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Toxoplasma gondii attachment to host cells is regulated by a calmodulin-like domain protein kinase
H Kieschnick1, T Wakefield, C A Narducci
1Division of Geographic Medicine, Department of Cell Biology, University of Alabama at Birmingham, Birmingham, Alabama 35294, USA.
Abstract:
The role of calcium-dependent protein kinases in the invasion of Toxoplasma gondii into its animal host cells was analyzed. KT5926, an inhibitor of calcium-dependent protein kinases in other systems, is known to block the motility of Toxoplasma tachyzoites and their attachment to host cells. In vivo, KT5926 blocks the phosphorylation of only three parasite proteins, and in parasite extracts only a single KT5926-sensitive protein kinase activity was detected. This activity was calcium-dependent but did not require calmodulin. In a search for calcium-dependent protein kinases in Toxoplasma, two members of the class of calmodulin-like domain protein kinases (CDPKs) were detected. TgCDPK2 was only expressed at the mRNA level in tachyzoites, but no protein was detected. TgCDPK1 protein was expressed in Toxoplasma tachyzoites and cofractionated precisely with the peak of KT5926-sensitive protein kinase activity. TgCDPK1 kinase activity was calcium-dependent but did not require calmodulin or phospholipids. TgCDPK1 was found to be inhibited effectively by KT5926 at concentrations that block parasite attachment to host cells. In vitro, TgCDPK1 phosphorylated three parasite proteins that migrated identical to the three KT5926-sensitive phosphoproteins detected in vivo. Based on these observations, a central role is suggested for TgCDPK1 in regulating Toxoplasma motility and host cell invasion.
Insights
Toxoplasma gondii invasion relies on calcium-dependent protein kinases. TgCDPK1, a specific kinase, regulates parasite motility and host cell attachment, making it a potential therapeutic target.
Area of Science:
- Parasitology
- Molecular Biology
- Cell Biology
Background:
- Toxoplasma gondii invasion into host cells is crucial for parasitic infection.
- Calcium-dependent protein kinases (CDPKs) are implicated in various cellular processes, including motility and invasion.
Purpose of the Study:
- To investigate the role of CDPKs in T. gondii host cell invasion.
- To identify specific CDPKs involved in parasite motility and attachment.
Main Methods:
- Utilized KT5926, a known inhibitor of CDPKs, to study its effects on T. gondii tachyzoites in vivo and in vitro.
- Analyzed protein phosphorylation patterns and kinase activity in parasite extracts.
- Identified and characterized T. gondii CDPKs, specifically TgCDPK1 and TgCDPK2.
- Assessed the calcium dependency and calmodulin independence of TgCDPK1 activity.
Main Results:
- KT5926 inhibited T. gondii tachyzoite motility and attachment, affecting the phosphorylation of three parasite proteins.
- A single, calcium-dependent, calmodulin-independent protein kinase activity sensitive to KT5926 was detected in parasite extracts.
- TgCDPK1 protein was expressed in tachyzoites and exhibited calcium-dependent kinase activity, inhibited by KT5926.
- TgCDPK1 phosphorylated the same three parasite proteins observed in vivo.
Conclusions:
- TgCDPK1 plays a central role in regulating T. gondii motility and host cell invasion.
- TgCDPK1 is a key mediator of parasite attachment and entry into host cells.
- TgCDPK1 represents a potential therapeutic target for controlling toxoplasmosis.