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© The Rockefeller University Press, 0021-9525/1998//403 $5.00
The Journal of Cell Biology, Volume 142, Number 2, , 1998 403-420

Pex20p of the Yeast Yarrowia lipolytica Is Required for the Oligomerization of Thiolase in the Cytosol and for Its Targeting to the Peroxisome

Department of Cell Biology and Anatomy, University of Alberta, Edmonton, Alberta T6G 2H7, Canada

Pex mutants are defective in peroxisome assembly. In the pex20-1 mutant strain of the yeast Yarrowia lipolytica, the peroxisomal matrix protein thiolase is mislocalized exclusively to the cytosol, whereas the import of other peroxisomal proteins is unaffected. The PEX20 gene was isolated by functional complementation of the pex20-1 strain and encodes a protein, Pex20p, of 424 amino acids (47,274 D). Despite its role in the peroxisomal import of thiolase, which is targeted by an amino-terminal peroxisomal targeting signal-2 (PTS2), Pex20p does not exhibit homology to Pex7p, which acts as the PTS2 receptor. Pex20p is mostly cytosolic, whereas 4–8% is associated with high-speed (200,000 g) pelletable peroxisomes. In the wild-type strain, all newly synthesized thiolase is associated with Pex20p in a heterotetrameric complex composed of two polypeptide chains of each protein. This association is independent of PTS2. Pex20p is required for both the oligomerization of thiolase in the cytosol and its targeting to the peroxisome. Our data suggest that monomeric Pex20p binds newly synthesized monomeric thiolase in the cytosol and promotes the formation of a heterotetrameric complex of these two proteins, which could further bind to the peroxisomal membrane. Translocation of the thiolase homodimer into the peroxisomal matrix would release Pex20p monomers back to the cytosol, thereby permitting a new cycle of binding-oligomerization-targeting-release for Pex20p and thiolase.

Key Words: biogenesis • import • translocation • chaperone • cross-linking

Abbreviations used in this paper: AOX, acyl-CoA oxidase; CAT, catalase; DSP, dithiobis(succinimidylproprionate); DSS, disuccinimidyl suberate; FUM, fumarase; G6PDH, glucose-6-phosphate dehydrogenase; HSP, high-speed (200,000 g) pelletable; ICL, isocitrate lyase; LSP, low-speed (20,000 g) pelletable; 20KgP, 20,000 g pellet; 20KgS, 20,000 g supernatant; 200KgP, 200,000 g pellet; 200KgS, 200,000 g supernatant (cytosol); MLS, malate synthase; PNS, postnuclear supernatant; PTS, peroxisomal targeting signal; THI, thiolase.

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