scholarly journals Distinct requirements for intra-ER sorting and budding of peroxisomal membrane proteins from the ER

2016 ◽  
Vol 212 (3) ◽  
pp. 335-348 ◽  
Author(s):  
Gaurav Agrawal ◽  
Scott N. Fassas ◽  
Zhi-Jie Xia ◽  
Suresh Subramani
Keyword(s):  
Endoplasmic Reticulum ◽  
Membrane Proteins ◽  
Ternary Complex ◽  
De Novo ◽  
Ring Domain ◽  
Peroxisome Biogenesis ◽  
Peroxisomal Membrane ◽  
Unique Role ◽  
Sorting Process

During de novo peroxisome biogenesis, importomer complex proteins sort via two preperoxisomal vesicles (ppVs). However, the sorting mechanisms segregating peroxisomal membrane proteins to the preperoxisomal endoplasmic reticulum (pER) and into ppVs are unknown. We report novel roles for Pex3 and Pex19 in intra–endoplasmic reticulum (ER) sorting and budding of the RING-domain peroxins (Pex2, Pex10, and Pex12). Pex19 bridged the interaction at the ER between Pex3 and RING-domain proteins, resulting in a ternary complex that was critical for the intra-ER sorting and subsequent budding of the RING-domain peroxins. Although the docking subcomplex proteins (Pex13, Pex14, and Pex17) also required Pex19 for budding from the ER, they sorted to the pER independently of Pex3 and Pex19 and were spatially segregated from the RING-domain proteins. We also discovered a unique role for Pex3 in sorting Pex10 and Pex12, but with the docking subcomplex. Our study describes an intra-ER sorting process that regulates segregation, packaging, and budding of peroxisomal importomer subcomplexes, thereby preventing their premature assembly at the ER.

Peroxisomal membrane proteins are properly targeted to peroxisomes in the absence of COPI- and COPII-mediated vesicular transport

2001 ◽  
Vol 114 (11) ◽  
pp. 2199-2204 ◽  
Author(s):  
Tineke Voorn-Brouwer ◽  
Astrid Kragt ◽  
Henk F. Tabak ◽  
Ben Distel
Keyword(s):  
Endoplasmic Reticulum ◽  
Membrane Proteins ◽  
Secretory Pathway ◽  
Human Fibroblasts ◽  
Vesicle Transport ◽  
Peroxisome Biogenesis ◽  
Peroxisomal Membrane ◽  
Classic Model ◽  
Early Secretory Pathway

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.

scholarly journals Mutants of the Yeast Yarrowia lipolyticaDefective in Protein Exit from the Endoplasmic Reticulum Are Also Defective in Peroxisome Biogenesis

1998 ◽  
Vol 18 (5) ◽  
pp. 2789-2803 ◽  
Author(s):  
Vladimir I. Titorenko ◽  
Richard A. Rachubinski
Keyword(s):  
Endoplasmic Reticulum ◽  
Oleic Acid ◽  
Membrane Proteins ◽  
Matrix Proteins ◽  
Secretory Proteins ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Peroxisome Biogenesis ◽  
Peroxisomal Membrane ◽  
Mutant Cells

ABSTRACT Mutations in the SEC238 and SRP54 genes of the yeast Yarrowia lipolytica not only cause temperature-sensitive defects in the exit of the precursor form of alkaline extracellular protease and of other secretory proteins from the endoplasmic reticulum and in protein secretion but also lead to temperature-sensitive growth in oleic acid-containing medium, the metabolism of which requires the assembly of functionally intact peroxisomes. The sec238A andsrp54KO mutations at the restrictive temperature significantly reduce the size and number of peroxisomes, affect the import of peroxisomal matrix and membrane proteins into the organelle, and significantly delay, but do not prevent, the exit of two peroxisomal membrane proteins, Pex2p and Pex16p, from the endoplasmic reticulum en route to the peroxisomal membrane. Mutations in the PEX1 and PEX6 genes, which encode members of the AAA family of N-ethylmaleimide-sensitive fusion protein-like ATPases, not only affect the exit of precursor forms of secretory proteins from the endoplasmic reticulum but also prevent the exit of the peroxisomal membrane proteins Pex2p and Pex16p from the endoplasmic reticulum and cause the accumulation of an extensive network of endoplasmic reticulum membranes. None of the peroxisomal matrix proteins tested associated with the endoplasmic reticulum in sec238A,srp54KO, pex1-1, and pex6KO mutant cells. Our data provide evidence that the endoplasmic reticulum is required for peroxisome biogenesis and suggest that inY. lipolytica, the trafficking of some membrane proteins, but not matrix proteins, to the peroxisome occurs via the endoplasmic reticulum, results in their glycosylation within the lumen of the endoplasmic reticulum, does not involve transport through the Golgi, and requires the products encoded by the SEC238, SRP54,PEX1, and PEX6 genes.

