic and Nuclear genetics, Receptors, Cytoplasmic and Nuclear metabolism
- Abstract
Rhizomelic chondrodysplasia punctata (RCDP) is a genetically heterogeneous, autosomal recessive disorder of peroxisomal metabolism that is clinically characterized by symmetrical shortening of the proximal long bones, cataracts, periarticular calcifications, multiple joint contractures, and psychomotor retardation. Most patients with RCDP have mutations in the PEX7 gene encoding peroxin 7, the cytosolic PTS2-receptor protein required for targeting a subset of enzymes to peroxisomes. These enzymes are deficient in cells of patients with RCDP, because of their mislocalization to the cytoplasm. We report the mutational spectrum in the PEX7 gene of 78 patients (including five pairs of sibs) clinically and biochemically diagnosed with RCDP type I. We found 22 different mutations, including 18 novel ones. Furthermore, we show by functional analysis that disease severity correlates with PEX7 allele activity: expression of eight different alleles from patients with severe RCDP failed to restore the targeting defect in RCDP fibroblasts, whereas two alleles found only in patients with mild disease complemented the targeting defect upon overexpression. Surprisingly, one of the mild alleles comprises a duplication of nucleotides 45-52, which is predicted to lead to a frameshift at codon 17 and an absence of functional peroxin 7. The ability of this allele to complement the targeting defect in RCDP cells suggests that frame restoration occurs, resulting in full-length functional peroxin 7, which leads to amelioration of the predicted severe phenotype. This was confirmed in vitro by expression of the eight-nucleotide duplication-containing sequence fused in different reading frames to the coding sequence of firefly luciferase in COS cells.
- Published
- 2002
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16. A role for Vps1p, actin, and the Myo2p motor in peroxisome abundance and inheritance in Saccharomyces cerevisiae.
- Author
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Hoepfner D, van den Berg M, Philippsen P, Tabak HF, and Hettema EH
- Subjects
- Actin Cytoskeleton metabolism, Bacterial Proteins genetics, Base Sequence, Carrier Proteins genetics, Gene Deletion, Green Fluorescent Proteins, Indicators and Reagents metabolism, Luminescent Proteins genetics, Membrane Proteins genetics, Microtubule-Associated Proteins genetics, Microtubules, Molecular Sequence Data, Mutagenesis physiology, Oligonucleotide Probes, Peroxisome-Targeting Signal 1 Receptor, Polymers metabolism, Receptors, Cytoplasmic and Nuclear genetics, Receptors, Cytoplasmic and Nuclear metabolism, Saccharomyces cerevisiae genetics, Vesicular Transport Proteins, Actins metabolism, Carrier Proteins metabolism, Cytoskeletal Proteins, GTP-Binding Proteins, Myosin Heavy Chains metabolism, Myosin Type V metabolism, Peroxisomes metabolism, Saccharomyces cerevisiae metabolism, Saccharomyces cerevisiae Proteins metabolism
- Abstract
In vivo time-lapse microscopy reveals that the number of peroxisomes in Saccharomyces cerevisiae cells is fairly constant and that a subset of the organelles are targeted and segregated to the bud in a highly ordered, vectorial process. The dynamin-like protein Vps1p controls the number of peroxisomes, since in a vps1Delta mutant only one or two giant peroxisomes remain. Analogous to the function of other dynamin-related proteins, Vps1p may be involved in a membrane fission event that is required for the regulation of peroxisome abundance. We found that efficient segregation of peroxisomes from mother to bud is dependent on the actin cytoskeleton, and active movement of peroxisomes along actin filaments is driven by the class V myosin motor protein, Myo2p: (a) peroxisomal dynamics always paralleled the polarity of the actin cytoskeleton, (b) double labeling of peroxisomes and actin cables revealed a close association between both, (c) depolymerization of the actin cytoskeleton abolished all peroxisomal movements, and (d) in cells containing thermosensitive alleles of MYO2, all peroxisome movement immediately stopped at the nonpermissive temperature. In addition, time-lapse videos showing peroxisome movement in wild-type and vps1Delta cells suggest the existence of various levels of control involved in the partitioning of peroxisomes.
- Published
- 2001
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17. Changes in mRNA expression profile underlie phenotypic adaptations in creatine kinase-deficient muscles.
- Author
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de Groof AJ, Smeets B, Groot Koerkamp MJ, Mul AN, Janssen EE, Tabak HF, and Wieringa B
- Subjects
- Animals, Creatine Kinase, Mitochondrial Form, Mice, Mice, Inbred C57BL, Mice, Knockout, Muscle, Skeletal physiology, Phenotype, Adaptation, Physiological genetics, Creatine Kinase genetics, Isoenzymes genetics, Muscle, Skeletal metabolism, RNA, Messenger genetics
- Abstract
We have studied the mechanisms that regulate the remodeling of the glycolytic, mitochondrial and structural network of muscles of creatine kinase M (M-CK)/sarcomeric mitochondrial creatine kinase (ScCKmit) knockout mice by comparison of wild-type and mutant mRNA profiles on cDNA arrays. The magnitudes of changes in mRNA levels were most prominent in M-CK/ScCKmit (CK(-/-)) double mutants but did never exceed those of previously observed changes in protein level for any protein examined. In gastrocnemius of CK(-/-) mice we measured a 2.5-fold increase in mRNA level for mitochondrial encoded cytochrome c oxidase (COX)-III which corresponds to the increase in protein content. The level of the nuclear encoded mRNAs for COX-IV, H(+)-ATP synthase-C, adenine nucleotide translocator-1 and insulin-regulatable glucose transporter-4 showed a 1.5-fold increase, also in agreement with protein data. In contrast, no concomitant up-regulation in mRNA and protein content was detected for the mitochondrial inorganic phosphate-carrier, voltage-dependent anion channel and certain glycolytic enzymes. Our results reveal that regulation of transcript level plays an important role, but it is not the only principle involved in the remodeling of mitochondrial and cytosolic design of CK(-/-) muscles.
- Published
- 2001
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18. Identification of a peroxisomal ATP carrier required for medium-chain fatty acid beta-oxidation and normal peroxisome proliferation in Saccharomyces cerevisiae.
- Author
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van Roermund CW, Drissen R, van Den Berg M, Ijlst L, Hettema EH, Tabak HF, Waterham HR, and Wanders RJ
- Subjects
- Adenosine Triphosphate metabolism, Carrier Proteins chemistry, Carrier Proteins genetics, Cell Fractionation, Fungal Proteins chemistry, Fungal Proteins genetics, Genes, Reporter genetics, Glucose metabolism, Humans, Immunoblotting, Lauric Acids metabolism, Luciferases genetics, Luciferases metabolism, Membrane Proteins chemistry, Membrane Proteins genetics, Membrane Proteins metabolism, Mitochondria chemistry, Mitochondria metabolism, Oleic Acid metabolism, Oxidation-Reduction, Peroxisomes chemistry, Peroxisomes ultrastructure, Recombinant Fusion Proteins genetics, Recombinant Fusion Proteins metabolism, Saccharomyces cerevisiae genetics, Saccharomyces cerevisiae ultrastructure, Carrier Proteins metabolism, Fatty Acids metabolism, Fungal Proteins metabolism, Nucleotide Transport Proteins, Peroxisomes metabolism, Saccharomyces cerevisiae physiology, Saccharomyces cerevisiae Proteins
- Abstract
We have characterized the role of YPR128cp, the orthologue of human PMP34, in fatty acid metabolism and peroxisomal proliferation in Saccharomyces cerevisiae. YPR128cp belongs to the mitochondrial carrier family (MCF) of solute transporters and is localized in the peroxisomal membrane. Disruption of the YPR128c gene results in impaired growth of the yeast with the medium-chain fatty acid (MCFA) laurate as a single carbon source, whereas normal growth was observed with the long-chain fatty acid (LCFA) oleate. MCFA but not LCFA beta-oxidation activity was markedly reduced in intact ypr128cDelta mutant cells compared to intact wild-type cells, but comparable activities were found in the corresponding lysates. These results imply that a transport step specific for MCFA beta-oxidation is impaired in ypr128cDelta cells. Since MCFA beta-oxidation in peroxisomes requires both ATP and CoASH for activation of the MCFAs into their corresponding coenzyme A esters, we studied whether YPR128cp is an ATP carrier. For this purpose we have used firefly luciferase targeted to peroxisomes to measure ATP consumption inside peroxisomes. We show that peroxisomal luciferase activity was strongly reduced in intact ypr128cDelta mutant cells compared to wild-type cells but comparable in lysates of both cell strains. We conclude that YPR128cp most likely mediates the transport of ATP across the peroxisomal membrane.
