Isolation and characterization of peroxisomal protein import (Pim−) mutants of Hansenula polymorpha

In the course of our studies on the molecular mechanisms involved in peroxisome biogenesis, we have isolated several mutants of the methylotrophic yeast Hansenula polymorpha impaired in the import of peroxisomal matrix proteins. These mutants are characterized by the presence of few small intact peroxisomes, while the bulk of the peroxisomal matrix protein is not imported and resides in the cytosol (Pim- phenotype). Genetic analysis of back-crossed mutants revealed five different complementation groups, which were designated PER1-PER5. Mapping studies to determine the linkage relationships indicated that the observed Pim- phenotypes were determined by single recessive nuclear mutations. The different mutants had comparable phenotypes: (i) they were impaired to utilize methanol as the sole source of carbon and energy but grew well on various other compounds, including nitrogen sources, the metabolism of which is known to be mediated by peroxisome-borne enzymes in wild-type cells; (ii) all peroxisomal enzymes tested were induced, assembled and activated as in wild-type cells although their activities varied between the different representative mutants; (iii) all peroxisomal proteins, whether constitutive or inducible, were found both in the cytosol and in the small peroxisomes. These results suggest that a general, major import mechanism is affected in all mutants.