Pex13p Is an SH3 Protein of the Peroxisome Membrane and a Docking Factor for the Predominantly Cytoplasmic PTSl Receptor

Import of newly synthesized PTS1 proteins into the peroxisome requires the PTS1 receptor (Pex5p), a predominantly cytoplasmic protein that cycles between the cytoplasm and peroxisome. We have identified Pex13p, a novel integral peroxisomal membrane from both yeast and humans that binds the PTS1 receptor via a cytoplasmically oriented SH3 domain. Although only a small amount of Pex5p is bound to peroxisomes at steady state (<5%), loss of Pex13p further reduces the amount of peroxisome-associated Pex5p by ~40-fold. Furthermore, loss of Pex13p eliminates import of peroxisomal matrix proteins that contain either the type-1 or type-2 peroxisomal targeting signal but does not affect targeting and insertion of integral peroxisomal membrane proteins. We conclude that Pexl3p functions as a docking factor for the predominantly cytoplasmic PTS1 receptor. C OMPARTMENTALIZATION of proteins within subcellular organelles is a hallmark of eukaryotic cells. Accordingly, eukaryotic cells have developed mechanisms for recognizing newly synthesized organellar proteins and directing them to their proper destination. In the case of protein import into peroxisomes, newly synthesized peroxisomal matrix proteins are distinguished from other cytoplasmic proteins by the presence of a peroxisomal targeting signal (PTS) 1 within their structure. The PTS1 consists of a COOH-terminal tripeptide of the sequence serine-lysine-leucine-cooH, or a conservative variant, and is used by almost all proteins destined for the peroxisome lumen (Gould et al., 1989; Subramani, 1993). A second signal, PTS2, also directs proteins to the peroxisome lumen but differs from PTS1 because it is found at the NH2 terminus and is used much less commonly (SubraPlease address all correspondence to either S.J. Gould, The Department of Biological Chemistry, The Johns Hopkins University School of Medicine, 725 North Wolfe Street, Baltimore, MD 21205. Tel.: (410) 955-3085. Fax: (410) 955-0215. E-Mail: [email protected] or D.I. Crane, The School of Biomolecular and Biomedical Science, Griffith University, Brisbane, QLD 4111, Australia. Tel.: 61 7 875 7253. Fax: 61 7 875 7656. E-Mail: [email protected] 1. Abbreviat ions used in this paper: dbEST, database of expressed sequence tags; GFP, green fluorescent protein; IPMP, integral peroxisomal membrane protein; MBP, maltose-binding protein: ORF, open reading frame; SH3, src homology-3: PTS, peroxisomal targeting signal; TPR, tetratricopeptide repeat. mani, 1993; Swinkels et al., 199l). Integral peroxisomal membrane proteins use neither PTS1 nor PTS2, but rather a distinct type of signal (McCammon et al., 1994; Dyer et al., 1996). Interestingly, proteins devoid of PTS1 and PTS2 can still be imported into the peroxisome lumen, provided that they oligomerize with a PTS1 or PTS2 protein before import (Glover et al., 1994; McNew and Goodman, 1994). The hypothesis that these oligomers may be translocated intact across the peroxisome membrane is supported by the observation that PTSl-coated gold particles can be imported into peroxisomes in vivo (Walton et al., 1995). Import of PTS1 and PTS2 proteins requires a host of peroxisome assembly factors, or peroxins (Distel et al., 1996), that include specific PTS receptors (Dodt et al., 1995; McCollum et al., 1993; Marzioch et al., 1994), two ATPases (Erdmann et al., 1991; Spong and Subramani, 1993; Yahraus et al., 1996), a ubiquitin-conjugating enzyme (Crane et al., 1994; Wiebel and Kunau, 1992), three zinc-binding integral peroxisomal membrane proteins (IPMPs) (Kalish et al., 1995, 1996; Kunau et al., 1993; Tsukamoto et al., 1991; Gould, S.J., unpublished observations), and several other novel proteins (Kunau et al., 1993; E1gersma et al., 1993). Identification of these factors has depended upon the isolation of yeast mutants deficient in import of peroxisomal proteins (Subramani, 1993). Significant contributions to understanding peroxisome assembly and peroxisomal protein import have also been obtained from analysis of the peroxisome biogenesis disorders (PBD). © The Rockefeller University Press, 0021-9525/96/10/85/11 $2.00 The Journal of Cell Biology, Volume 135, Number I, October 1996 85-95 85 on F ebuary 3, 2013 jcb.rress.org D ow nladed fom Published October 1, 1996