The AAA peroxins Pex1p and Pex6p function as dislocases for the ubiquitinated peroxisomal import receptor Pex5p

2008 ◽  
Vol 36 (1) ◽  
pp. 99-104 ◽  
Author(s):  
Harald W. Platta ◽  
Mykhaylo O. Debelyy ◽  
Fouzi El Magraoui ◽  
Ralf Erdmann
Keyword(s):  
Matrix Protein ◽  
Protein Import ◽  
Matrix Proteins ◽  
Genetic Dissection ◽  
Cellular Functions ◽  
Peroxisomal Membrane ◽  
Aaa Atpase ◽  
Ubiquitin Conjugating Enzyme ◽  
Core Components ◽  
Import Receptor

The discovery of the peroxisomal ATPase Pex1p triggered the beginning of the research on AAA (ATPase associated with various cellular activities) proteins and the genetic dissection of peroxisome biogenesis. Peroxisomes are virtually ubiquitous organelles, which are connected to diverse cellular functions. The highly diverse and adaptive character of peroxisomes is accomplished by modulation of their enzyme content, which is mediated by dynamically operating protein-import machineries. The import of matrix proteins into the peroxisomal lumen has been described as the ATP-consuming step, but the corresponding reaction, as well as the ATPase responsible, had been obscure for nearly 15 years. Recent work using yeast and human fibroblast cells has identified the peroxisomal AAA proteins Pex1p and Pex6p as mechano-enzymes and core components of a complex which dislocates the cycling import receptor Pex5p from the peroxisomal membrane back to the cytosol. This AAA-mediated process is regulated by the ubiquitination status of the receptor. Pex4p [Ubc10p (ubiquitin-conjugating enzyme 10)]-catalysed mono-ubiquitination of Pex5p primes the receptor for recycling, thereby enabling further rounds of matrix protein import, whereas Ubc4p-catalysed polyubiquitination targets Pex5p to proteasomal degradation.

scholarly journals PEX3 functions as a PEX19 docking factor in the import of class I peroxisomal membrane proteins

2004 ◽  
Vol 164 (6) ◽  
pp. 863-875 ◽  
Author(s):  
Yi Fang ◽  
James C. Morrell ◽  
Jacob M. Jones ◽  
Stephen J. Gould
Keyword(s):  
Membrane Proteins ◽  
Matrix Protein ◽  
Protein Import ◽  
Class Ii ◽  
Class I ◽  
Peroxisomal Membrane ◽  
Direct Role ◽  
Conserved Motif ◽  
Peroxisome Membrane ◽  
Import Receptor

PEX19 is a chaperone and import receptor for newly synthesized, class I peroxisomal membrane proteins (PMPs). PEX19 binds these PMPs in the cytoplasm and delivers them to the peroxisome for subsequent insertion into the peroxisome membrane, indicating that there may be a PEX19 docking factor in the peroxisome membrane. Here we show that PEX3 is required for PEX19 to dock at peroxisomes, interacts specifically with the docking domain of PEX19, and is required for recruitment of the PEX19 docking domain to peroxisomes. PEX3 is also sufficient to dock PEX19 at heterologous organelles and binds PEX19 via a conserved motif that is essential for this docking activity and for PEX3 function in general. Not surprisingly, transient inhibition of PEX3 abrogates class I PMP import but has no effect on class II PMP import or peroxisomal matrix protein import. Taken together, these results suggest that PEX3 plays a selective, essential, and direct role in PMP import as a docking factor for PEX19.

scholarly journals Ubiquitination of the peroxisomal import receptor Pex5p

2004 ◽  
Vol 384 (1) ◽  
pp. 37-45 ◽  
Author(s):  
Harald W. PLATTA ◽  
Wolfgang GIRZALSKY ◽  
Ralf ERDMANN
Keyword(s):  
Protein Import ◽  
Ring Finger ◽  
Proteasomal Degradation ◽  
Peroxisomal Membrane ◽  
Peroxisomal Biogenesis ◽  
Peroxisomal Targeting ◽  
Ubiquitin Conjugating Enzyme ◽  
Targeting Signal ◽  
Import Receptor ◽  
In Cells

Proteins harbouring a peroxisomal targeting signal of type 1 (PTS1) are recognized by the import receptor Pex5p in the cytosol which directs them to a docking and translocation complex at the peroxisomal membrane. We demonstrate the ubiquitination of Pex5p in cells lacking components of the peroxisomal AAA (ATPases associated with various cellular activities) or Pex4p–Pex22p complexes of the peroxisomal protein import machinery and in cells affected in proteasomal degradation. In cells lacking components of the Pex4p–Pex22p complex, mono-ubiquitinated Pex5p represents the major modification, while in cells lacking components of the AAA complex polyubiquitinated forms are most prominent. Ubiquitination of Pex5p is shown to take place exclusively at the peroxisomal membrane after the docking step, and requires the presence of the RING-finger peroxin Pex10p. Mono- and poly-ubiquitination are demonstrated to depend on the ubiquitin-conjugating enzyme Ubc4p, suggesting that the ubiquitinated forms of Pex5p are targeted for proteasomal degradation. Accumulation of ubiquitinated Pex5p in proteasomal mutants demonstrates that the ubiquitination of Pex5p also takes place in strains which are not affected in peroxisomal biogenesis, indicating that the ubiquitination of Pex5p represents a genuine stage in the Pex5p receptor cycle.