- Published
- 2001
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19. Peroxisomal membrane proteins are properly targeted to peroxisomes in the absence of COPI- and COPII-mediated vesicular transport.
- Author
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Voorn-Brouwer T, Kragt A, Tabak HF, and Distel B
- Subjects
- Brefeldin A pharmacology, COP-Coated Vesicles drug effects, Cells, Cultured, Coat Protein Complex I metabolism, Endoplasmic Reticulum drug effects, Endoplasmic Reticulum metabolism, Fibroblasts, Humans, Intracellular Membranes drug effects, Intracellular Membranes metabolism, Lipoproteins biosynthesis, Lipoproteins genetics, Lipoproteins metabolism, Membrane Proteins biosynthesis, Membrane Proteins genetics, Microscopy, Fluorescence, Peroxins, Peroxisomal Biogenesis Factor 2, Peroxisomes drug effects, Protein Transport drug effects, ATP-Binding Cassette Transporters, COP-Coated Vesicles metabolism, Coat Protein Complex I antagonists & inhibitors, Fungal Proteins, Membrane Proteins metabolism, Peroxisomes metabolism
- Abstract
The classic model for peroxisome biogenesis states that new peroxisomes arise by the fission of pre-existing ones and that peroxisomal matrix and membrane proteins are recruited directly from the cytosol. Recent studies challenge this model and suggest that some peroxisomal membrane proteins might traffic via the endoplasmic reticulum to peroxisomes. We have studied the trafficking in human fibroblasts of three peroxisomal membrane proteins, Pex2p, Pex3p and Pex16p, all of which have been suggested to transit the endoplasmic reticulum before arriving in peroxisomes. Here, we show that targeting of these peroxisomal membrane proteins is not affected by inhibitors of COPI and COPII that block vesicle transport in the early secretory pathway. Moreover, we have obtained no evidence for the presence of these peroxisomal membrane proteins in compartments other than peroxisomes and demonstrate that COPI and COPII inhibitors do not affect peroxisome morphology or integrity. Together, these data fail to provide any evidence for a role of the endoplasmic reticulum in peroxisome biogenesis.
- Published
- 2001
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20. Recognition of peroxisomal targeting signal type 1 by the import receptor Pex5p.
- Author
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Klein AT, Barnett P, Bottger G, Konings D, Tabak HF, and Distel B
- Subjects
- Amino Acid Sequence, Base Sequence, DNA Primers, Models, Molecular, Molecular Sequence Data, Mutagenesis, Peroxisome-Targeting Signal 1 Receptor, Protein Binding, Protein Conformation, Receptors, Cytoplasmic and Nuclear chemistry, Receptors, Cytoplasmic and Nuclear genetics, Sequence Homology, Amino Acid, Peroxisomes metabolism, Receptors, Cytoplasmic and Nuclear metabolism
- Abstract
We have studied how Pex5p recognizes peroxisomal targeting signal type 1 (PTS1)-containing proteins. A randomly mutagenized pex5 library was screened in a two-hybrid setup for mutations that disrupted the interaction with the PTS1 protein Mdh3p or for suppressor mutations that could restore the interaction with Mdh3p containing a mutation in its PTS1. All mutations localized in the tetratricopeptide repeat (TPR) domain of Pex5p. The Pex5p TPR domain was modeled based on the crystal structure of a related TPR protein. Mapping of the mutations on this structural model revealed that some of the loss-of-interaction mutations consisted of substitutions in alpha-helices of TPRs with bulky amino acids, probably resulting in local misfolding and thereby indirectly preventing binding of PTS1 proteins. The other loss-of-interaction mutations and most suppressor mutations localized in short, exposed, intra-repeat loops of TPR2, TPR3, and TPR6, which are predicted to mediate direct interaction with PTS1 amino acids. Additional site-directed mutants at conserved positions in intra-repeat loops underscored the importance of the loops of TPR2 and TPR3 for PTS1 interaction. Based on the mutational analysis and the structural model, we put forward a model as to how PTS1 proteins are selected by Pex5p.
- Published
- 2001
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21. The peroxisomal membrane protein Pex13p shows a novel mode of SH3 interaction.
- Author
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Barnett P, Bottger G, Klein AT, Tabak HF, and Distel B
- Subjects
- Alanine metabolism, Amino Acid Motifs, Amino Acid Sequence, Binding Sites, Binding, Competitive, Cell Division, Dose-Response Relationship, Drug, Escherichia coli metabolism, Glutathione Transferase metabolism, Ligands, Membrane Proteins genetics, Models, Molecular, Molecular Sequence Data, Mutagenesis, Site-Directed, Peroxisome-Targeting Signal 1 Receptor, Proline metabolism, Protein Binding, Protein Structure, Secondary, Receptors, Cytoplasmic and Nuclear chemistry, Receptors, Cytoplasmic and Nuclear genetics, Receptors, Cytoplasmic and Nuclear metabolism, Recombinant Fusion Proteins metabolism, Sequence Homology, Amino Acid, Suppression, Genetic, Two-Hybrid System Techniques, Membrane Proteins chemistry, Membrane Proteins metabolism, Peroxisomes metabolism, src Homology Domains
- Abstract
Src homology 3 (SH3) domains are small non-catalytic protein modules capable of mediating protein-protein interactions by binding to proline-X-X-proline (P-X-X-P) motifs. Here we demonstrate that the SH3 domain of the integral peroxisomal membrane protein Pex13p is able to bind two proteins, one of which, Pex5p, represents a novel non-P-X-X-P ligand. Using alanine scanning, two-hybrid and in vitro interaction analysis, we show that an alpha-helical element in Pex5p is necessary and sufficient for SH3 interaction. Sup pressor analysis using Pex5p mutants located in this alpha-helical element allowed the identification of a unique site of interaction for Pex5p on the Pex13p-SH3 domain that is distinct from the classical P-X-X-P binding pocket. On the basis of a structural model of the Pex13p-SH3 domain we show that this interaction probably takes place between the RT- and distal loops. Thus, the Pex13p-SH3-Pex5p interaction establishes a novel mode of SH3 interaction.
- Published
- 2000
- Full Text
- View/download PDF
22. Saccharomyces cerevisiae PTS1 receptor Pex5p interacts with the SH3 domain of the peroxisomal membrane protein Pex13p in an unconventional, non-PXXP-related manner.
