title: Identification of Toxoplasma gondii cAMP dependent protein kinase and its role in the tachyzoite growth.
abstract: BACKGROUND
  cAMP-dependent protein kinase (PKA) has been implicated in the asexual stage of the Toxoplasma gondii life cycle through assaying the effect of a PKA-specific inhibitor on its growth rate. Since inhibition of the host cell PKA cannot be ruled out, a more precise evaluation of the role of PKA, as well as characterization of the kinase itself, is necessary.
  
  
  RESULTS
  The inhibitory effects of two PKA inhibitors, H89, an ATP-competitive chemical inhibitor, and PKI, a substrate-competitive mammalian natural peptide inhibitor, were estimated. In the in vitro kinase assay, the inhibitory effect of PKI on a recombinant T. gondii PKA catalytic subunit (TgPKA-C) was weaker compared to that on mammalian PKA-C. In a tachyzoite growth assay, PKI had little effect on the growth of tachyzoites, whereas H89 strongly inhibited it. Moreover, T. gondii PKA regulatory subunit (TgPKA-R)-overexpressing tachyzoites showed a significant growth defect.
  
  
  CONCLUSIONS
  Our data suggest that PKA plays an important role in the growth of tachyzoites, and the inhibitory effect of substrate-competitive inhibitor PKI on T. gondii PKA was low compared to that of the ATP competitive inhibitor H89.
authors.affiliation: Department of Veterinary Microbiology, Faculty of Agriculture, The University of Tokyo, Bunkyo-ku, Tokyo, Japan.
authors.forename: Hitomi
authors.initials: H
authors.lastname: Kurokawa
authors.forename: Kentaro
authors.initials: K
authors.lastname: Kato
authors.forename: Tatsuya
authors.initials: T
authors.lastname: Iwanaga
authors.forename: Tatsuki
authors.initials: T
authors.lastname: Sugi
authors.forename: Atsushi
authors.initials: A
authors.lastname: Sudo
authors.forename: Kyousuke
authors.initials: K
authors.lastname: Kobayashi
authors.forename: Haiyan
authors.initials: H
authors.lastname: Gong
authors.forename: Hitoshi
authors.initials: H
authors.lastname: Takemae
authors.forename: Frances C
authors.initials: FC
authors.lastname: Recuenco
authors.forename: Taisuke
authors.initials: T
authors.lastname: Horimoto
authors.forename: Hiroomi
authors.initials: H
authors.lastname: Akashi
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D000595:Amino Acid Sequence; D001667:Binding, Competitive; D020134:Catalytic Domain; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015971:Gene Expression Regulation, Enzymologic; D007546:Isoquinolines / Q000494:pharmacology; D008969:Molecular Sequence Data; D010455:Peptides / Q000494:pharmacology; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D013449:Sulfonamides / Q000494:pharmacology; D014122:Toxoplasma / Q000201:enzymology* / Q000254:growth & development*
chemical_list: D007546:Isoquinolines; D010455:Peptides; D047428:Protein Kinase Inhibitors; D013449:Sulfonamides; C061479:protein kinase inhibitor peptide; D000255:Adenosine Triphosphate; D017868:Cyclic AMP-Dependent Protein Kinases; C063509:N-(2-(4-bromocinnamylamino)ethyl)-5-isoquinolinesulfonamide