The ubiquitin-conjugating enzyme Pex4p of Hansenula polymorpha is required for efficient functioning of the PTS1 import machinery.
We have cloned the Hansenula polymorpha PEX4 gene by functional complementation of a peroxisome-deficient mutant. The PEX4 translation product, Pex4p, is a member of the ubiquitin-conjugating enzyme family. In H.polymorpha, Pex4p is a constitutive, low abundance protein. Both the original mutant and the pex4 deletion strain (Deltapex4) showed a specific defect in import of peroxisomal matrix proteins containing a C-terminal targeting signal (PTS1) and of malate synthase, whose targeting signal is not yet known. Import of the PTS2 protein amine oxidase and the insertion of the peroxisomal membrane proteins Pex3p and Pex14p was not disturbed in Deltapex4 cells. The PTS1 protein import defect in Deltapex4 cells could be suppressed by overproduction of the PTS1 receptor, Pex5p, in a dose-response related manner. In such cells, Pex5p is localized in the cytosol and in peroxisomes. The peroxisome-bound Pex5p specifically accumulated at the inner surface of the peroxisomal membrane and thus differed from Pex5p in wild-type peroxisomes, which is localized throughout the matrix. We hypothesize that in H. polymorpha Pex4p plays an essential role for normal functioning of Pex5p, possibly in mediating recycling of Pex5p from the peroxisome to the cytosol.
van der Klei
I J
IJ
Eukaryotic Microbiology, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Biological Centre, Kerklaan 30, 9751 NN Haren, The Netherlands. ijvdklei@biol.rug.nl
Hilbrands
R E
RE
Kiel
J A
JA
Rasmussen
S W
SW
Cregg
J M
JM
Veenhuis
M
M
D000595:Amino Acid Sequence; D001692:Biological Transport; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism*; D007425:Intracellular Membranes / Q000378:metabolism; D008025:Ligases / Q000378:metabolism*; D008830:Microbodies / Q000378:metabolism; D008969:Molecular Sequence Data; D016296:Mutagenesis; D000074435:Peroxisome-Targeting Signal 1 Receptor; D010843:Pichia / Q000201:enzymology* / Q000235:genetics; D018160:Receptors, Cytoplasmic and Nuclear / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D014452:Ubiquitins / Q000378:metabolism*
D005656:Fungal Proteins; D000074435:Peroxisome-Targeting Signal 1 Receptor; D018160:Receptors, Cytoplasmic and Nuclear; D014452:Ubiquitins; D008025:Ligases