Pex5p is the receptor for the peroxisomal targeting signal 1 (PTS1) that consists of a C-terminal tripeptide (consensus (S/A/C)(K/R/H)(L/M)). Hexadecapeptides recognized by Pex5p from Homo sapiens and Saccharomyces cerevisiae were identified by screening a two-hybrid peptide library, and the targeting ability of the peptides was demonstrated using the green fluorescent protein as reporter. The PTS1 receptors recognized in a species-specific manner a broad range of C-terminal tripeptides, and these are reported herein. In addition, residues upstream of the tripeptide influenced the strength of the interaction in the two-hybrid system as well as in an in vitro competition assay. In peptides interacting with the human protein, hydrophobic residues were found with high frequency especially at positions -2 and -5, whereas peptides interacting with S. cerevisiae Pex5p were more hydrophilic and frequently contained arginine at position -2. In instances where the terminal tripeptide deviated from the consensus, upstream residues exerted a greater influence on the ability of the hexadecapeptides to bind Pex5p.

Dynamic changes of membrane structure are intrinsic to organelle morphogenesis and homeostasis. Ectopic expression of proteins of the PEX11 family from yeast, plant or human lead to the formation of juxtaposed elongated peroxisomes (JEPs),which is evocative of an evolutionary conserved function of these proteins in membrane tubulation. Microscopic examinations reveal that JEPs are composed of independent elongated peroxisomes with heterogeneous distribution of matrix proteins. We established the homo- and heterodimerization properties of the human PEX11 proteins and their interaction with the fission factor hFis1, which is known to recruit the GTPase DRP1 to the peroxisomal membrane. We show that excess of hFis1 but not of DRP1 is sufficient to fragment JEPs into normal round-shaped organelles, and illustrate the requirement of microtubules for JEP formation. Our results demonstrate that PEX11-induced JEPs represent intermediates in the process of peroxisome membrane proliferation and that hFis1 is the limiting factor for progression. Hence, we propose a model for a conserved role of PEX11 proteins in peroxisome maintenance through peroxisome polarization, membrane elongation and segregation.