Vector-mediated overexpression of catalase A in the yeast Saccharomyces cerevisiae induces inclusion body formation.
To study the morphological effects of overexpression of catalase A in yeast, the gene coding for catalase A was introduced into Saccharomyces cerevisiae on a multicopy vector. After induction of microbody biogenesis and catalase A expression by growth on oleic acid as sole carbon source, cells were analyzed by immunofluorescence and immunoelectron microscopy. In addition, overexpression of catalase A was studied by quantitative immunoblotting and by activity measurement. Quantitative immunoblotting resulted in a 16-fold difference between immunoreactive material from transformed and non-transformed cells. An 18-fold increase of enzyme activity was measured in transformed cells due to overexpression of catalase A from plasmid pAH521. Immunofluorescent staining of semithin sections of Lowicryl HM20-embedded cells with anti-catalase localized peroxisomes and--at a low percentage--larger particles. By immunoelectron microscopy, these larger structures could be identified as agranular, electron-dense aggregates which are morphologically clearly distinct from the cytoplasm and not bounded by a membrane. These structures, which have been named inclusion bodies, contain catalase A but not other peroxisomal enzymes like thiolase. These findings suggest that cells are capable of compensating for overproduced proteins by formation of particular types of structures.
Binder
M
M
Institut für Tumorbiologie-Krebsforschung, Universität, Wien/Austria.
Schanz
M
M
Hartig
A
A
D000101:Acetyl-CoA C-Acetyltransferase / Q000276:immunology / Q000378:metabolism; D002374:Catalase / Q000096:biosynthesis / Q000276:immunology / Q000378:metabolism*; D005822:Genetic Vectors; D002479:Inclusion Bodies / Q000201:enzymology / Q000648:ultrastructure*; D008830:Microbodies / Q000201:enzymology / Q000648:ultrastructure; D008856:Microscopy, Fluorescence; D016253:Microscopy, Immunoelectron; D019301:Oleic Acid; D009829:Oleic Acids / Q000378:metabolism; D010957:Plasmids; D011994:Recombinant Proteins / Q000096:biosynthesis; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics; D014170:Transformation, Genetic
D009829:Oleic Acids; D011994:Recombinant Proteins; D019301:Oleic Acid; D002374:Catalase; D000101:Acetyl-CoA C-Acetyltransferase