- Author
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Bottger G, Barnett P, Klein AT, Kragt A, Tabak HF, and Distel B
- Subjects
- Amino Acid Motifs, Amino Acid Sequence, Binding Sites, Carrier Proteins genetics, Carrier Proteins metabolism, Membrane Proteins genetics, Membrane Transport Proteins, Molecular Sequence Data, Mutation, Peroxins, Peroxisome-Targeting Signal 1 Receptor, Protein Transport, Receptors, Cytoplasmic and Nuclear genetics, Saccharomyces cerevisiae genetics, Two-Hybrid System Techniques, Membrane Proteins metabolism, Peroxisomes metabolism, Receptors, Cytoplasmic and Nuclear metabolism, Repressor Proteins, Saccharomyces cerevisiae metabolism, Saccharomyces cerevisiae Proteins, src Homology Domains
- Abstract
A number of peroxisome-associated proteins have been described that are involved in the import of proteins into peroxisomes, among which is the receptor for peroxisomal targeting signal 1 (PTS1) proteins Pex5p, the integral membrane protein Pex13p, which contains an Src homology 3 (SH3) domain, and the peripheral membrane protein Pex14p. In the yeast Saccharomyces cerevisiae, both Pex5p and Pex14p are able to bind Pex13p via its SH3 domain. Pex14p contains the classical SH3 binding motif PXXP, whereas this sequence is absent in Pex5p. Mutation of the conserved tryptophan in the PXXP binding pocket of Pex13-SH3 abolished interaction with Pex14p, but did not affect interaction with Pex5p, suggesting that Pex14p is the classical SH3 domain ligand and that Pex5p binds the SH3 domain in an alternative way. To identify the SH3 binding site in Pex5p, we screened a randomly mutagenized PEX5 library for loss of interaction with Pex13-SH3. Such mutations were all located in a small region in the N-terminal half of Pex5p. One of the altered residues (F208) was part of the sequence W(204)XXQF(208), that is conserved between Pex5 proteins of different species. Site-directed mutagenesis of Trp204 confirmed the essential role of this motif in recognition of the SH3 domain. The Pex5p mutants could only partially restore PTS1-protein import in pex5Delta cells in vivo. In vitro binding studies showed that these Pex5p mutants failed to interact with Pex13-SH3 in the absence of Pex14p, but regained their ability to bind in the presence of Pex14p, suggesting the formation of a heterotrimeric complex consisting of Pex5p, Pex14p, and Pex13-SH3. In vivo, these Pex5p mutants, like wild-type Pex5p, were still found to be associated with peroxisomes. Taken together, this indicates that in the absence of Pex13-SH3 interaction, other protein(s) is able to bind Pex5p at the peroxisome; Pex14p is a likely candidate for this function.
- Published
- 2000
- Full Text
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23. Pex11p plays a primary role in medium-chain fatty acid oxidation, a process that affects peroxisome number and size in Saccharomyces cerevisiae.
- Author
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van Roermund CW, Tabak HF, van Den Berg M, Wanders RJ, and Hettema EH
- Subjects
- Acyl-CoA Oxidase, Coenzyme A Ligases isolation & purification, Coenzyme A Ligases metabolism, Membrane Proteins genetics, Mutation, Oleic Acid metabolism, Oxidation-Reduction, Oxidoreductases genetics, Peroxins, Peroxisomes ultrastructure, Saccharomyces cerevisiae ultrastructure, Fatty Acids metabolism, Membrane Proteins metabolism, Peroxisomes physiology, Repressor Proteins, Saccharomyces cerevisiae metabolism, Saccharomyces cerevisiae Proteins
- Abstract
The Saccharomyces cerevisiae peroxisomal membrane protein Pex11p has previously been implicated in peroxisome proliferation based on morphological observations of PEX11 mutant cells. Pex11p-deficient cells fail to increase peroxisome number in response to growth on fatty acids and instead accumulate a few giant peroxisomes. We report that mutants deficient in genes required for medium-chain fatty acid (MCFA) beta-oxidation display the same phenotype as Pex11p-deficient cells. Upon closer inspection, we found that Pex11p is required for MCFA beta-oxidation. Disruption of the PEX11 gene results in impaired formation of MCFA-CoA esters as measured in intact cells, whereas their formation is normal in cell lysates. The sole S. cerevisiae MCFA-CoA synthetase (Faa2p) remains properly localized to the inner leaflet of the peroxisomal membrane in PEX11 mutant cells. Therefore, the in vivo latency of MCFA activation observed in Pex11p-deficient cells suggests that Pex11p provides Faa2p with substrate. When PEX11 mutant cells are shifted from glucose to oleate-containing medium, we observed an immediate deficiency in beta-oxidation of MCFAs whereas giant peroxisomes and a failure to increase peroxisome abundance only became apparent much later. Our observations suggest that the MCFA oxidation pathway regulates the level of a signaling molecule that modulates the number of peroxisomal structures in a cell.
- Published
- 2000
- Full Text
- View/download PDF
24. Caenorhabditis elegans has a single pathway to target matrix proteins to peroxisomes.
- Author
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Motley AM, Hettema EH, Ketting R, Plasterk R, and Tabak HF
- Subjects
- Alkyl and Aryl Transferases genetics, Animals, Caenorhabditis elegans cytology, Caenorhabditis elegans genetics, Genes, Reporter genetics, Humans, Microscopy, Fluorescence, Peroxisomal Targeting Signal 2 Receptor, Protein Transport, Recombinant Fusion Proteins metabolism, Alkyl and Aryl Transferases metabolism, Caenorhabditis elegans physiology, Peroxisomes metabolism, Protein Sorting Signals genetics, Receptors, Cytoplasmic and Nuclear metabolism
- Abstract
All eukaryotes so far studied, including animals, plants, yeasts and trypanosomes, have two pathways to target proteins to peroxisomes. These two pathways are specific for the two types of peroxisome targeting signal (PTS) present on peroxisomal matrix proteins. Remarkably, the complete genome sequence of Caenorhabditis elegans lacks the genes encoding proteins specific for the PTS2 targeting pathway. Here we show, by expression of green fluorescent protein (GFP) reporters for both pathways, that the PTS2 pathway is indeed absent in C. elegans. Lack of this pathway in man causes severe disease due to mislocalization of PTS2-containing proteins. This raises the question as to how C. elegans has accommodated the absence of the PTS2 pathway. We found by in silico analysis that C. elegans orthologues of PTS2-containing proteins have acquired a PTS1. We propose that switching of targeting signals has allowed the PTS2 pathway to be lost in the phylogenetic lineage leading to C. elegans.
- Published
- 2000
- Full Text
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25. Transport of fatty acids and metabolites across the peroxisomal membrane.
- Author
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Hettema EH and Tabak HF
- Subjects
- ATP Binding Cassette Transporter, Subfamily D, Member 1, ATP-Binding Cassette Transporters genetics, Acyl Coenzyme A metabolism, Biological Transport, Cytoplasm metabolism, Membrane Proteins genetics, Oxidation-Reduction, Saccharomyces cerevisiae, ATP-Binding Cassette Transporters metabolism, Fatty Acids metabolism, Intracellular Membranes metabolism, Peroxisomes metabolism
- Abstract
The peroxisomal membrane forms a permeability barrier for a wide variety of metabolites required for and formed during fatty acid beta-oxidation. To communicate with the cytoplasm and mitochondria, peroxisomes need dedicated proteins to transport such hydrophilic molecules across their membranes. Genetic and biochemical studies in the yeast Saccharomyces cerevisiae have identified enzymes for redox shuttles as well as the first peroxisomal membrane transporter. This peroxisomal ATP-binding cassette transporter (Pat) is highly homologous to the gene mutated in X-linked adrenoleukodystrophy (X-ALD). The yeast Pat is required for import of activated fatty acids into peroxisomes suggesting that this is the primary defect in X-ALD.
