Receptor recognition by the peroxisomal AAA complex depends on the presence of the ubiquitin moiety and is mediated by Pex1p

Schwerter, Daniel; Grimm, Immanuel; Girzalsky, Wolfgang; Erdmann, Ralf

doi:10.1074/jbc.ra118.003936

Cited by 15 publications

(15 citation statements)

References 82 publications

(93 reference statements)

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“…The three modules of Ant1p contain hydrophilic loops showing an excess of positively charged residues, suggesting that the essential requirements for mitochondrial targeting should be fulfilled. In fact, we found that in wild type yeast cells, an EFGP-Ant1p fusion protein co-localized with peroxisomes visualized with reporter protein mCherry-SKL [ 61 ]. However, the same protein showed a clear mitochondrial localization in the cells of a pex19Δ deletion strain (Additional file 5 : Fig.…”

Section: Resultsmentioning

confidence: 99%

The selectivity filter of the mitochondrial protein import machinery

et al. 2020

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Background The uptake of newly synthesized nuclear-encoded mitochondrial proteins from the cytosol is mediated by a complex of mitochondrial outer membrane proteins comprising a central pore-forming component and associated receptor proteins. Distinct fractions of proteins initially bind to the receptor proteins and are subsequently transferred to the pore-forming component for import. The aim of this study was the identification of the decisive elements of this machinery that determine the specific selection of the proteins that should be imported. Results We identified the essential internal targeting signal of the members of the mitochondrial metabolite carrier proteins, the largest protein family of the mitochondria, and we investigated the specific recognition of this signal by the protein import machinery at the mitochondrial outer surface. We found that the outer membrane import receptors facilitated the uptake of these proteins, and we identified the corresponding binding site, marked by cysteine C141 in the receptor protein Tom70. However, in tests both in vivo and in vitro, the import receptors were neither necessary nor sufficient for specific recognition of the targeting signals. Although these signals are unrelated to the amino-terminal presequences that mediate the targeting of other mitochondrial preproteins, they were found to resemble presequences in their strict dependence on a content of positively charged residues as a prerequisite of interactions with the import pore. Conclusions The general import pore of the mitochondrial outer membrane appears to represent not only the central channel of protein translocation but also to form the decisive general selectivity filter in the uptake of the newly synthesized mitochondrial proteins.

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Section: Resultsmentioning

confidence: 99%

The selectivity filter of the mitochondrial protein import machinery

et al. 2020

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“…The recycling process requires activities of other membrane bound peroxins, ubiquitinlabeling of the receptor as well as ATP-hydrolysis. The releasing process is initiated by cysteine-dependent monoubiquitination at the N-termini of Pex5p or Pex18p (Platta et al, 2007;El Magraoui et al, 2013;Schwerter et al, 2018). In the cytosol, several ways exist to remove the thioether-linked ubiquitin from the receptor (Grou et al, 2009;Debelyy et al, 2011).…”

Section: Introductionmentioning

confidence: 99%

Membrane Processing and Steady-State Regulation of the Alternative Peroxisomal Import Receptor Pex9p

Rudowitz

Erdmann

Schliebs

2020

Front. Cell Dev. Biol.

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Import of peroxisomal matrix proteins with a type 1 peroxisomal targeting signal (PTS1) in Saccharomyces cerevisiae is facilitated by cytosolic import receptors Pex5p and Pex9p. While Pex5p has a broad specificity for all PTS1 proteins independent of the growth conditions, Pex9p is only expressed in fatty-acid containing media to mediate peroxisomal import of the two malate synthases, Mls1p and Mls2p, as well as the glutathione transferase Gto1p. Pex5p-cargo complexes dock at the peroxisomal membrane, translocate their cargo-protein via a transient pore and are recycled into the cytosol for a further round of import. The processing of Pex5p has been shown to require a complex network of interactions with other membrane-bound peroxins, as well as decoration with ubiquitin as signal for its ATP-dependent release and recycling. Here, we show that the alternative receptor Pex9p requires the same set of interacting peroxins to mediate peroxisomal import of Mls1p. However, while Pex5p is rather stable, Pex9p is rapidly degraded during its normal life cycle. The steady-state regulation of Pex9p, combining oleate-induced expression with high turnover rates resembles that of Pex18p, one of the two co-receptors of the PTS2-dependent targeting pathway into peroxisomes. Both Pex9p-and Pex18p-dependent import routes serve the fast metabolic adaptation to changes of carbon sources in baker's yeast. By sequence similarities, we identified another Pex9p homolog in the human pathogenic fungus Candida glabrata, in which similar metabolic reprogramming strategies are crucial for survival of the pathogen.

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“…Thus, this would support the idea that the PEX1 binding engages the ubiquitin part of H 6 -PEX5-Ub-Strep. This has also been shown recently using a photo-crosslinking approach in an in vitro export assay 31 or in an assay with purified yeast proteins 53 . Thus, it seems likely that H 6 -PEX5-Ub-Strep provides binding-sites for the docking and REM proteins.…”

Section: Discussionmentioning

confidence: 54%

“…While this manuscript was under review similar findings, i.e . the result that the AAA ATPases Pex1p and Pex6p interact with N-terminally linked ubiquitin-Pex5p fusion protein were reported for the yeast proteins 53 .…”

Section: Discussionmentioning

confidence: 84%

Chemically monoubiquitinated PEX5 binds to the components of the peroxisomal docking and export machinery

Hagmann

Sommer

Fabian

et al. 2018

Sci Rep

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Peroxisomal matrix proteins contain either a peroxisomal targeting sequence 1 (PTS1) or a PTS2 that are recognized by the import receptors PEX5 and PEX7, respectively. PEX5 transports the PTS1 proteins and the PEX7/PTS2 complex to the docking translocation module (DTM) at the peroxisomal membrane. After cargo release PEX5 is monoubiquitinated and extracted from the peroxisomal membrane by the receptor export machinery (REM) comprising PEX26 and the AAA ATPases PEX1 and PEX6. Here, we investigated the protein interactions of monoubiquitinated PEX5 with the docking proteins PEX13, PEX14 and the REM. “Click” chemistry was used to synthesise monoubiquitinated recombinant PEX5. We found that monoubiquitinated PEX5 binds the PEX7/PTS2 complex and restores PTS2 protein import in vivo in ΔPEX5 fibroblasts. In vitro pull-down assays revealed an interaction of recombinant PEX5 and monoubiquitinated PEX5 with PEX13, PEX14 and with the REM components PEX1, PEX6 and PEX26. The interactions with the docking proteins were independent of the PEX5 ubiquitination status whereas the interactions with the REM components were increased when PEX5 is ubiquitinated.

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Receptor recognition by the peroxisomal AAA complex depends on the presence of the ubiquitin moiety and is mediated by Pex1p

Cited by 15 publications

References 82 publications

The selectivity filter of the mitochondrial protein import machinery

The selectivity filter of the mitochondrial protein import machinery

Membrane Processing and Steady-State Regulation of the Alternative Peroxisomal Import Receptor Pex9p

Chemically monoubiquitinated PEX5 binds to the components of the peroxisomal docking and export machinery

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