title: Eci1p uses a PTS1 to enter peroxisomes: either its own or that of a partner, Dci1p.
abstract: Saccharomyces cerevisiae delta3,delta2-enoyl-CoA isomerase (Eci1p), encoded by ECI1, is an essential enzyme for the betaoxidation of unsaturated fatty acids. It has been reported, as well as confirmed in this study, to be a peroxisomal protein. Unlike many other peroxisomal proteins, Ecilp possesses both a peroxisome targeting signal type 1 (PTS1)-like signal at its carboxy-terminus (-HRL) and a PTS2-like signal at its amino-terminus (RIEGPFFIIHL). We have found that peroxisomal targeting of a fusion protein consisting of Eci1p in front of green fluorescent protein (GFP) is not dependent on Pex7p (the PTS2 receptor), ruling out a PTS2 mechanism, but is dependent on Pex5p (the PTS1 receptor). This Pex5p-dependence was unexpected, since the putative PTS1 of Ecilp is not at the C-terminus of the fusion protein; indeed, deletion of this signal (-HRL-) from the fusion did not affect the Pex5p-dependent targeting. Consistent with this, Pex5p interacted in two-hybrid assays with both Eci1p and Eci1PdeltaHRL. Ecilp-GFP targeting and Eci1pdeltaHRL interaction were abolished by replacement of Pex5p with Pex5p(N495K), a point-mutated Pex5p that specifically abolishes the PTS1 protein import pathway. Thus, Eci1p peroxisomal targeting does require the Pex5p-dependent PTS1 pathway, but does not require a PTS1 of its own. By disruption of ECI1 and DCI1, we found that Dci1p, a peroxisomal PTS1 protein that shares 50% identity with Eci1p, is necessary for Eci1p-GFP targeting. This suggests that the Pex5p-dependent import of Eci1p-GFP is due to interaction and co-import with Dci1p. Despite the dispensability of the C-terminal HRL for import in wild-type cells, we have also shown that this tripeptide can function as a PTS1, albeit rather weakly, and is essential for targeting in the absence of Dci1p. Thus, Eci1p can be targeted to peroxisomes by its own PTS1 or as a hetero-oligomer with Dcilp. These data demonstrate a novel, redundant targeting pathway for Eci1p.
authors: Department of Cell Biology and Anatomy, Mount Sinai School of Medicine, New York, NY 10029-6574, USA
authors: X
authors: X
authors: Yang
authors: P E
authors: PE
authors: Purdue
authors: P B
authors: PB
authors: Lazarow
mesh_terms: D019748:Carbon-Carbon Double Bond Isomerases / Q000235:genetics / Q000378:metabolism*; D003470:Culture Media; D064006:Dodecenoyl-CoA Isomerase; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism*; D019301:Oleic Acid / Q000378:metabolism; D000074437:Peroxisomal Targeting Signal 2 Receptor; D000074435:Peroxisome-Targeting Signal 1 Receptor; D020675:Peroxisomes / Q000378:metabolism*; D017354:Point Mutation; D011485:Protein Binding; D018160:Receptors, Cytoplasmic and Nuclear / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000254:growth & development / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D020798:Two-Hybrid System Techniques
chemical_list: D003470:Culture Media; D008164:Luminescent Proteins; D008565:Membrane Proteins; D000074437:Peroxisomal Targeting Signal 2 Receptor; D000074435:Peroxisome-Targeting Signal 1 Receptor; D018160:Receptors, Cytoplasmic and Nuclear; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; D049452:Green Fluorescent Proteins; D019301:Oleic Acid; D019748:Carbon-Carbon Double Bond Isomerases; C120150:DCI1 protein, S cerevisiae; D064006:Dodecenoyl-CoA Isomerase