- Published
- 2000
- Full Text
- View/download PDF
26. Nuclear receptors arose from pre-existing protein modules during evolution.
- Author
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Barnett P, Tabak HF, and Hettema EH
- Subjects
- Amino Acid Sequence, Animals, Humans, Molecular Sequence Data, Proteins chemistry, Receptors, Cytoplasmic and Nuclear chemistry, Sequence Homology, Amino Acid, Evolution, Molecular, Proteins genetics, Receptors, Cytoplasmic and Nuclear genetics
- Published
- 2000
- Full Text
- View/download PDF
27. In silicio search for genes encoding peroxisomal proteins in Saccharomyces cerevisiae.
- Author
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Kal AJ, Hettema EH, van den Berg M, Koerkamp MG, van Ijlst L, Distel B, and Tabak HF
- Subjects
- Amino Acid Sequence, Molecular Sequence Data, Sequence Alignment, Sequence Analysis, DNA, Genome, Fungal, Peroxisomes genetics, Saccharomyces cerevisiae genetics
- Abstract
The biogenesis of peroxisomes involves the synthesis of new proteins that after, completion of translation, are targeted to the organelle by virtue of peroxisomal targeting signals (PTS). Two types of PTSs have been well characterized for import of matrix proteins (PTS1 and PTS2). Induction of the genes encoding these matrix proteins takes place in oleate-containing medium and is mediated via an oleate response element (ORE) present in the region preceding these genes. The authors have searched the yeast genome for OREs preceding open reading frames (ORFs), and for ORFs that contain either a PTS1 or PTS2. Of the ORFs containing an ORE, as well as either a PTS1 or a PTS2, many were known to encode bona fide peroxisomal matrix proteins. In addition, candidate genes were identified as encoding putative new peroxisomal proteins. For one case, subcellular location studies validated the in silicio prediction. This gene encodes a new peroxisomal thioesterase.
- Published
- 2000
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28. Transactions at the peroxisomal membrane.
- Author
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Distel B, Braakman I, Elgersma Y, and Tabak HF
- Subjects
- Biological Transport, Endoplasmic Reticulum metabolism, Mutation, Proteins metabolism, Saccharomyces cerevisiae genetics, Saccharomyces cerevisiae metabolism, Intracellular Membranes metabolism, Peroxisomes metabolism
- Published
- 2000
- Full Text
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29. Molecular characterization of carnitine-dependent transport of acetyl-CoA from peroxisomes to mitochondria in Saccharomyces cerevisiae and identification of a plasma membrane carnitine transporter, Agp2p.
- Author
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van Roermund CW, Hettema EH, van den Berg M, Tabak HF, and Wanders RJ
- Subjects
- Biological Transport genetics, Carnitine Acyltransferases metabolism, Carnitine O-Acetyltransferase genetics, Carnitine O-Acetyltransferase metabolism, Carrier Proteins metabolism, Cloning, Molecular, Fungal Proteins, Genes, Fungal, Membrane Proteins chemistry, Microscopy, Immunoelectron, Mutation, Oleic Acid metabolism, Saccharomyces cerevisiae metabolism, Acetyl Coenzyme A metabolism, Amino Acid Transport Systems, Carnitine metabolism, Carrier Proteins genetics, Cell Membrane chemistry, Membrane Proteins genetics, Mitochondria metabolism, Peroxisomes metabolism, Saccharomyces cerevisiae genetics, Saccharomyces cerevisiae Proteins, Symporters
- Abstract
In Saccharomyces cerevisiae, beta-oxidation of fatty acids is confined to peroxisomes. The acetyl-CoA produced has to be transported from the peroxisomes via the cytoplasm to the mitochondrial matrix in order to be degraded to CO(2) and H(2)O. Two pathways for the transport of acetyl-CoA to the mitochondria have been proposed. The first involves peroxisomal conversion of acetyl-CoA into glyoxylate cycle intermediates followed by transport of these intermediates to the mitochondria. The second pathway involves peroxisomal conversion of acetyl-CoA into acetylcarnitine, which is subsequently transported to the mitochondria. Using a selective screen, we have isolated several mutants that are specifically affected in the second pathway, the carnitine-dependent acetyl-CoA transport from the peroxisomes to the mitochondria, and assigned these CDAT mutants to three different complementation groups. The corresponding genes were identified using functional complementation of the mutants with a genomic DNA library. In addition to the previously reported carnitine acetyl-CoA transferase (CAT2), we identified the genes for the yeast orthologue of the human mitochondrial carnitine acylcarnitine translocase (YOR100C or CAC) and for a transport protein (AGP2) required for carnitine transport across the plasma membrane.
- Published
- 1999
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30. Peroxisomes: simple in function but complex in maintenance.
- Author
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Tabak HF, Braakman I, and Distel B
- Subjects
- Animals, Biological Transport, Humans, Peroxisomes genetics, Protein Folding, Proteins metabolism, Rats, Peroxisomes metabolism
- Abstract
Peroxisomes compartmentalize part of the anabolic and catabolic pathways and reactions of the cell. Dysfunction of a single peroxisomal enzyme or loss of the whole peroxisomal compartment causes sporadic, but serious, human diseases. Genetic studies in various yeasts have identified PEX genes, which are required for the maintenance of complete peroxisomes. Mutations in PEX genes have proved to be the molecular cause of several human diseases, particularly those involving loss of organelles. Peroxisomes have several properties that distinguish them from other organelles, including the import of folded proteins from the cytosol by an unknown mechanism. By discussing recent highlights from the field of peroxisome research, we aim to share with the general readership our excitement as well as the many mysteries still surrounding peroxisome function and maintenance.
- Published
- 1999
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31. Import of proteins into peroxisomes.
- Author
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Hettema EH, Distel B, and Tabak HF
- Subjects
- Membrane Proteins chemistry, Peroxisomal Targeting Signal 2 Receptor, Peroxisome-Targeting Signal 1 Receptor, Protein Folding, Receptors, Cytoplasmic and Nuclear chemistry, Signal Transduction, Intracellular Membranes chemistry, Microbodies chemistry, Proteins chemistry
- Abstract
Peroxisomes are organelles that confine an important set of enzymes within their single membrane boundaries. In man, a wide variety of genetic disorders is caused by loss of peroxisome function. In the most severe cases, the clinical phenotype indicates that abnormalities begin to appear during embryological development. In less severe cases, the quality of life of adults is affected. Research on yeast model systems has contributed to a better understanding of peroxisome formation and maintenance. This framework of knowledge has made it possible to understand the molecular basis of most of the peroxisome biogenesis disorders. Interestingly, most peroxisome biogenesis disorders are caused by a failure to target peroxisomal proteins to the organellar matrix or membrane, which classifies them as protein targeting diseases. Here we review recent fundamental research on peroxisomal protein targeting and discuss a few burning questions in the field concerning the origin of peroxisomes.
- Published
- 1999
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32. Enlargement of the endoplasmic reticulum membrane in Saccharomyces cerevisiae is not necessarily linked to the unfolded protein response via Ire1p.
- Author
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Stroobants AK, Hettema EH, van den Berg M, and Tabak HF
- Subjects
- Adaptation, Biological, Endoplasmic Reticulum ultrastructure, Galactose metabolism, Gene Deletion, Membrane Proteins biosynthesis, Myo-Inositol-1-Phosphate Synthase biosynthesis, Oleic Acid metabolism, Phosphoproteins biosynthesis, Endoplasmic Reticulum physiology, Fungal Proteins metabolism, Membrane Glycoproteins metabolism, Microbodies physiology, Protein Folding, Protein Serine-Threonine Kinases, Saccharomyces cerevisiae physiology, Saccharomyces cerevisiae Proteins
- Abstract
Conditions that stress the endoplasmic reticulum (ER) in Saccharomyces cerevisiae can elicit a combination of an unfolded protein response (UPR) and an inositol response (IR). This results in increased synthesis of ER protein-folding factors and of enzymes participating in phospholipid biosynthesis. It was suggested that in cells grown on glucose or galactose medium, the UPR and the IR are linked and controlled by the ER stress sensor Ire1p. However, our studies suggest that during growth on oleate the IR is controlled both by an Ire1p-dependent pathway and by an Ire1p-independent pathway.
- Published
- 1999
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33. Dynamics of gene expression revealed by comparison of serial analysis of gene expression transcript profiles from yeast grown on two different carbon sources.