scholarly journals Endoplasmic Reticulum-Associated Secretory Proteins Sec20p, Sec39p, and Dsl1p Are Involved in Peroxisome Biogenesis

2009 ◽  
Vol 8 (6) ◽  
pp. 830-843 ◽  
Author(s):  
Ryan J. Perry ◽  
Fred D. Mast ◽  
Richard A. Rachubinski
Keyword(s):  
Endoplasmic Reticulum ◽  
Secretory Pathway ◽  
Matrix Protein ◽  
Fluorescent Protein ◽  
De Novo ◽  
Secretory Proteins ◽  
Peroxisome Biogenesis ◽  
Gfp Expression ◽  
Peroxisomal Membrane ◽  
Green Fluorescent

ABSTRACT Two pathways have been identified for peroxisome formation: (i) growth and division and (ii) de novo synthesis. Recent experiments determined that peroxisomes originate at the endoplasmic reticulum (ER). Although many proteins have been implicated in the peroxisome biogenic program, no proteins in the eukaryotic secretory pathway have been identified as having roles in peroxisome formation. Using the yeast Saccharomyces cerevisiae regulatable Tet promoter Hughes clone collection, we found that repression of the ER-associated secretory proteins Sec20p and Sec39p resulted in mislocalization of the peroxisomal matrix protein chimera Pot1p-green fluorescent protein (GFP) to the cytosol. Likewise, the peroxisomal membrane protein chimera Pex3p-GFP localized to tubular-vesicular structures in cells suppressed for Sec20p, Sec39p, and Dsl1p, which form a complex at the ER. Loss of Sec39p attenuated formation of Pex3p-derived peroxisomal structures following galactose induction of Pex3p-GFP expression from the GAL1 promoter. Expression of Sec20p, Sec39p, and Dsl1p was moderately increased in yeast grown under conditions that proliferate peroxisomes, and Sec20p, Sec39p, and Dsl1p were found to cofractionate with peroxisomes and colocalize with Pex3p-monomeric red fluorescent protein under these conditions. Our results show that SEC20, SEC39, and DSL1 are essential secretory genes involved in the early stages of peroxisome assembly, and this work is the first to identify and characterize an ER-associated secretory machinery involved in peroxisome biogenesis.

scholarly journals The Peroxin Pex19p Interacts with Multiple, Integral Membrane Proteins at the Peroxisomal Membrane

2000 ◽  
Vol 149 (6) ◽  
pp. 1171-1178 ◽  
Author(s):  
William B. Snyder ◽  
Antonius Koller ◽  
A. Jobu Choy ◽  
Suresh Subramani
Keyword(s):  
Membrane Proteins ◽  
Multiple Integral ◽  
Integral Membrane Proteins ◽  
Peroxisome Biogenesis ◽  
Yeast Two Hybrid ◽  
Peroxisomal Membrane ◽  
Site Of Action ◽  
Peroxisome Membrane ◽  
Two Hybrid ◽  
New Protein

Pex19p is a protein required for the early stages of peroxisome biogenesis, but its precise function and site of action are unknown. We tested the interaction between Pex19p and all known Pichia pastoris Pex proteins by the yeast two-hybrid assay. Pex19p interacted with six of seven known integral peroxisomal membrane proteins (iPMPs), and these interactions were confirmed by coimmunoprecipitation. The interactions were not reduced upon inhibition of new protein synthesis, suggesting that they occur with preexisting, and not newly synthesized, pools of iPMPs. By mapping the domains in six iPMPs that interact with Pex19p and the iPMP sequences responsible for targeting to the peroxisome membrane (mPTSs), we found the majority of these sites do not overlap. Coimmunoprecipitation of Pex19p from fractions that contain peroxisomes or cytosol revealed that the interactions between predominantly cytosolic Pex19p and the iPMPs occur in the organelle pellet that contains peroxisomes. These data, taken together, suggest that Pex19p may have a chaperone-like role at the peroxisome membrane and that it is not the receptor for targeting of iPMPs to the peroxisome.

scholarly journals Four distinct secretory pathways serve protein secretion, cell surface growth, and peroxisome biogenesis in the yeast Yarrowia lipolytica.

