Role of the PAS1 gene of Pichia pastoris in peroxisome biogenesis.
Several groups have reported the cloning and sequencing of genes involved in the biogenesis of yeast peroxisomes. Yeast strains bearing mutations in these genes are unable to grow on carbon sources whose metabolism requires peroxisomes, and these strains lack morphologically normal peroxisomes. We report the cloning of Pichia pastoris PAS1, the homologue (based on a high level of protein sequence similarity) of the Saccharomyces cerevisiae PAS1. We also describe the creation and characterization of P. pastoris pas1 strains. Electron microscopy on the P. pastoris pas1 cells revealed that they lack morphologically normal peroxisomes, and instead contain membrane-bound structures that appear to be small, mutant peroxisomes, or "peroxisome ghosts." These "ghosts" proliferated in response to induction on peroxisome-requiring carbon sources (oleic acid and methanol), and they were distributed to daughter cells. Biochemical analysis of cell lysates revealed that peroxisomal proteins are induced normally in pas1 cells. Peroxisome ghosts from pas1 cells were purified on sucrose gradients, and biochemical analysis showed that these ghosts, while lacking several peroxisomal proteins, did import varying amounts of several other peroxisomal proteins. The existence of detectable peroxisome ghosts in P. pastoris pas1 cells, and their ability to import some proteins, stands in contrast with the results reported by Erdmann et al. (1991) for the S. cerevisiae pas1 mutant, in which they were unable to detect peroxisome-like structures. We discuss the role of PAS1 in peroxisome biogenesis in light of the new information regarding peroxisome ghosts in pas1 cells.
Department of Biology, University of California, San Diego, La Jolla 92093-0322.
J A
JA
Heyman
E
E
Monosov
S
S
Subramani
D000074183:ATPases Associated with Diverse Cellular Activities; D000251:Adenosine Triphosphatases; D000595:Amino Acid Sequence; D001483:Base Sequence; D002458:Cell Fractionation; D002499:Centrifugation, Density Gradient; D003001:Cloning, Molecular; D003470:Culture Media; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D005800:Genes, Fungal; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D000432:Methanol / Q000378:metabolism; D008830:Microbodies / Q000378:metabolism* / Q000648:ultrastructure; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D019301:Oleic Acid; D009829:Oleic Acids / Q000378:metabolism; D010843:Pichia / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism / Q000648:ultrastructure; D012441:Saccharomyces cerevisiae / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins; D017386:Sequence Homology, Amino Acid; D033921:Vesicular Transport Proteins
D003470:Culture Media; D005656:Fungal Proteins; D008565:Membrane Proteins; D009829:Oleic Acids; D029701:Saccharomyces cerevisiae Proteins; D033921:Vesicular Transport Proteins; D019301:Oleic Acid; D000251:Adenosine Triphosphatases; C064404:SEC18 protein, S cerevisiae; D000074183:ATPases Associated with Diverse Cellular Activities; C511586:PEX1 protein, S cerevisiae; D000432:Methanol