- Author
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Kal AJ, van Zonneveld AJ, Benes V, van den Berg M, Koerkamp MG, Albermann K, Strack N, Ruijter JM, Richter A, Dujon B, Ansorge W, and Tabak HF
- Subjects
- Cytosol metabolism, Fungal Proteins genetics, Fungal Proteins metabolism, Gene Library, Genetic Techniques, Glucose metabolism, Heat-Shock Proteins genetics, Heat-Shock Proteins metabolism, Microbodies genetics, Microbodies metabolism, Mitochondria genetics, Mitochondria metabolism, Models, Statistical, Mutation, Oleic Acid metabolism, Transcription Factors genetics, Transcription Factors metabolism, Transcription, Genetic, Carbon metabolism, Gene Expression Regulation, Fungal, Saccharomyces cerevisiae genetics, Saccharomyces cerevisiae metabolism, Saccharomyces cerevisiae Proteins
- Abstract
We describe a genome-wide characterization of mRNA transcript levels in yeast grown on the fatty acid oleate, determined using Serial Analysis of Gene Expression (SAGE). Comparison of this SAGE library with that reported for glucose grown cells revealed the dramatic adaptive response of yeast to a change in carbon source. A major fraction (>20%) of the 15,000 mRNA molecules in a yeast cell comprised differentially expressed transcripts, which were derived from only 2% of the total number of approximately 6300 yeast genes. Most of the mRNAs that were differentially expressed code for enzymes or for other proteins participating in metabolism (e.g., metabolite transporters). In oleate-grown cells, this was exemplified by the huge increase of mRNAs encoding the peroxisomal beta-oxidation enzymes required for degradation of fatty acids. The data provide evidence for the existence of redox shuttles across organellar membranes that involve peroxisomal, cytoplasmic, and mitochondrial enzymes. We also analyzed the mRNA profile of a mutant strain with deletions of the PIP2 and OAF1 genes, encoding transcription factors required for induction of genes encoding peroxisomal proteins. Induction of genes under the immediate control of these factors was abolished; other genes were up-regulated, indicating an adaptive response to the changed metabolism imposed by the genetic impairment. We describe a statistical method for analysis of data obtained by SAGE.
- Published
- 1999
- Full Text
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34. A tale of tags: report on a HUGO/EU SAGE workshop, 29 January-1 February 1999, Hilversum, The Netherlands.
- Author
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Baas F and Tabak HF
- Subjects
- Animals, Humans, Transcription, Genetic, Gene Expression, Genetic Techniques, RNA, Messenger genetics, RNA, Messenger metabolism
- Published
- 1999
- Full Text
- View/download PDF
35. The cytosolic DnaJ-like protein djp1p is involved specifically in peroxisomal protein import.
- Author
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Hettema EH, Ruigrok CC, Koerkamp MG, van den Berg M, Tabak HF, Distel B, and Braakman I
- Subjects
- Amino Acid Sequence, Base Sequence, Biological Transport, Active, Cloning, Molecular, Cytosol metabolism, DNA Primers genetics, Escherichia coli genetics, Escherichia coli metabolism, Escherichia coli Proteins, Fungal Proteins genetics, Gene Deletion, Gene Expression, Genes, Fungal, Green Fluorescent Proteins, HSP40 Heat-Shock Proteins, HSP70 Heat-Shock Proteins metabolism, Heat-Shock Proteins genetics, Luminescent Proteins genetics, Luminescent Proteins metabolism, Microscopy, Immunoelectron, Molecular Sequence Data, Phenotype, Saccharomyces cerevisiae genetics, Saccharomyces cerevisiae ultrastructure, Sequence Homology, Amino Acid, Fungal Proteins metabolism, Heat-Shock Proteins metabolism, Microbodies metabolism, Saccharomyces cerevisiae metabolism
- Abstract
The Saccharomyces cerevisiae DJP1 gene encodes a cytosolic protein homologous to Escherichia coli DnaJ. DnaJ homologues act in conjunction with molecular chaperones of the Hsp70 protein family in a variety of cellular processes. Cells with a DJP1 gene deletion are viable and exhibit a novel phenotype among cytosolic J-protein mutants in that they have a specific impairment of only one organelle, the peroxisome. The phenotype was also unique among peroxisome assembly mutants: peroxisomal matrix proteins were mislocalized to the cytoplasm to a varying extent, and peroxisomal structures failed to grow to full size and exhibited a broad range of buoyant densities. Import of marker proteins for the endoplasmic reticulum, nucleus, and mitochondria was normal. Furthermore, the metabolic adaptation to a change in carbon source, a complex multistep process, was unaffected in a DJP1 gene deletion mutant. We conclude that Djp1p is specifically required for peroxisomal protein import.
- Published
- 1998
- Full Text
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36. Molecular basis of rhizomelic chondrodysplasia punctata type I: high frequency of the Leu-292 stop mutation in 38 patients.
- Author
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Brites P, Motley A, Hogenhout E, Hettema E, Wijburg F, Heijmans HS, Tabak HF, Distel B, and Wanders RJ
- Subjects
- Gene Frequency, Humans, Peroxisomal Targeting Signal 2 Receptor, Chondrodysplasia Punctata, Rhizomelic genetics, Leucine genetics, Mutation, Receptors, Cytoplasmic and Nuclear genetics
- Published
- 1998
- Full Text
- View/download PDF
37. Peroxisomal beta-oxidation of polyunsaturated fatty acids in Saccharomyces cerevisiae: isocitrate dehydrogenase provides NADPH for reduction of double bonds at even positions.
- Author
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van Roermund CW, Hettema EH, Kal AJ, van den Berg M, Tabak HF, and Wanders RJ
- Subjects
- Amino Acid Sequence, Arachidonic Acid pharmacology, Carbon pharmacology, Cell Division drug effects, Chemical Phenomena, Chemistry, Physical, Cytosol enzymology, Fatty Acid Desaturases analysis, Fatty Acid Desaturases drug effects, Fatty Acids pharmacology, Gene Expression Regulation, Fungal, Genes, Fungal genetics, Genes, Fungal physiology, Hydrogen Bonding, Isocitrate Dehydrogenase metabolism, Isomerism, Linoleic Acid pharmacology, Microbodies genetics, Microbodies ultrastructure, Mitochondria enzymology, NADP metabolism, NADP pharmacology, Oleic Acid pharmacology, Oleic Acids pharmacology, Oxidation-Reduction, Peroxisome-Targeting Signal 1 Receptor, Receptors, Cytoplasmic and Nuclear metabolism, Saccharomyces cerevisiae genetics, Saccharomyces cerevisiae ultrastructure, Sequence Homology, Amino Acid, Substrate Specificity drug effects, Fatty Acids, Unsaturated metabolism, Microbodies enzymology, Oxidoreductases Acting on CH-CH Group Donors, Saccharomyces cerevisiae enzymology
- Abstract
The beta-oxidation of saturated fatty acids in Saccharomyces cerevisiae is confined exclusively to the peroxisomal compartment of the cell. Processing of mono- and polyunsaturated fatty acids with the double bond at an even position requires, in addition to the basic beta-oxidation machinery, the contribution of the NADPH-dependent enzyme 2,4-dienoyl-CoA reductase. Here we show by biochemical cell fractionation studies that this enzyme is a typical constituent of peroxisomes. As a consequence, the beta-oxidation of mono- and polyunsaturated fatty acids with double bonds at even positions requires stoichiometric amounts of intraperoxisomal NADPH. We suggest that NADP-dependent isocitrate dehydrogenase isoenzymes function in an NADP redox shuttle across the peroxisomal membrane to keep intraperoxisomal NADP reduced. This is based on the finding of a third NADP-dependent isocitrate dehydrogenase isoenzyme, Idp3p, next to the already known mitochondrial and cytosolic isoenzymes, which turned out to be present in the peroxisomal matrix. Our proposal is strongly supported by the observation that peroxisomal Idp3p is essential for growth on the unsaturated fatty acids arachidonic, linoleic and petroselinic acid, which require 2, 4-dienoyl-CoA reductase activity. On the other hand, growth on oleate which does not require 2,4-dienoyl-CoA reductase, and NADPH is completely normal in Deltaidp3 cells.
