Cysteine-specific ubiquitination protects the peroxisomal import receptor Pex5p against proteasomal degradation. Issue 3 (22nd June 2015)

• Record Type: Journal Article
• Authors: Schwartzkopff, Benjamin; Platta, Harald W.; Hasan, Sohel; Girzalsky, Wolfgang; Erdmann, Ralf
• Is Part Of: Bioscience reports. Volume 35:Issue 3(2015)
• Publication Date: 2015-06-22
• Subjects: peroxisome biogenesis -- Pex5p -- protein import -- type 1 peroxisomal targeting signal (PTS1)-receptor -- ubiquitination; Molecular biology -- Periodicals; Cytology -- Periodicals; 572.8
• DOI: 10.1042/BSR20150103
• Languages: English
• ISSNs: 0144-8463

Abstract

Abstract : Peroxisomal import receptors cycle between the peroxisomal membrane and the cytosol. A monoubiquitinated cysteine is required for efficient recycling of the peroxisomal import receptor Pex5p and prevents the protein from polyubiquitination, which leads to a rapid degradation of the protein. Abstract : Peroxisomal matrix protein import is mediated by dynamic import receptors, which cycle between the peroxisomal membrane and the cytosol. Proteins with a type 1 peroxisomal targeting signal (PTS1) are bound by the import receptor Pex5p in the cytosol and guided to the peroxisomal membrane. After cargo translocation into the peroxisomal matrix, the receptor is released from the membrane back to the cytosol in an ATP-dependent manner by the AAA-type ATPases Pex1p and Pex6p. These mechanoenzymes recognize ubiquitinated Pex5p-species as substrates for membrane extraction. The PTS1-receptor is either polyubiquitinated via peptide bonds at two certain lysines and results in proteasomal degradation or monoubiquitinated via a thioester-bond at a conserved cysteine, which enables the recycling of Pex5p and further rounds of matrix protein import. To investigate the physiological relevance of the conserved N-terminal cysteine of Pex5p, the known target amino acids for ubiquitination were substituted by site-directed mutagenesis. In contrast with Pex5pC6A, Pex5pC6K turned out to be functional in PTS1 import and utilization of oleic acid, independent of the lysines at position 18 and 24. In contrast with wild-type Pex5p, Pex5pC6K displays an ubiquitination pattern, similar to the polyubiquitination pattern of Pex4p or Pex22p mutant strains. Moreover, Pex5pC6K displays a significantly reduced steady-state level when the deubiquitinating enzyme Ubp15p is missing. Thus, our results indicate that not the cysteine residue but the position of ubiquitination is important for Pex5p function. The presence of the cysteine prevents polyubiquitination and rapid degradation of Pex5p.