The peroxisome biogenesis factors Pex4p, Pex22p, Pex1p, and Pex6p act in the terminal steps of peroxisomal matrix protein import

Peroxisomes are independent organelles found in virtually all eukaryotic ce lls. Genetic studies have identified more than 20 PEX genes that are requir ed for peroxisome biogenesis. The role of most PEX gene products, peroxins, remains to be determined, but a variety of studies have established that P ex5p binds the type 1 peroxisomal targeting signal and is the import recept or for most newly synthesized peroxisomal matrix proteins. The steady-state abundance of Pex5p is unaffected in most per mutants of the yeast Pichia p astoris but is severely reduced in pex4 and pex22 mutants and moderately re duced in pex1 and pex6 mutants. We used these subphenotypes to determine th e epistatic relationships among several groups of pex mutants. Our results demonstrate that Pex4p acts after the peroxisome membrane synthesis factor Pex3p, the Pex5p docking factors Pex13p and Pex14p, the matrix protein impo rt factors Pex8p, Pex10p, and Pex12p, and two other peroxins, Pex2p and Pex 17p. Pex22p and the interacting AAA ATPases Pex1p and Pex6p were also found to act after Pex10p. Furthermore, Pex1p and Pex6p were found to act upstre am of Pex4p and Pex22p. These results suggest that Pex1p, Pex4p, Pex6p, and Pex22p act late in peroxisomal matrix protein import, after matrix protein translocation. This hypothesis is supported by the phenotypes of the corre sponding mutant strains. As has been shown previously for P. pastoris pert, pex6, and pex22 mutant cells, we show here that pex4 Delta mutant cells co ntain peroxisomal membrane protein-containing peroxisomes that import resid ual amounts of peroxisomal matrix proteins.