- Published
- 1998
- Full Text
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38. Overexpression of Pex15p, a phosphorylated peroxisomal integral membrane protein required for peroxisome assembly in S.cerevisiae, causes proliferation of the endoplasmic reticulum membrane.
- Author
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Elgersma Y, Kwast L, van den Berg M, Snyder WB, Distel B, Subramani S, and Tabak HF
- Subjects
- Amino Acid Sequence, Cloning, Molecular, Endoplasmic Reticulum ultrastructure, Fungal Proteins biosynthesis, Genes, Fungal, Glycoside Hydrolases biosynthesis, Intracellular Membranes ultrastructure, Membrane Proteins chemistry, Membrane Proteins metabolism, Microbodies ultrastructure, Molecular Sequence Data, Phosphoproteins chemistry, Phosphoproteins metabolism, Phosphorylation, Recombinant Fusion Proteins biosynthesis, Recombinant Fusion Proteins chemistry, Saccharomyces cerevisiae genetics, Saccharomyces cerevisiae ultrastructure, Sequence Alignment, Sequence Homology, Amino Acid, beta-Fructofuranosidase, Endoplasmic Reticulum physiology, Intracellular Membranes physiology, Membrane Proteins biosynthesis, Microbodies physiology, Phosphoproteins biosynthesis, Saccharomyces cerevisiae physiology, Saccharomyces cerevisiae Proteins
- Abstract
We have cloned PEX15 which is required for peroxisome biogenesis in Saccharomyces cerevisiae. pex15Delta cells are characterized by the cytosolic accumulation of peroxisomal matrix proteins containing a PTS1 or PTS2 import signal, whereas peroxisomal membrane proteins are present in peroxisomal remnants. PEX15 encodes a phosphorylated, integral peroxisomal membrane protein (Pex15p). Using multiple in vivo methods to determine the topology, Pex15p was found to be a tail-anchored type II (Ncyt-Clumen) peroxisomal membrane protein with a single transmembrane domain near its carboxy-terminus. Overexpression of Pex15p resulted in impaired peroxisome assembly, and caused profound proliferation of the endoplasmic reticulum (ER) membrane. The lumenal carboxy-terminal tail of Pex15p protrudes into the lumen of these ER membranes, as demonstrated by its O-glycosylation. Accumulation in the ER was also observed at an endogenous expression level when Pex15p was fused to the N-terminus of mature invertase. This resulted in core N-glycosylation of the hybrid protein. The lumenal C-terminal tail of Pex15p is essential for targeting to the peroxisomal membrane. Furthermore, the peroxisomal membrane targeting signal of Pex15p overlaps with an ER targeting signal on this protein. These results indicate that Pex15p may be targeted to peroxisomes via the ER, or to both organelles.
- Published
- 1997
- Full Text
- View/download PDF
39. Transport of activated fatty acids by the peroxisomal ATP-binding-cassette transporter Pxa2 in a semi-intact yeast cell system.
- Author
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Verleur N, Hettema EH, van Roermund CW, Tabak HF, and Wanders RJ
- Subjects
- Adenosine Triphosphate metabolism, Adrenoleukodystrophy etiology, Biological Transport, Cell Membrane Permeability, Digitonin pharmacology, Fungal Proteins metabolism, Humans, Oxidation-Reduction, Protoplasts metabolism, ATP-Binding Cassette Transporters metabolism, Acyl Coenzyme A metabolism, Fatty Acids metabolism, Microbodies metabolism, Saccharomyces cerevisiae metabolism, Saccharomyces cerevisiae Proteins
- Abstract
In the yeast Saccharomyces cerevisiae, fatty acid beta-oxidation is restricted to peroxisomes. Previous studies have shown two possible routes by which fatty acids enter the peroxisome. The first route involves transport of medium-chain fatty acids across the peroxisomal membrane as free fatty acids, followed by activation within the peroxisome by Faa2p, an acyl-CoA synthetase. The second route involves transport of long-chain fatty acids. Long-chain fatty acids enter the peroxisome via a route that involves activation in the extraperoxisomal space, followed by transport across the peroxisomal membrane. It has been suggested that this transport is dependent upon the peroxisomal ATP-binding-cassette transporters Pxa1p and Pxa2p. In this paper we investigated whether Pxa2p is directly responsible for the transport of C18:1-CoA, a long-chain acyl-CoA ester. Using protoplasts in which the plasma membrane has been selectively permeabilised by digitonin, we show that C18:1-CoA, but not C8:0-CoA, enters the peroxisome via Pxa2p, in an ATP-dependent fashion. The results obtained may contribute to the elucidation of the primary defect in the human disease X-linked adrenoleukodystrophy.
- Published
- 1997
- Full Text
- View/download PDF
40. Cytosolic aspartate aminotransferase encoded by the AAT2 gene is targeted to the peroxisomes in oleate-grown Saccharomyces cerevisiae.
- Author
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Verleur N, Elgersma Y, Van Roermund CW, Tabak HF, and Wanders RJ
- Subjects
- Amino Acid Sequence, Aspartate Aminotransferases biosynthesis, Base Sequence, Biological Transport, Cloning, Molecular, Enzyme Induction, Microscopy, Electron, Molecular Sequence Data, Saccharomyces cerevisiae genetics, Saccharomyces cerevisiae growth & development, Subcellular Fractions enzymology, Aspartate Aminotransferases genetics, Cytosol enzymology, Microbodies enzymology, Oleic Acid pharmacology, Saccharomyces cerevisiae enzymology
- Abstract
Fatty acid beta-oxidation in peroxisomes requires the continued uptake of fatty acids or their derivatives into peroxisomes and export of beta-oxidation products plus oxidation of NADH to NAD. In an earlier study we provided evidence for the existence of an NAD(H) redox shuttle in which peroxisomal malate dehydrogenase plays a pivotal role. In analogy to the NAD(H)-redox-shuttle systems in mitochondria we have investigated whether a malate/aspartate shuttle is operative in peroxisomes. The results described in this paper show that peroxisomes of oleate-grown Saccharomyces cerevisiae contain aspartate aminotransferase (AAT) activity. Whereas virtually all cellular AAT activity was peroxisomal in oleate-grown cells, we found that in glucose-grown cells most of the AAT activity resided in the cytosol. We demonstrate that the gene AAT2 codes for the cytosolic and peroxisomal AAT activities. Disruption of the AAT2 gene did not affect growth on oleate. Furthermore beta-oxidation of palmitate was normal. These results indicate that AAT2 is not essential for the peroxisomal NAD(H) redox shuttle.
- Published
- 1997
- Full Text
- View/download PDF
41. A heterodimer of the Zn2Cys6 transcription factors Pip2p and Oaf1p controls induction of genes encoding peroxisomal proteins in Saccharomyces cerevisiae.
- Author
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Rottensteiner H, Kal AJ, Hamilton B, Ruis H, and Tabak HF
- Subjects
- Cloning, Molecular, Dimerization, Fungal Proteins genetics, Genes, Fungal, Oleic Acid metabolism, Protein Binding, Transcription Factors genetics, Fungal Proteins metabolism, Gene Expression Regulation, Fungal, Microbodies metabolism, Saccharomyces cerevisiae genetics, Saccharomyces cerevisiae Proteins, Transcription Factors metabolism
- Abstract
In the yeast Saccharomyces cerevisiae, two transcriptional activators belonging to the Zn2Cys6 protein family, Pip2p and Oaf1p, are involved in fatty-acid-dependent induction of genes encoding peroxisomal proteins. This induction is mediated via an upstream activation sequence called the oleate-response element (ORE). DNA-bandshift experiments with ORE probes and epitope-tagged proteins showed that two binary complexes occurred: in wild-type cells the major complex consisted of a Pip2p x Oaf1p heterodimer, but in cells in which Oaf1p was overexpressed an Oaf1p homodimer was also observed. The genes encoding Oaf1p and Pip2p were controlled in different ways. The OAF1 gene was constitutively expressed, while the PIP2 gene was induced upon growth on oleate, giving rise to positive autoregulatory control. We have shown that the Pip2p x Oaf1p heterodimer is responsible for the strong expression of the genes encoding peroxisomal proteins upon growth on oleate. Pip2p and Oaf1p form an example of a heterodimere of yeast Zn2Cys6 zinc-finger proteins binding to DNA.
