Toxoplasma gondii catalase: are there peroxisomes in toxoplasma?
The intracellular protozoan parasite Toxoplasma gondii, like all members of the phylum Apicomplexa, is known to possess many organelles: in addition to mitochondria and the compartments of the secretory pathway, there is a reduced chloroplast (the apicoplast) and the phylum-specific components of the apical complex: dense granules, micronemes and rhoptries. Conspicuously missing so far are microbodies, organelles that can be found in nearly all eukaryotic organisms. Microbodies show a large variation with regard to their size, number and contents, depending on the organism and cell type. One marker enzyme of this single membrane-bound organelle is catalase, which is responsible for the degradation of hydrogen peroxide to water and oxygen. The EST project in T. gondii revealed the existence of two overlapping clones which showed similarity with catalase, and these were used to clone the corresponding gene. The predicted sequence of T. gondii catalase has -AKM at the C terminus, which falls within the consensus of the PTS1 peroxisomal targeting signal. Southern blot analysis confirmed the presence of a single copy gene. Northern and western blot analyses showed that the catalase gene is transcribed and translated. Immunofluorescence assays using an antibody raised against a catalase peptide identified a distinct structure towards the apical end, but other catalase-specific antibodies failed to confirm this localisation. Cell fractionations indicated that the majority of the enzyme was in the cytosol. The fusion of the C-terminal twelve amino acids, including AKM, or the canonical peroxisomal targeting signal, -SKL, to GFP resulted in predominantly cytosolic localization in T. gondii. There was therefore no evidence for membrane-bound peroxisomes in Toxoplasma.
Ding
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Zentrum für Molekulare Biologie Heidelberg, Im Neuenheimer Feld 282, Germany. soldati@zmbh.uni-heidelberg.de
Clayton
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Soldati
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D000596:Amino Acids / Q000378:metabolism; D000818:Animals; D002374:Catalase / Q000378:metabolism*; D003001:Cloning, Molecular; D003593:Cytoplasm / Q000378:metabolism; D049452:Green Fluorescent Proteins; D007202:Indicators and Reagents; D008164:Luminescent Proteins; D008969:Molecular Sequence Data; D020675:Peroxisomes / Q000201:enzymology*; D017386:Sequence Homology, Amino Acid; D014122:Toxoplasma / Q000201:enzymology*
D000596:Amino Acids; D007202:Indicators and Reagents; D008164:Luminescent Proteins; D049452:Green Fluorescent Proteins; D002374:Catalase