The peroxisomal protein import machinery – a case report of transient ubiquitination with a new flavor

Abstract

The peroxisomal protein import machinery displays remarkable properties. Be it its capacity to accept already folded proteins as substrates, its complex architecture or its energetics, almost every aspect of this machinery seems unique. The list of unusual properties is still growing as shown by the recent finding that one of its central components, Pex5p, is transiently monoubiquitinated at a cysteine residue. However, the data gathered in recent years also suggest that the peroxisomal import machinery is not that exclusive and similarities with p97/Cdc48-mediated processes and with multisubunit RING-E3 ligases are starting to emerge. Here, we discuss these data trying to distill the principles by which this complex machinery operates.

A Cargo-centered Perspective on the PEX5 Receptor-mediated Peroxisomal Protein Import Pathway

“…Indeed, the import efficiencies and behaviors of [ 35 S]SCPx upon the peroxisome fractionation experiments were the same when using recombinant PEX5 or PEX5(C11A). Thus, contrary to previous hypotheses (23,33), neither ubiquitination of PEX5 at the DTM nor the ATP-dependent extraction of monoubiquitinated PEX5 from the DTM plays a role in the cargo protein translocation steps.…”

“…According to this model, the driving force for the protein translocation step resides in the strong protein-protein interactions that are established between PEX5 on one side and components of the DTM on the other; ATP hydrolysis is necessary only at later steps, to extract PEX5 from the DTM, thus resetting the protein transport system (see Ref. 23 and references cited therein). Although essentially all the data on PEX5 collected since then using several experimental systems are compatible with such a model (35,36), the fact remained that no direct evidence showing ATP-independent import of a peroxisomal matrix protein was available for many years.…”

“…According to current models (23)(24)(25)(26), after binding newly synthesized matrix proteins in the cytosol, PEX5 interacts with the peroxisomal docking/translocation module (DTM), a multisubunit protein assembly comprising five core components: PEX13 and PEX14 and the RING finger peroxins PEX2, PEX10, and PEX12 (27)(28)(29). This interaction results in the insertion of PEX5 into the DTM, with PEX5 acquiring a transmembrane topology (Refs.…”

Plant Peroxisomes: Biogenesis and Function

“…In yeast, Pex5p reexport requires the three RING finger peroxins Pex2p, Pex10p, and Pex12p, the ubiquitin-conjugating enzyme Pex4p and its membrane anchor Pex22p, and the two AAA ATPases Pex1p and Pex6p, which are tethered to the membrane by Pex15p. The prevailing model (Figure 3) invokes Pex5p monoubiquitination by Pex4p (E2) and Pex12p (E3) and ATPdependent dislocation of ubiquitinated Pex5p from the membrane via Pex1p and Pex6p (Grou et al, 2009). Although there is no direct evidence for PEX5 ubiquitination in plants, the machinery is conserved.…”

Identification of Ubiquitin-specific Protease 9X (USP9X) as a Deubiquitinase Acting on Ubiquitin-Peroxin 5 (PEX5) Thioester Conjugate

“…According to current models (21)(22)(23)(24), PEX5 recognizes newly synthesized proteins in the cytosol and targets them to the docking/translocation machinery (DTM), 7 a multisubunit protein complex present in the peroxisomal membrane (25,26). Here, the PEX5-cargo protein complex becomes inserted into the DTM with the concomitant translocation of the cargo protein across the organelle membrane (27,28).…”