- Published
- 1997
- Full Text
- View/download PDF
42. Rhizomelic chondrodysplasia punctata is a peroxisomal protein targeting disease caused by a non-functional PTS2 receptor.
- Author
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Motley AM, Hettema EH, Hogenhout EM, Brites P, ten Asbroek AL, Wijburg FA, Baas F, Heijmans HS, Tabak HF, Wanders RJ, and Distel B
- Subjects
- Amino Acid Sequence, Animals, Base Sequence, Cells, Cultured, Cloning, Molecular, DNA, Complementary genetics, Fibroblasts, Gene Expression, Humans, Mice, Molecular Sequence Data, Mutation, Peroxisomal Targeting Signal 2 Receptor, Polymorphism, Single-Stranded Conformational, Receptors, Cytoplasmic and Nuclear metabolism, Receptors, Cytoplasmic and Nuclear physiology, Recombinant Fusion Proteins, Sequence Homology, Amino Acid, Chondrodysplasia Punctata, Rhizomelic genetics, Receptors, Cytoplasmic and Nuclear genetics
- Abstract
Rhizomelic chondrodysplasia punctata (RCDP) is an autosomal recessive disease characterized clinically by a disproportionately short stature primarily affecting the proximal parts of the extremities, typical dysmorphic facial appearance, congenital contractures and severe growth and mental retardation. Although some patients have single enzyme deficiencies, the majority of RCDP patients (86%) belong to a single complementation group (CG11, also known as complementation group I, Amsterdam nomenclature). Cells from CG11 show a tetrad of biochemical abnormalities: a deficiency of i) dihydroxyacetonephosphate acyltransferase, ii) alkyldihydroxyacetonephosphate synthase, iii) phytanic acid alpha-oxidation and iv) inability to import peroxisomal thiolase. These deficiencies indicate involvement of a component required for correct targeting of these peroxisomal proteins. Deficiencies in peroxisomal targeting are also found in Saccharomyces cerevisiae pex5 and pex7 mutants, which show differential protein import deficiencies corresponding to two peroxisomal targeting sequences (PTS1 and PTS2). These mutants lack their PTS1 and PTS2 receptors, respectively. Like S. cerevisiae pex cells, RCDP cells from CG11 cannot import a PTS2 reporter protein. Here we report the cloning of PEX7 encoding the human PTS2 receptor, based on its similarity to two yeast orthologues. All RCDP patients from CG11 with detectable PEX7 mRNA were found to contain mutations in PEX7. A mutation resulting in C-terminal truncation of PEX7 cosegregates with the disease and expression of PEX7 in RCDP fibroblasts from CG11 rescues the PTS2 protein import deficiency. These findings prove that mutations in PEX7 cause RCDP, CG11.
- Published
- 1997
- Full Text
- View/download PDF
43. Proteins involved in peroxisome biogenesis and functioning.
- Author
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Elgersma Y and Tabak HF
- Subjects
- Biological Transport, Fungal Proteins genetics, Fungal Proteins physiology, Mutation, Yeasts genetics, Yeasts physiology, Microbodies physiology, Proteins physiology
- Published
- 1996
- Full Text
- View/download PDF
44. Analysis of the carboxyl-terminal peroxisomal targeting signal 1 in a homologous context in Saccharomyces cerevisiae.
- Author
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Elgersma Y, Vos A, van den Berg M, van Roermund CW, van der Sluijs P, Distel B, and Tabak HF
- Subjects
- Blotting, Western, Electrophoresis, Polyacrylamide Gel, Epitopes, Malate Dehydrogenase genetics, Malate Dehydrogenase metabolism, Microbodies enzymology, Microscopy, Electron, Peroxisome-Targeting Signal 1 Receptor, Subcellular Fractions enzymology, Protein Sorting Signals metabolism, Receptors, Cytoplasmic and Nuclear metabolism, Saccharomyces cerevisiae metabolism
- Abstract
Most peroxisomal matrix proteins contain a carboxyl-terminal tripeptide that directs them to peroxisomes. Within limits, these amino acids may be varied, without loss of function. The specificity of this peroxisomal targeting signal (PTS1) is remarkable considering its small size and its relaxed consensus sequence. Moreover, several peroxisomal proteins have a PTS1-like signal that does not fit the reported consensus sequence. Because many of these PTS1 variants seem to be functional in a species-dependent or protein context-dependent manner, we investigated the PTS1 requirements in a homologous context, using Saccharomyces cerevisiae and endogenous peroxisomal malate dehydrogenase (MDH3). Peroxisomal import of the MDH3-PTS1 variants was tested qualitatively by the ability to complement the Deltamdh3 mutant and quantitatively by subcellular fractionation. We observed efficient import of MDH3 into peroxisomes with a large variety of PTS1 tripeptides. Many of these variants do not fit the observed PTS1 requirements for heterologously expressed proteins, which suggests that additional domains in the protein may be of decisive importance whether or not a certain PTS1 variant is recognized by the components of the peroxisomal import machinery. Because we show that dimerization of MDH3 precedes import into the organelle, these domains are most likely conformational domains.
- Published
- 1996
- Full Text
- View/download PDF
45. The SH3 domain of the Saccharomyces cerevisiae peroxisomal membrane protein Pex13p functions as a docking site for Pex5p, a mobile receptor for the import PTS1-containing proteins.
- Author
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Elgersma Y, Kwast L, Klein A, Voorn-Brouwer T, van den Berg M, Metzig B, America T, Tabak HF, and Distel B
- Subjects
- 3-Hydroxyacyl CoA Dehydrogenases metabolism, Acetyl-CoA C-Acetyltransferase analysis, Amino Acid Sequence, Base Sequence, Biological Transport, Catalase metabolism, Cloning, Molecular, Cytosol chemistry, Fibroblasts, Genes, Fungal genetics, Humans, Intracellular Membranes chemistry, Ligands, Membrane Proteins analysis, Membrane Proteins genetics, Microbodies chemistry, Molecular Sequence Data, Peroxisome-Targeting Signal 1 Receptor, Point Mutation, Receptors, Cytoplasmic and Nuclear analysis, Receptors, Cytoplasmic and Nuclear genetics, Recombinant Fusion Proteins metabolism, Sequence Deletion, Membrane Proteins metabolism, Microbodies metabolism, Receptors, Cytoplasmic and Nuclear metabolism, Saccharomyces cerevisiae genetics, Saccharomyces cerevisiae Proteins, src Homology Domains genetics
- Abstract
We identified a Saccharomyces cerevisiae peroxisomal membrane protein, Pex13p, that is essential for protein import. A point mutation in the COOH-terminal Src homology 3 (SH3) domain of Pex13p inactivated the protein but did not affect its membrane targeting. A two-hybrid screen with the SH3 domain of Pex13p identified Pex5p, a receptor for proteins with a type I peroxisomal targeting signal (PTS1), as its ligand. Pex13p SH3 interacted specifically with Pex5p in vitro. We determined, furthermore, that Pex5p was mainly present in the cytosol and only a small fraction was associated with peroxisomes. We therefore propose that Pex13p is a component of the peroxisomal protein import machinery onto which the mobile Pex5p receptor docks for the delivery of the selected PTS1 protein.