1997 ◽  
Vol 17 (9) ◽  
pp. 5210-5226 ◽  
Author(s):  
V I Titorenko ◽  
D M Ogrydziak ◽  
R A Rachubinski
Keyword(s):  
Endoplasmic Reticulum ◽  
Cell Wall ◽  
Plasma Membrane ◽  
Protein Secretion ◽  
Peroxisome Biogenesis ◽  
Peroxisomal Membrane ◽  
Mycelial Form ◽  
Secretory Pathways ◽  
Associated Proteins ◽  
Yeast Yarrowia Lipolytica

We have identified and characterized mutants of the yeast Yarrowia lipolytica that are deficient in protein secretion, in the ability to undergo dimorphic transition from the yeast to the mycelial form, and in peroxisome biogenesis. Mutations in the SEC238, SRP54, PEX1, PEX2, PEX6, and PEX9 genes affect protein secretion, prevent the exit of the precursor form of alkaline extracellular protease from the endoplasmic reticulum, and compromise peroxisome biogenesis. The mutants sec238A, srp54KO, pex2KO, pex6KO, and pex9KO are also deficient in the dimorphic transition from the yeast to the mycelial form and are affected in the export of only plasma membrane and cell wall-associated proteins specific for the mycelial form. Mutations in the SEC238, SRP54, PEX1, and PEX6 genes prevent or significantly delay the exit of two peroxisomal membrane proteins, Pex2p and Pex16p, from the endoplasmic reticulum en route to the peroxisomal membrane. Mutations in the PEX5, PEX16, and PEX17 genes, which have previously been shown to be essential for peroxisome biogenesis, affect the export of plasma membrane and cell wall-associated proteins specific for the mycelial form but do not impair exit from the endoplasmic reticulum of either Pex2p and Pex16p or of proteins destined for secretion. Biochemical analyses of these mutants provide evidence for the existence of four distinct secretory pathways that serve to deliver proteins for secretion, plasma membrane and cell wall synthesis during yeast and mycelial modes of growth, and peroxisome biogenesis. At least two of these secretory pathways, which are involved in the export of proteins to the external medium and in the delivery of proteins for assembly of the peroxisomal membrane, diverge at the level of the endoplasmic reticulum.

scholarly journals Inhibitors of Copi and Copii Do Not Block PEX3-Mediated Peroxisome Synthesis

2000 ◽  
Vol 149 (7) ◽  
pp. 1345-1360 ◽  
Author(s):  
Sarah T. South ◽  
Katherine A. Sacksteder ◽  
Xiaoling Li ◽  
Yifei Liu ◽  
Stephen J. Gould
Keyword(s):  
Membrane Proteins ◽  
Zellweger Syndrome ◽  
Brefeldin A ◽  
Membrane Traffic ◽  
Dominant Negative ◽  
Peroxisome Biogenesis ◽  
Membrane Synthesis ◽  
Peroxisomal Membrane ◽  
Peroxisome Membrane ◽  
Inactivating Mutations

In humans, defects in peroxisome biogenesis are the cause of lethal diseases typified by Zellweger syndrome. Here, we show that inactivating mutations in human PEX3 cause Zellweger syndrome, abrogate peroxisome membrane synthesis, and result in reduced abundance of peroxisomal membrane proteins (PMPs) and/or mislocalization of PMPs to the mitochondria. Previous studies have suggested that PEX3 may traffic through the ER en route to the peroxisome, that the COPI inhibitor, brefeldin A, leads to accumulation of PEX3 in the ER, and that PEX3 overexpression alters the morphology of the ER. However, we were unable to detect PEX3 in the ER at early times after expression. Furthermore, we find that inhibition of COPI function by brefeldin A has no effect on trafficking of PEX3 to peroxisomes and does not inhibit PEX3-mediated peroxisome biogenesis. We also find that inhibition of COPII-dependent membrane traffic by a dominant negative SAR1 mutant fails to block PEX3 transport to peroxisomes and PEX3-mediated peroxisome synthesis. Based on these results, we propose that PEX3 targeting to peroxisomes and PEX3-mediated peroxisome membrane synthesis may occur independently of COPI- and COPII-dependent membrane traffic.

