This Article Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Liu, H Articles by Cregg, J M Search for Related Content PubMed PubMed Citation Articles by Liu, H Articles by Cregg, J M

research-article

An efficient screen for peroxisome-deficient mutants of Pichia pastoris.

Department of Chemical and Biological Sciences, Oregon Graduate Institute of Science and Technology, Beaverton 97006-1999.

ABSTRACT

We describe a rapid and efficient screen for peroxisome-deficient (per) mutants in the yeast Pichia pastoris. The screen relies on the unusual ability of P. pastoris to grow on two carbon sources, methanol and oleic acid, both of which absolutely require peroxisomes to be metabolized. A collection of 280 methanol utilization-defective (Mut-) P. pastoris mutants was isolated, organized into 46 complementation groups, and tested for those that were also oleate-utilization defective (Out-) but still capable of growth on ethanol and glucose. Mutants in 10 groups met this phenotypic description, and 8 of these were observed by electron microscopy to be peroxisome deficient (Per-). In each per mutant, Mut-, Out-, and Per- phenotypes were tightly linked and therefore were most likely due to a mutation at a single locus. Subcellular fractionation experiments indicated that the peroxisomal marker enzyme catalase was mislocalized to the cytosol in both methanol- and oleate-induced cultures of the mutants. In contrast, alcohol oxidase, a peroxisomal methanol utilization pathway enzyme, was virtually absent from per mutant cells. The relative ease of per mutant isolation in P. pastoris, in conjunction with well-developed procedures for its molecular and genetic manipulation, makes this organism an attractive system for studies on peroxisome biogenesis.

This article has been cited by other articles:

• Nazarko, T. Y., Polupanov, A. S., Manjithaya, R. R., Subramani, S., Sibirny, A. A. (2007). The Requirement of Sterol Glucoside for Pexophagy in Yeast Is Dependent on the Species and Nature of Peroxisome Inducers. Mol. Biol. Cell 18: 106-118 [Abstract] [Full Text]  
• Lin-Cereghino, G. P., Godfrey, L., de la Cruz, B. J., Johnson, S., Khuongsathiene, S., Tolstorukov, I., Yan, M., Lin-Cereghino, J., Veenhuis, M., Subramani, S., Cregg, J. M. (2006). Mxr1p, a Key Regulator of the Methanol Utilization Pathway and Peroxisomal Genes in Pichia pastoris. Mol. Cell. Biol. 26: 883-897 [Abstract] [Full Text]  
• Johnson, M. A., Waterham, H. R., Ksheminska, G. P., Fayura, L. R., Cereghino, J. L., Stasyk, O. V., Veenhuis, M., Kulachkovsky, A. R., Sibirny, A. A., Cregg, J. M. (1999). Positive Selection of Novel Peroxisome Biogenesis-Defective Mutants of the Yeast Pichia pastoris. Genetics 151: 1379-1391 [Abstract] [Full Text]  
• Elgersma, Y., Elgersma-Hooisma, M., Wenzel, T., McCaffery, J. M., Farquhar, M. G., Subramani, S. (1998). A Mobile PTS2 Receptor for Peroxisomal Protein Import in Pichia pastoris. J. Cell Biol. 140: 807-820 [Abstract] [Full Text]  
• SUBRAMANI, S. (1998). Components Involved in Peroxisome Import, Biogenesis, Proliferation, Turnover, and Movement. Physiol. Rev. 78: 171-188 [Abstract] [Full Text]  
• Waterham, H. R., Russell, K. A., Vries, Y. d., Cregg, J. M. (1997). Peroxisomal Targeting, Import, and Assembly of Alcohol Oxidase in Pichia pastoris. J. Cell Biol. 139: 1419-1431 [Abstract] [Full Text]  
• Eitzen, G. A., Titorenko, V. I., Smith, J. J., Veenhuis, M., Szilard, R. K., Rachubinski, R. A. (1996). The Yarrowia lipolytica Gene PAY5 Encodes a Peroxisomal Integral Membrane Protein Homologous to the Mammalian Peroxisome Assembly Factor PAF-1. J. Biol. Chem. 271: 20300-20306 [Abstract] [Full Text]  
• Wiemer, ErikA.C., Luers, GeorgH., Faber, K., Wenzel, T., Veenhuis, M., Subramani, S. (1996). Isolation and Characterization of Pas2p, a Peroxisomal Membrane Protein Essential for Peroxisome Biogenesis in the Methylotrophic Yeast Pichia pastoris. J. Biol. Chem. 271: 18973-18980 [Abstract] [Full Text]  
• Liu, H., Tan, X., Russell, K. A., Veenhuis, M., Cregg, J. M. (1995). PER3, a Gene Required for Peroxisome Biogenesis in Pichia pastoris, Encodes a Peroxisomal Membrane Protein Involved in Protein Import. J. Biol. Chem. 270: 10940-10951 [Abstract] [Full Text]  
• Eitzen, G. A., Aitchison, J. D., Szilard, R. K., Veenhuis, M., Nuttley, W. M., Rachubinski, R. A. (1995). The Yarrowia lipolytica Gene PAY2 Encodes a 42-kDa Peroxisomal Integral Membrane Protein Essential for Matrix Protein Import and Peroxisome Enlargement but Not for Peroxisome Membrane Proliferation. J. Biol. Chem. 270: 1429-1436 [Abstract] [Full Text]  
• Slawecki, M., Dodt, G, Steinberg, S, Moser, A., Moser, H., Gould, S. (1995). Identification of three distinct peroxisomal protein import defects in patients with peroxisome biogenesis disorders. J. Cell Sci. 108: 1817-1829 [Abstract]