- Published
- 1996
- Full Text
- View/download PDF
46. A unified nomenclature for peroxisome biogenesis factors.
- Author
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Distel B, Erdmann R, Gould SJ, Blobel G, Crane DI, Cregg JM, Dodt G, Fujiki Y, Goodman JM, Just WW, Kiel JA, Kunau WH, Lazarow PB, Mannaerts GP, Moser HW, Osumi T, Rachubinski RA, Roscher A, Subramani S, Tabak HF, Tsukamoto T, Valle D, van der Klei I, van Veldhoven PP, and Veenhuis M
- Subjects
- Animals, Fungal Proteins, Humans, Microbodies, Proteins, Terminology as Topic
- Published
- 1996
- Full Text
- View/download PDF
47. The ABC transporter proteins Pat1 and Pat2 are required for import of long-chain fatty acids into peroxisomes of Saccharomyces cerevisiae.
- Author
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Hettema EH, van Roermund CW, Distel B, van den Berg M, Vilela C, Rodrigues-Pousada C, Wanders RJ, and Tabak HF
- Subjects
- Base Sequence, Biological Transport, Active, Blotting, Northern, Blotting, Western, Coenzyme A Ligases metabolism, Cytosol metabolism, Electrophoresis, Polyacrylamide Gel, Fungal Proteins metabolism, Microscopy, Immunoelectron, Molecular Sequence Data, Oleic Acid, Oleic Acids metabolism, Saccharomyces cerevisiae genetics, ATP-Binding Cassette Transporters metabolism, Fatty Acids metabolism, Microbodies metabolism, Saccharomyces cerevisiae metabolism
- Abstract
Peroxisomes of Saccharomyces cerevisiae are the exclusive site of fatty acid beta-oxidation. We have found that fatty acids reach the peroxisomal matrix via two independent pathways. The subcellular site of fatty acid activation varies with chain length of the substrate and dictates the pathway of substrate entry into peroxisomes. Medium-chain fatty acids are activated inside peroxisomes hby the acyl-CoA synthetase Faa2p. On the other hand, long-chain fatty acids are imported from the cytosolic pool of activated long-chain fatty acids via Pat1p and Pat2p, peroxisomal membrane proteins belonging to the ATP binding cassette transporter superfamily. Pat1p and Pat2p are the first examples of membrane proteins involved in metabolite transport across the peroxisomal membrane.
- Published
- 1996
48. Pip2p: a transcriptional regulator of peroxisome proliferation in the yeast Saccharomyces cerevisiae.
- Author
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Rottensteiner H, Kal AJ, Filipits M, Binder M, Hamilton B, Tabak HF, and Ruis H
- Subjects
- Amino Acid Sequence, Base Sequence, Cloning, Molecular, DNA, Fungal, Fungal Proteins metabolism, Molecular Sequence Data, Oleic Acid, Oleic Acids biosynthesis, Oleic Acids physiology, Saccharomyces cerevisiae metabolism, Saccharomyces cerevisiae ultrastructure, Sequence Homology, Amino Acid, Sequence Homology, Nucleic Acid, Transcription Factors metabolism, Fungal Proteins genetics, Microbodies physiology, Saccharomyces cerevisiae genetics, Saccharomyces cerevisiae Proteins, Trans-Activators physiology, Transcription Factors genetics
- Abstract
In Saccharomyces cerevisiae, peroxisomes are the exclusive site for the degradation of fatty acids. Upon growth with the fatty acid oleic acid as sole carbon source, not only are the enzymes of beta-oxidation and catalase A induced, but also the peroxisomal compartment as a whole increases in volume and the number of organelles per cell rises. We previously identified a cis-acting DNA sequence [oleate response element (ORE)] involved in induction of genes encoding peroxisomal proteins. The aim of our investigation was to test whether a single mechanism acting via the ORE coordinates the events necessary for the proliferation of an entire organelle. Here we report the cloning and characterization of the oleate-specific transcriptional activator protein Pip2p (pip: peroxisome induction pathway). Pip2p contains a typical Zn(2)-Cys(6) cluster domain and binds to OREs. A pip2 deletion strain is impaired in growth on oleate as sole carbon source and the induction of beta-oxidation enzymes is abolished. Moreover, only a few, small peroxisomes per cell can be detected. These results indicate that fatty acids activate Pip2p, which in turn activates the transcription of genes encoding beta-oxidation components and acts as the crucial activator of peroxisomes.
- Published
- 1996
49. Non-rhizomelic and rhizomelic chondrodysplasia punctata within a single complementation group.
- Author
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Motley AM, Tabak HF, Smeitink JA, Poll-The BT, Barth PG, and Wanders RJ
- Subjects
- Acetyl-CoA C-Acetyltransferase metabolism, Acyltransferases deficiency, Acyltransferases metabolism, Biological Transport genetics, Fibroblasts enzymology, Fibroblasts ultrastructure, Genetic Complementation Test, Humans, Microbodies enzymology, Phenotype, Protein Sorting Signals metabolism, Recombinant Fusion Proteins metabolism, Transferases deficiency, Transferases metabolism, Alkyl and Aryl Transferases, Chondrodysplasia Punctata genetics, Chondrodysplasia Punctata, Rhizomelic genetics, Genetic Heterogeneity
- Abstract
Several patients have been described recently who suffer from a non-rhizomelic type of chondrodysplasia punctata (CDP), but who show all the biochemical abnormalities characteristic of the rhizomelic form of chondrodysplasia punctata (RCDP), a peroxisomal disorder. We have used protease protection experiments and microinjection of reporter-protein-encoding expression plasmids to show that peroxisomal thiolase fails to be imported into peroxisomes in cells from non-rhizomelic CDP patients, as has already been found in cells from classical RCDP patients. Furthermore, complementation analysis after somatic cell fusion indicates that the non-rhizomelic CDP patients are impaired in the same gene as classical RCDP patients. We conclude that defects in a single gene can give rise to both clinical phenotypes.
- Published
- 1996
- Full Text
- View/download PDF
50. The immunohistochemical localization of the non-specific lipid transfer protein (sterol carrier protein-2) in rat small intestine enterocytes.
- Author
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Wouters FS, Markman M, de Graaf P, Hauser H, Tabak HF, Wirtz KW, and Moorman AF
- Subjects
- Amino Acid Sequence, Animals, Carrier Proteins chemistry, Carrier Proteins metabolism, Humans, Immunoblotting, Immunoenzyme Techniques, Intestinal Absorption, Isoelectric Point, Lipid Metabolism, Male, Microbodies chemistry, Microvilli chemistry, Molecular Sequence Data, Molecular Weight, Rats, Rats, Wistar, Carrier Proteins analysis, Intestine, Small chemistry, Plant Proteins
- Abstract
A 13 kDa protein was isolated from rabbit small intestine brush-border membrane vesicles that was postulated to be involved in intestinal phosphatidylcholine (PC) and cholesterol uptake. This protein has cholesterol and PC-transfer activity in vitro (Turnhofer, H. et al. (1991) Biochim. Biophys. Acta 1064, 275-286) and has a molecular mass and isoelectric point similar to that of the non-specific lipid transfer protein (nsL-TP, identical to sterol carrier protein-2). In addition, the first 28 N-terminal amino acid residues of the 13 kDa protein are nearly identical to nsL-TP from different species (Lipka, G. et al. (1995) J. Biol. Chem. 270, 5917-5925). In view of its possible role in intestinal lipid absorption, the localization of nsL-TP in rat small intestine was investigated using immunohistochemistry and immunoblotting. It is shown that nsLTP is predominantly localized in a subapical zone of the enterocyte but not in the brush-border membrane, thereby excluding a role in lipid uptake of this protein at the level of the plasma membrane. nsL-TP co-localized with the peroxisomal marker PMP70, underscoring earlier observations that nsL-TP is a peroxisomal protein. nsL-TP was found to be present along the entire length of the small intestine. The 58 kDa cross-reactive protein that was recently identified as a peroxisomal thiolase was shown to be present only in a small segment approximately halfway down the jejunum. The close apposition of the peroxisomes with the apical membrane and the discrete distribution of the 58 kDa protein may indicate that these organelles play a role in the intracellular processing of absorbed lipids.
- Published
- 1995
- Full Text
- View/download PDF
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