scholarly journals Reevaluation of the role of Pex1 and dynamin-related proteins in peroxisome membrane biogenesis

2015 ◽  
Vol 211 (5) ◽  
pp. 1041-1056 ◽  
Author(s):  
Alison M. Motley ◽  
Paul C. Galvin ◽  
Lakhan Ekal ◽  
James M. Nuttall ◽  
Ewald H. Hettema
Keyword(s):  
Protein Delivery ◽  
Matrix Protein ◽  
Protein Import ◽  
De Novo ◽  
Primary Role ◽  
Peroxisome Biogenesis ◽  
Peroxisomal Membrane ◽  
Related Proteins ◽  
Peroxisome Membrane

A recent model for peroxisome biogenesis postulates that peroxisomes form de novo continuously in wild-type cells by heterotypic fusion of endoplasmic reticulum–derived vesicles containing distinct sets of peroxisomal membrane proteins. This model proposes a role in vesicle fusion for the Pex1/Pex6 complex, which has an established role in matrix protein import. The growth and division model proposes that peroxisomes derive from existing peroxisomes. We tested these models by reexamining the role of Pex1/Pex6 and dynamin-related proteins in peroxisome biogenesis. We found that induced depletion of Pex1 blocks the import of matrix proteins but does not affect membrane protein delivery to peroxisomes; markers for the previously reported distinct vesicles colocalize in pex1 and pex6 cells; peroxisomes undergo continued growth if fission is blocked. Our data are compatible with the established primary role of the Pex1/Pex6 complex in matrix protein import and show that peroxisomes in Saccharomyces cerevisiae multiply mainly by growth and division.

scholarly journals Biogenesis of peroxisomes: immunocytochemical investigation of peroxisomal membrane proteins in proliferating rat liver peroxisomes and in catalase-negative membrane loops.

1989 ◽  
Vol 108 (6) ◽  
pp. 2221-2231 ◽  
Author(s):  
E Baumgart ◽  
A Völkl ◽  
T Hashimoto ◽  
H D Fahimi
Keyword(s):  
Endoplasmic Reticulum ◽  
Membrane Proteins ◽  
Smooth Endoplasmic Reticulum ◽  
Gradient Centrifugation ◽  
Normal Liver ◽  
Membrane System ◽  
Peroxisomal Membrane ◽  
Lowicryl K4m ◽  
Membrane Reservoir ◽  
Liver Peroxisomes

Treatment of rats with a new hypocholesterolemic drug BM 15766 induces proliferation of peroxisomes in pericentral regions of the liver lobule with distinct alterations of the peroxisomal membrane (Baumgart, E., K. Stegmeier, F. H. Schmidt, and H. D. Fahimi. 1987. Lab. Invest. 56:554-564). We have used ultrastructural cytochemistry in conjunction with immunoblotting and immunoelectron microscopy to investigate the effects of this drug on peroxisomal membranes. Highly purified peroxisomal fractions were obtained by Metrizamide gradient centrifugation from control and treated rats. Immunoblots prepared from such peroxisomal fractions incubated with antibodies to 22-, 26-, and 70-kD peroxisomal membrane proteins revealed that the treatment with BM 15766 induced only the 70-kD protein. In sections of normal liver embedded in Lowicryl K4M, all three membrane proteins of peroxisomes could be localized by the postembedding technique. The strongest labeling was obtained with the 22-kD antibody followed by the 70-kD and 26-kD antibodies. In treated animals, double-membraned loops with negative catalase reaction in their lumen, resembling smooth endoplasmic reticulum segments as well as myelin-like figures, were noted in the proximity of some peroxisomes. Serial sectioning revealed that the loops seen at some distance from peroxisomes in the cytoplasm were always continuous with the peroxisomal membranes. The double-membraned loops were consistently negative for glucose-6-phosphatase, a marker for endoplasmic reticulum, but were distinctly labeled with antibodies to peroxisomal membrane proteins. Our observations indicate that these membranous structures are part of the peroxisomal membrane system. They could provide a membrane reservoir for the proliferation of peroxisomes and the expansion of this intracellular compartment.

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