scholarly journals Peroxisomes exhibit compromised structure and matrix protein content in SARS-CoV-2-infected cells

2021 ◽  
pp. mbc.E21-02-0074
Author(s):  
Barbara Knoblach ◽  
Ray Ishida ◽  
Tom C. Hobman ◽  
Richard A. Rachubinski
Keyword(s):  
Protein Content ◽  
Matrix Protein ◽  
Protein Import ◽  
Structural Integrity ◽  
Matrix Proteins ◽  
Peroxisome Biogenesis ◽  
Peroxisomal Membrane ◽  
Antiviral Signaling ◽  
Infected Cells ◽  
Novel Coronavirus

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a novel coronavirus that has triggered global health and economic crises. Here we report the effects of SARS-CoV-2 infection on peroxisomes of human cell lines, Huh-7 and SK-N-SH. Peroxisomes undergo dramatic changes in morphology in SARS-CoV-2-infected cells. Rearrangement of peroxisomal membranes is followed by redistribution of peroxisomal matrix proteins to the cytosol, resulting in a dramatic decrease in the numbers of mature peroxisomes. The SARS-CoV-2 ORF14 protein was shown to interact physically with human PEX14, a peroxisomal membrane protein required for matrix protein import and peroxisome biogenesis. Given the important roles of peroxisomes in innate immunity, SARS-CoV-2 may directly target peroxisomes, resulting in loss of peroxisome structural integrity, matrix protein content and ability to function in antiviral signaling. [Media: see text]

scholarly journals Yeast pex1 cells contain peroxisomal ghosts that import matrix proteins upon reintroduction of Pex1

2015 ◽  
Vol 211 (5) ◽  
pp. 955-962 ◽  
Author(s):  
Kèvin Knoops ◽  
Rinse de Boer ◽  
Anita Kram ◽  
Ida J. van der Klei
Keyword(s):  
Cross Sections ◽  
Matrix Protein ◽  
Protein Import ◽  
Electron Tomography ◽  
High Resolution Electron Microscopy ◽  
Matrix Proteins ◽  
Peroxisomal Membrane ◽  
Resolution Electron ◽  
Microscopy Techniques ◽  
Aaa Atpases

Pex1 and Pex6 are two AAA-ATPases that play a crucial role in peroxisome biogenesis. We have characterized the ultrastructure of the Saccharomyces cerevisiae peroxisome-deficient mutants pex1 and pex6 by various high-resolution electron microscopy techniques. We observed that the cells contained peroxisomal membrane remnants, which in ultrathin cross sections generally appeared as double membrane rings. Electron tomography revealed that these structures consisted of one continuous membrane, representing an empty, flattened vesicle, which folds into a cup shape. Immunocytochemistry revealed that these structures lack peroxisomal matrix proteins but are the sole sites of the major peroxisomal membrane proteins Pex2, Pex10, Pex11, Pex13, and Pex14. Upon reintroduction of Pex1 in Pex1-deficient cells, these peroxisomal membrane remnants (ghosts) rapidly incorporated peroxisomal matrix proteins and developed into peroxisomes. Our data support earlier views that Pex1 and Pex6 play a role in peroxisomal matrix protein import.

scholarly journals The Peroxisome Biogenesis Factors Pex4p, Pex22p, Pex1p, and Pex6p Act in the Terminal Steps of Peroxisomal Matrix Protein Import

2000 ◽  
Vol 20 (20) ◽  
pp. 7516-7526 ◽  
Author(s):  
Cynthia S. Collins ◽  
Jennifer E. Kalish ◽  
James C. Morrell ◽  
J. Michael McCaffery ◽  
Stephen J. Gould
Keyword(s):  
Matrix Protein ◽  
Protein Import ◽  
Protein Translocation ◽  
Matrix Proteins ◽  
Peroxisome Biogenesis ◽  
Peroxisomal Membrane ◽  
The Matrix ◽  
Peroxisomal Targeting ◽  
Targeting Signal ◽  
Mutant Cells

