Targeting of the 22 kDa integral peroxisomal membrane protein

Pause, Birgit; Saffrich, Rainer; Hunziker, Andreas; Ansorge, Wilhelm; Just, Wilhelm W.

doi:10.1016/s0014-5793(00)01332-6

Cited by 43 publications

(40 citation statements)

References 32 publications

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“…Interestingly, the Pex19p-binding site motif resembles a motif of hydrophobic and basic amino acids that has been implicated recently in targeting of mammalian PMPs (Pause et al, 2000;Brosius et al, 2002;Biermanns et al, 2003;Landgraf et al, 2003). This opens the possibility that also in higher eukaryotes Pex19p binds the targeting-specific regions of mPTSs, which however needs to be studied in future work.…”

Section: Discussionmentioning

confidence: 93%

“…The targeting signals of PMPs (mPTS) that direct and insert PMPs into the peroxisomal membrane have been determined for a number of PMPs of several species. Despite some differences, a picture emerged from these studies of a targeting signal consisting of one or more transmembrane domains in conjunction with a short sequence, which contains either a cluster of basic residues or a mixture of basic and hydrophobic amino acids (Dyer et al, 1996;Baerends et al, 2000b;Pause et al, 2000;Honsho and Fujiki, 2001;Wang et al, 2001;Honsho et al, 2002;Biermanns et al, 2003;Murphy et al, 2003). Although some similarities among these basic sequences have been noted, a clear consensus could not yet be deduced.…”

Section: Introductionmentioning

confidence: 99%

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Peroxisomal Membrane Proteins Contain Common Pex19p-binding Sites that Are an Integral Part of Their Targeting Signals

Rottensteiner

Kramer

Lorenzen

et al. 2004

MBoC

161

196

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Targeting of peroxisomal membrane proteins (PMPs) is a multistep process that requires not only recognition of PMPs in the cytosol but also their insertion into the peroxisomal membrane. As a consequence, targeting signals of PMPs (mPTS) are rather complex. A candidate protein for the PMP recognition event is Pex19p, which interacts with most PMPs. However, the respective Pex19p-binding sites are ill-defined and it is currently disputed whether these sites are contained within mPTS. By using synthetic peptide scans and yeast two-hybrid analyses, we determined and characterized Pex19p-binding sites in Pex11p and Pex13p, two PMPs from Saccharomyces cerevisiae. The sites turned out to be composed of a short helical motif with a minimal length of 11 amino acids. With the acquired data, it proved possible to predict and experimentally verify Pex19p-binding sites in several other PMPs by applying a pattern search and a prediction matrix. A peroxisomally targeted Pex13p fragment became mislocalized to the endoplasmic reticulum in the absence of its Pex19p-binding site. By adding the heterologous binding site of Pex11p, peroxisomal targeting of the Pex13p fragment was restored. We conclude that Pex19p-binding sites are well-defined entities that represent an essential part of the mPTS. INTRODUCTIONPeroxisomes are ubiquitous organelles of eukaryotic cells, whose proteins are imported posttranslationally. Matrix proteins are directed to peroxisomes by either of two targeting signals, a C-terminal PTS1 or an N-terminal PTS2. The topogenesis of peroxisomal membrane proteins (PMPs) is accomplished by yet another mechanism, because most of the peroxin mutants, characterized by their defect in the biogenesis of peroxisomes, exhibit a block in matrix protein import, but do import PMPs normally (Lazarow and Fujiki, 1985;Gould and Valle, 2000;Subramani et al., 2000;Purdue and Lazarow, 2001;Eckert and Erdmann, 2003). To date, only three peroxins with a potential role in PMP targeting have been identified, namely Pex3p (Hettema et al., 2000;South et al., 2000), Pex16p in mammals (South and Gould, 1999;Honsho et al., 2002), and Pex19p (Gö tte et al., 1998;Matsuzono et al., 1999;Snyder et al., 1999;Soukupova et al., 1999). In cells lacking any of these proteins, PMPs are either degraded or mistargeted to other subcellular compartments such as mitochondria, the endoplasmic reticulum (ER) and membranes of unknown origin (Ghaedi et al., 2000;Hettema et al., 2000;Sacksteder et al., 2000).In accordance with a distinct pathway, PMPs use neither PTS1 nor PTS2. The targeting signals of PMPs (mPTS) that direct and insert PMPs into the peroxisomal membrane have been determined for a number of PMPs of several species. Despite some differences, a picture emerged from these studies of a targeting signal consisting of one or more transmembrane domains in conjunction with a short sequence, which contains either a cluster of basic residues or a mixture of basic and hydrophobic amino acids (Dyer et al., 1996;Baerends et al., 2000b;Pause et al., 2000;...

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Section: Discussionmentioning

confidence: 93%

Section: Introductionmentioning

confidence: 99%

Peroxisomal Membrane Proteins Contain Common Pex19p-binding Sites that Are an Integral Part of Their Targeting Signals

Rottensteiner

Kramer

Lorenzen

et al. 2004

MBoC

161

196

View full text Add to dashboard Cite

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“…via the endoplasmic reticulum; see below). Thus, although there are no doubts that at least PMP22 and PMP70 are inserted into the peroxisomal membrane directly from the cytosol in liver cells (31,49), we reasoned that no assumptions should be made when studying the biogenesis pathway of a given PMP for the first time. These reservations are justified by recent data showing that some peroxisomal proteins reach the organelle membrane after traveling first through the endoplasmic reticulum (23, 50 -55).…”

Section: Discussionmentioning

confidence: 99%

The Import Competence of a Peroxisomal Membrane Protein Is Determined by Pex19p before the Docking Step

