Use of the Kinase Inhibitor Analog 1NM-PP1 Reveals a Role for Toxoplasma gondii CDPK1 in the Invasion Step

Sugi, Tatsuki; Kato, Kentaro; Kobayashi, Kanae; Watanabe, Shumpei; Kurokawa, Hitomi; Gong, Haiyan; Pandey, Kishor; Takemae, Hitoshi; Akashi, Hiroomi

doi:10.1128/ec.00351-09

Cited by 54 publications

(65 citation statements)

References 25 publications

(22 reference statements)

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“…Support for this model is provided by inhibition of PKG with Compound 1 (15,17), and selective inhibition of CDPK1 with pyrazolopyrimidine inhibitors (8,47). Both classes of compounds offer promise for development of alternative therapeutics, given the essential nature of these kinases and the potent and specific inhibition offered by these respective chemical scaffolds (48,49).…”

Section: Discussionmentioning

confidence: 99%

Serum Albumin Stimulates Protein Kinase G-dependent Microneme Secretion in Toxoplasma gondii

Brown

Lourido

Sibley

2016

Journal of Biological Chemistry

103

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Microneme secretion is essential for motility, invasion, and egress in apicomplexan parasites. Although previous studies indicate that Ca 2؉ and cGMP control microneme secretion, little is known about how these pathways are naturally activated. Here we have developed genetically encoded indicators for Ca 2؉ and microneme secretion to better define the signaling pathways that regulate these processes in Toxoplasma gondii. We found that microneme secretion was triggered in vitro by exposure to a single host protein, serum albumin. The natural agonist serum albumin induced microneme secretion in a protein kinase G-dependent manner that correlated with increased cGMP levels. Surprisingly, serum albumin acted independently of elevated Ca 2؉ and yet it was augmented by artificial agonists that raise Ca 2؉ , such as ethanol. Furthermore, although ethanol elevated intracellular Ca 2؉ , it alone was unable to trigger secretion without the presence of serum or serum albumin. This dichotomy was recapitulated by zaprinast, a phosphodiesterase inhibitor that elevated cGMP and separately increased Ca 2؉ in a protein kinase G-independent manner leading to microneme secretion. Taken together, these findings reveal that microneme secretion is centrally controlled by protein kinase G and that this pathway is further augmented by elevation of intracellular Ca 2؉ .Toxoplasma gondii is an important opportunistic pathogen and model organism for studying the biology of members of the phylum Apicomplexa (1). Micronemes are specialized secretory vesicles present in all motile stages of apicomplexan parasites (reviewed in Ref. 2). The majority of internal microneme (MIC) 3 proteins (i.e. cargo) consist of adhesive proteins that translocate to the surface of the parasite following the regulated fusion of the organelle with the apical plasma membrane.

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Section: Discussionmentioning

confidence: 99%

Serum Albumin Stimulates Protein Kinase G-dependent Microneme Secretion in Toxoplasma gondii

Brown

Lourido

Sibley

2016

Journal of Biological Chemistry

103

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“…A Tet-inducible knockout of TgCDPK1 blocks the secretion of micronemes, which results in impaired gliding motility, invasion, and egress (Lourido et al, 2010). Several specific inhibitors were designed against TgCDPK1, including pyrazolopyrimidine derivatives for which a gatekeeper mutant was shown to restore microneme secretion in the presence of the drug (Johnson et al, 2012;Kieschnick et al, 2001;Ojo et al, 2010;Sugi et al, 2010).…”

Section: Microneme Secretion and Egressmentioning

confidence: 99%

Does protein phosphorylation govern host cell entry and egress by the Apicomplexa?

Jacot

Soldati‐Favre

2012

International Journal of Medical Microbiology

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a b s t r a c tMembers of the phylum Apicomplexa are responsible for a wide range of diseases in humans and animals. The absence of an effective vaccine or safe curing drugs and the continuous emergence of resistant parasites to available treatments impose a high demand on the identification of novel targets for intervention against the apicomplexans. Protein kinases are considered attractive potential therapeutic targets not only against cancers but also to combat infectious diseases. The scope and aim of this review is to report on the recent progress in dissecting the impact of protein phosphorylation in regulating motility and invasion.

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“…BKIs selectively inhibit analog-sensitive kinases, which contain a small amino acid at the gate site of the ATP binding pocket and are rare in mammalian genomes [1]. Using in silico prediction, we found that T. gondii calcium-dependent protein kinase 1 (TgCDPK1), which has a glycine residue at the gate site of the ATP binding pocket, was the main T. gondii analog-sensitive kinase [12]. In silico prediction, an in vitro culture assay using transgenic parasites, and a structural analysis of TgCDPK1 all showed that TgCDPK1 is a target of BKI [6,8,12].…”

mentioning

confidence: 99%

“…Using in silico prediction, we found that T. gondii calcium-dependent protein kinase 1 (TgCDPK1), which has a glycine residue at the gate site of the ATP binding pocket, was the main T. gondii analog-sensitive kinase [12]. In silico prediction, an in vitro culture assay using transgenic parasites, and a structural analysis of TgCDPK1 all showed that TgCDPK1 is a target of BKI [6,8,12]. TgCDPK1 is involved in parasite attachment, invasion, egress, and secretion in vitro [5,6].…”

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confidence: 99%

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1NM-PP1 Treatment of Mice Infected with Toxoplasma gondii

Sugi

Kato

Kobayashi

et al. 2011

The Journal of Veterinary Medical Science

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ABSTRACT. Bumped kinase inhibitors (BKIs) target analog-sensitive kinases, which the genomes of mammals rarely encode. Previously, we demonstrated that a BKI effectively suppressed the in vitro replication of Toxoplasma gondii, the causative pathogen of toxoplasmosis, by targeting T. gondii calcium-dependent protein kinase 1 (TgCDPK1) (Eukaryotic Cell, 9: 667-670). Here, we examined whether the BKI 1NM-PP1 reduced parasite replication in vivo. A high dose of 1NM-PP1, by intraperitoneal injection, just before the parasite inoculation effectively reduced the parasite load in the brains, livers, and lungs of T. gondii-infected mice, however, a low dose of 1NM-PP1 with oral administration didn't change the survival rates of infected mice.KEY WORDS: 1NM-PP1, bumped kinase inhibitor, drug, protein kinase, Toxoplasma gondii.

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Use of the Kinase Inhibitor Analog 1NM-PP1 Reveals a Role for Toxoplasma gondii CDPK1 in the Invasion Step

Cited by 54 publications

References 25 publications

Serum Albumin Stimulates Protein Kinase G-dependent Microneme Secretion in Toxoplasma gondii

Serum Albumin Stimulates Protein Kinase G-dependent Microneme Secretion in Toxoplasma gondii

Does protein phosphorylation govern host cell entry and egress by the Apicomplexa?

1NM-PP1 Treatment of Mice Infected with Toxoplasma gondii

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