ABSTRACT Peroxisomes are independent organelles found in virtually all eukaryotic cells. Genetic studies have identified more than 20PEX genes that are required for peroxisome biogenesis. The role of most PEX gene products, peroxins, remains to be determined, but a variety of studies have established that Pex5p binds the type 1 peroxisomal targeting signal and is the import receptor for most newly synthesized peroxisomal matrix proteins. The steady-state abundance of Pex5p is unaffected in mostpex mutants of the yeast Pichia pastorisbut is severely reduced in pex4 andpex22 mutants and moderately reduced in pex1and pex6 mutants. We used these subphenotypes to determine the epistatic relationships among several groups ofpex mutants. Our results demonstrate that Pex4p acts after the peroxisome membrane synthesis factor Pex3p, the Pex5p docking factors Pex13p and Pex14p, the matrix protein import factors Pex8p, Pex10p, and Pex12p, and two other peroxins, Pex2p and Pex17p. Pex22p and the interacting AAA ATPases Pex1p and Pex6p were also found to act after Pex10p. Furthermore, Pex1p and Pex6p were found to act upstream of Pex4p and Pex22p. These results suggest that Pex1p, Pex4p, Pex6p, and Pex22p act late in peroxisomal matrix protein import, after matrix protein translocation. This hypothesis is supported by the phenotypes of the corresponding mutant strains. As has been shown previously for P. pastoris pex1,pex6, and pex22 mutant cells, we show here thatpex4Δ mutant cells contain peroxisomal membrane protein-containing peroxisomes that import residual amounts of peroxisomal matrix proteins.

scholarly journals Peroxisome Synthesis in the Absence of Preexisting Peroxisomes

1999 ◽  
Vol 144 (2) ◽  
pp. 255-266 ◽  
Author(s):  
Sarah T. South ◽  
Stephen J. Gould
Keyword(s):  
Membrane Proteins ◽  
Membrane Protein ◽  
Matrix Protein ◽  
Protein Import ◽  
Zellweger Syndrome ◽  
Matrix Proteins ◽  
Peroxisomal Membrane ◽  
Peroxisomal Membrane Protein ◽  
The Matrix ◽  
Inactivating Mutations

Zellweger syndrome and related diseases are caused by defective import of peroxisomal matrix proteins. In all previously reported Zellweger syndrome cell lines the defect could be assigned to the matrix protein import pathway since peroxisome membranes were present, and import of integral peroxisomal membrane proteins was normal. However, we report here a Zellweger syndrome patient (PBD061) with an unusual cellular phenotype, an inability to import peroxisomal membrane proteins. We also identified human PEX16, a novel integral peroxisomal membrane protein, and found that PBD061 had inactivating mutations in the PEX16 gene. Previous studies have suggested that peroxisomes arise from preexisting peroxisomes but we find that expression of PEX16 restores the formation of new peroxisomes in PBD061 cells. Peroxisome synthesis and peroxisomal membrane protein import could be detected within 2–3 h of PEX16 injection and was followed by matrix protein import. These results demonstrate that peroxisomes do not necessarily arise from division of preexisting peroxisomes. We propose that peroxisomes may form by either of two pathways: one that involves PEX11-mediated division of preexisting peroxisomes, and another that involves PEX16-mediated formation of peroxisomes in the absence of preexisting peroxisomes.

scholarly journals Pex22p of Pichia pastoris, Essential for Peroxisomal Matrix Protein Import, Anchors the Ubiquitin-Conjugating Enzyme, Pex4p, on the Peroxisomal Membrane

1999 ◽  
Vol 146 (1) ◽  
pp. 99-112 ◽  
Author(s):  
Antonius Koller ◽  
William B. Snyder ◽  
Klaas Nico Faber ◽  
Thibaut J. Wenzel ◽  
Linda Rangell ◽  
...  
Keyword(s):  
Pichia Pastoris ◽  
Membrane Protein ◽  
Matrix Protein ◽  
Protein Import ◽  
Hypothetical Protein ◽  
Peroxisomal Membrane ◽  
Peroxisomal Membrane Protein ◽  
Ubiquitin Conjugating Enzyme ◽  
Conjugating Enzyme

We isolated a Pichia pastoris mutant that was unable to grow on the peroxisome-requiring media, methanol and oleate. Cloning the gene by complementation revealed that the encoded protein, Pex22p, is a new peroxin. A Δpex22 strain does not grow on methanol or oleate and is unable to import peroxisomal matrix proteins. However, this strain targets peroxisomal membrane proteins to membranes, most likely peroxisomal remnants, detectable by fluorescence and electron microscopy. Pex22p, composed of 187 amino acids, is an integral peroxisomal membrane protein with its NH2 terminus in the matrix and its COOH terminus in the cytosol. It contains a 25–amino acid peroxisome membrane-targeting signal at its NH2 terminus. Pex22p interacts with the ubiquitin-conjugating enzyme Pex4p, a peripheral peroxisomal membrane protein, in vivo, and in a yeast two-hybrid experiment. Pex22p is required for the peroxisomal localization of Pex4p and in strains lacking Pex22p, the Pex4p is cytosolic and unstable. Therefore, Pex22p anchors Pex4p at the peroxisomal membrane. Strains that do not express Pex4p or Pex22p have similar phenotypes and lack Pex5p, suggesting that Pex4p and Pex22p act at the same step in peroxisome biogenesis. The Saccharomyces cerevisiae hypothetical protein, Yaf5p, is the functional homologue of P. pastoris Pex22p.

