title: The peroxisomal targeting sequence type 1 receptor, Pex5p, and the peroxisomal import efficiency of alanine:glyoxylate aminotransferase.
abstract: Unlike most organellar proteins, some peroxisomal proteins are often found in significant amounts in the cytosol. Such apparent import inefficiency is very marked in guinea pig (Cavia porcellus) hepatocytes in which the cytosolic levels of two peroxisomal proteins, catalase and alanine:glyoxylate aminotransferase (AGT), are much higher than those found in human (Homo sapiens) hepatocytes, for example. In an attempt to provide an explanation for this phenomenon, we have cloned the guinea pig CpPEX5 gene, which encodes the peroxisomal targeting sequence type 1 (PTS1) import receptor Pex5p, and functionally compared it with its human homologue, HsPex5p. Our results showed the following: (1) CpPEX5, like its human homologue, encodes two splice variants differing by the presence or absence of an internal region of 37 amino acids; (2) both variants were expressed in all guinea pig tissues studied; (3) both variants were equally able to complement peroxisomal import of PTS1 proteins in microinjected Deltapex5 human fibroblasts; (4) CpPex5p was as efficient as HsPex5p in mediating the peroxisomal import of proteins possessing the consensus PTS1, Ser-Lys-Leu, but much less efficient in mediating the import of proteins possessing non-consensus PTS1s (i.e. Lys-Lys-Leu of human AGT and Ala-Asn-Leu of human catalase); (5) reporter proteins with the consensus PTS1, Ser-Lys-Leu, inhibited the peroxisomal import of endogenous catalase, whereas AGT with the non-consensus Lys-Lys-Leu did not; (6) high concentrations of HsPex5p, but not CpPex5p, markedly inhibited the import of AGT, but not catalase or proteins ending in Ser-Lys-Leu; and (7) in the yeast two-hybrid system, AGT-Ser-Lys-Leu interacted with the tetratricopeptide repeat domain of HsPex5p, but AGT-Lys-Lys-Leu did not. In addition, AGT-Ser-Lys-Leu was targeted to peroxisomes in Saccharomyces cerevisiae, whereas AGT-Lys-Lys-Leu was not. These data suggest that the inefficient peroxisomal import of AGT and catalase in guinea pig cells is due to the inefficiency with which CpPex5p mediates the peroxisomal import of proteins containing non-consensus PTS1s. They also suggest that the non-consensus PTS1 of human AGT might interact with HsPex5p very differently compared with the consensus PTS1, Ser-Lys-Leu.
authors.affiliation: MRC Laboratory for Molecular Cell Biology and Department of Biology, University College London, Gower Street, London WC1E 6BT, U.K.
authors.forename: T G
authors.initials: TG
authors.lastname: Knott
authors.forename: G M
authors.initials: GM
authors.lastname: Birdsey
authors.forename: K E
authors.initials: KE
authors.lastname: Sinclair
authors.forename: I M
authors.initials: IM
authors.lastname: Gallagher
authors.forename: P E
authors.initials: PE
authors.lastname: Purdue
authors.forename: C J
authors.initials: CJ
authors.lastname: Danpure
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002374:Catalase / Q000378:metabolism; D002478:Cells, Cultured; D003001:Cloning, Molecular; D017931:DNA Primers; D006801:Humans; D008969:Molecular Sequence Data; D000074435:Peroxisome-Targeting Signal 1 Receptor; D020675:Peroxisomes / Q000201:enzymology / Q000378:metabolism*; D021381:Protein Transport; D018160:Receptors, Cytoplasmic and Nuclear / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D000637:Transaminases / Q000378:metabolism*; D020798:Two-Hybrid System Techniques
chemical_list: D017931:DNA Primers; D000074435:Peroxisome-Targeting Signal 1 Receptor; D018160:Receptors, Cytoplasmic and Nuclear; D002374:Catalase; D000637:Transaminases; C019635:Alanine-glyoxylate transaminase