Pinto

Grou

Alencastre

et al. 2006

Journal of Biological Chemistry

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Biogenesis of the mammalian peroxisomal membrane requires the action of Pex3p and Pex16p, two proteins present in the organelle membrane, and Pex19p, a protein that displays a dual subcellular distribution (peroxisomal and cytosolic). Pex19p interacts with most peroxisomal intrinsic membrane proteins, but whether this property reflects its role as an import receptor for this class of proteins or a chaperone-like function in the assembly/disassembly of peroxisomal membrane proteins has been the subject of much controversy. Here, we describe an in vitro system particularly suited to address this issue. It is shown that insertion of a reporter protein into the peroxisomal membrane is a Pex3p-dependent process that does not require ATP/GTP hydrolysis. The system can be programmed with recombinant versions of Pex19p, allowing us to demonstrate that Pex19p-cargo protein complexes formed in the absence of peroxisomes are the substrates for the peroxisomal docking/ insertion machinery. Data suggesting that cargo-loaded Pex19p displays a much higher affinity for Pex3p than Pex19p alone are also provided. These results suggest that soluble Pex19p participates in the targeting of newly synthesized peroxisomal membrane proteins to the organelle membrane and support the existence of a cargo-induced peroxisomal targeting mechanism for Pex19p.

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“…In addition to the demonstrated importance of matrix-oriented basic clusters in targeting to the peroxisomal membrane, a short cytosol-oriented region adjacent to TMD1 of rat Pmp22 has been shown to contain a peroxisomal targeting signal (14). Similarly, data suggest that a cytosolic region adjacent to the membrane span of Pex15p is required for peroxisomal localization (11).…”

Section: A Quantitative Assay For Peroxisomal Localization-mentioning

confidence: 97%

“…Similarly, a basic cluster from the cotton seed ascorbate peroxidase (APX), fused to a synthetic membrane span and carrier protein, targets to punctate and reticular structures, although peroxisomal localization was not demonstrated (13). In contrast, a cytosol-oriented region of 22 amino acids in rat Pmp22 can target a different fragment of the protein, which by itself is not sufficient for targeting, to the peroxisomal membrane (14). Because some peroxisomal membrane proteins integrate into peroxisomal precursor organelles before arriving at peroxisomes (15), a more complex targeting signal may be required to accommodate such a pathway.…”

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confidence: 99%

Discrete Targeting Signals Direct Pmp47 to Oleate-induced Peroxisomes in Saccharomyces cerevisiae

Wang

Unruh

Goodman

2001

Journal of Biological Chemistry

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Pmp47 is a peroxisomal membrane protein consisting of six transmembrane domains (TMDs). We previously showed that the second matrix loop containing a basic cluster of amino acids is important for peroxisomal targeting, and similar basic targeting motifs have been found in other peroxisomal membrane proteins. However, this basic cluster by itself targets to peroxisomes very poorly. We have developed a sensitive quantitative localization assay based on the targeting of Pmp47-GFP fusion proteins to identify the important elements of the basic cluster and to search for other targeting information on Pmp47. Our data suggest that side-chain structure and position as well as charge are important for targeting by the basic cluster. Analysis of other regions of Pmp47 indicates that all TMDs except TMD2 can be eliminated or substituted without significant loss of targeting. TMD2 plus an adjacent cytoplasmic-oriented sequence is crucial for targeting. Cytoplasmic-oriented sequences from two other peroxisomal membrane proteins, ScPex15p and ScPmp22, could partially substitute for the analogous sequence in Pmp47. Targeting with high fidelity to oleate-induced peroxisomes required the following elements: the cytoplasmic-oriented sequence and TMD2, a short matrix loop containing a basic cluster, and a membrane-anchoring TMD.Peroxisomes consist of a dense matrix and surrounding membrane. Proteins are targeted to the peroxisomal matrix from the cytosol by virtue of peroxisomal targeting signals (PTSs) 1 (1). Most matrix proteins contain a PTS1 sequence at their extreme carboxyl terminus consisting of three amino acids identical or related to the tripeptide SKL. A few matrix proteins contain a PTS2 sequence close to the amino terminus, which is often cleaved upon entering the peroxisomal assembly pathway. Receptors for PTS1 and PTS2 have been identified, as have docking proteins for the PTS receptors on the peroxisomal surface (2, 3). Receptor and cargo may translocate through the membrane before receptors are recycled to the cytosol (4, 5).Integral membrane proteins lack these PTS sequences and must be targeted by alternative sequences. We have used Pmp47 from the methylotrophic yeast Candida boidinii as a model protein to understand targeting to the peroxisome membrane. Pmp47 shares homology with members of the mitochondrial carrier protein family (6). These proteins span the membrane six times, and sequence similarity within them suggests that the family was derived from an ancient gene triplication. Pmp47 is important in the metabolism of medium-chain fatty acids and may transport ATP (7), and it is also critical for the import of dihydroxyacetone synthase, an abundant matrix protein (8). Pmp47 contains two loops of amino acids on the matrix side and three on the cytoplasmic side of the membrane. We previously identified a targeting motif contained within the second matrix loop that we termed the mPTS (9). We showed that the most important attribute of the targeting loop is a basic cluster of amino acids of the sequence KIKKR...

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Targeting of the 22 kDa integral peroxisomal membrane protein

Cited by 43 publications

References 32 publications

Peroxisomal Membrane Proteins Contain Common Pex19p-binding Sites that Are an Integral Part of Their Targeting Signals

Peroxisomal Membrane Proteins Contain Common Pex19p-binding Sites that Are an Integral Part of Their Targeting Signals

The Import Competence of a Peroxisomal Membrane Protein Is Determined by Pex19p before the Docking Step

Discrete Targeting Signals Direct Pmp47 to Oleate-induced Peroxisomes in Saccharomyces cerevisiae

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