Metabolic control of peroxisome abundance

1999 ◽  
Vol 112 (10) ◽  
pp. 1579-1590 ◽  
Author(s):  
C.C. Chang ◽  
S. South ◽  
D. Warren ◽  
J. Jones ◽  
A.B. Moser ◽  
...  
Keyword(s):  
Metabolic Control ◽  
Matrix Protein ◽  
Protein Import ◽  
Zellweger Syndrome ◽  
Mutant Gene ◽  
Peroxisomal Membrane ◽  
Peroxisomal Targeting ◽  
Targeting Signal ◽  
Complementation Groups ◽  
Peroxisomal Targeting Signal

Zellweger syndrome and related disorders represent a group of lethal, genetically heterogeneous diseases. These peroxisome biogenesis disorders (PBDs) are characterized by defective peroxisomal matrix protein import and comprise at least 10 complementation groups. The genes defective in seven of these groups and more than 90% of PBD patients are now known. Here we examine the distribution of peroxisomal membrane proteins in fibroblasts from PBD patients representing the seven complementation groups for which the mutant gene is known. Peroxisomes were detected in all PBD cells, indicating that the ability to form a minimal peroxisomal structure is not blocked in these mutants. We also observed that peroxisome abundance was reduced fivefold in PBD cells that are defective in the PEX1, PEX5, PEX12, PEX6, PEX10, and PEX2 genes. These cell lines all display a defect in the import of proteins with the type-1 peroxisomal targeting signal (PTS1). In contrast, peroxisome abundance was unaffected in cells that are mutated in PEX7 and are defective only in the import of proteins with the type-2 peroxisomal targeting signal. Interestingly, a fivefold reduction in peroxisome abundance was also observed for cells lacking either of two PTS1-targeted peroxisomal beta-oxidation enzymes, acyl-CoA oxidase and 2-enoyl-CoA hydratase/D-3-hydroxyacyl-CoA dehydrogenase. These results indicate that reduced peroxisome abundance in PBD cells may be caused by their inability to import these PTS1-containing enzymes. Furthermore, the fact that peroxisome abundance is influenced by peroxisomal 105-oxidation activities suggests that there may be metabolic control of peroxisome abundance.

scholarly journals Autophagy Inhibitors Do Not Restore Peroxisomal Functions in Cells With the Most Common Peroxisome Biogenesis Defect

2021 ◽  
Vol 9 ◽  
Author(s):  
Femke C. C. Klouwer ◽  
Kim D. Falkenberg ◽  
Rob Ofman ◽  
Janet Koster ◽  
Démi van Gent ◽  
...  
Keyword(s):  
Matrix Protein ◽  
Protein Import ◽  
Therapeutic Option ◽  
Cell Types ◽  
Matrix Proteins ◽  
Peroxisome Biogenesis ◽  
Metabolic Functions ◽  
Minimal Improvement ◽  
Different Cell Types ◽  
Autophagy Inhibition

Peroxisome biogenesis disorders within the Zellweger spectrum (PBD-ZSDs) are most frequently associated with the c.2528G>A (p.G843D) mutation in the PEX1 gene (PEX1-G843D), which results in impaired import of peroxisomal matrix proteins and, consequently, defective peroxisomal functions. A recent study suggested that treatment with autophagy inhibitors, in particular hydroxychloroquine, would be a potential therapeutic option for PBD-ZSD patients carrying the PEX1-G843D mutation. Here, we studied whether autophagy inhibition by chloroquine, hydroxychloroquine and 3-methyladenine indeed can improve peroxisomal functions in four different cell types with the PEX1-G843D mutation, including primary patient cells. Furthermore, we studied whether autophagy inhibition may be the mechanism underlying the previously reported improvement of peroxisomal functions by L-arginine in PEX1-G843D cells. In contrast to L-arginine, we observed no improvement but a worsening of peroxisomal metabolic functions and peroxisomal matrix protein import by the autophagy inhibitors, while genetic knock-down of ATG5 and NBR1 in primary patient cells resulted in only a minimal improvement. Our results do not support the use of autophagy inhibitors as potential treatment for PBD-ZSD patients, whereas L-arginine remains a therapeutically promising compound.

Sign in / Sign up

Export Citation Format

Share Document