title: Two internal type II NADH dehydrogenases of Toxoplasma gondii are both required for optimal tachyzoite growth.
abstract: In many apicomplexan parasites the entry of electrons from NADH into the electron transport chain is governed by type II NADH dehydrogenases (NDH2s) instead of a canonical complex I. Toxoplasma gondii expresses two NDH2 isoforms, TgNDH2-I and TgNDH2-II with no indication for stage-specific regulation. We dissected the orientation of both isoforms by using a split GFP assay and a protease protection assay after selective membrane permeabilization. The two approaches revealed that both TgNDH2 isoforms are internal enzymes facing with their active sites to the mitochondrial matrix. Single knockout mutants displayed a decreased replication rate and a reduced mitochondrial membrane potential, which were both more severe in the Tgndh2-II-deleted than in the Tgndh2-I-deleted mutant. Complementation with a myc-tagged, ectopic copy of the deleted gene restored the growth rate and the mitochondrial membrane potential. However, an overexpression of the remaining intact isoform could not restore the phenotype, suggesting that the two TgNDH2 isoforms are non-redundant and possess functional differences. Together, our studies indicate that although TgNDH2-I and TgNDH2-II are individually non-essential, the expression of both internal isoforms is required to maintain the mitochondrial physiology in T. gondii tachyzoites.
authors.affiliation: Institute of Medical Microbiology, University Medical Center Göttingen, Kreuzbergring 57, Göttingen D-37075, Germany.
authors.forename: San San
authors.initials: SS
authors.lastname: Lin
authors.forename: Uwe
authors.initials: U
authors.lastname: Gross
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Bohne
mesh_terms: D002460:Cell Line; D006801:Humans; D007527:Isoenzymes / Q000378:metabolism; D009245:NADH Dehydrogenase / Q000235:genetics / Q000378:metabolism*; D015800:Protozoan Proteins / Q000235:genetics / Q000378:metabolism*; D017384:Sequence Deletion; D014122:Toxoplasma / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development*; D014123:Toxoplasmosis / Q000469:parasitology*
chemical_list: D007527:Isoenzymes; D015800:Protozoan Proteins; C088908:NADH dehydrogenase II; D009245:NADH Dehydrogenase