ROP18 Is a Rhoptry Kinase Controlling the Intracellular Proliferation of Toxoplasma gondii

Hiba El Hajj,Stefan T Arold,Jean François Dubremetz,Gilles Labesse,Henri Vial,Maryse Lebrun,Kasturi Haldar

doi:10.1371/journal.ppat.0030014

Hiba El Hajj, Stefan T Arold + Show 5 more

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https://doi.org/10.1371/journal.ppat.0030014

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Abstract

Toxoplasma gondii is an obligate intracellular parasite for which the discharge of apical organelles named rhoptries is a key event in host cell invasion. Among rhoptry proteins, ROP2, which is the prototype of a large protein family, is translocated in the parasitophorous vacuole membrane during invasion. The ROP2 family members are related to protein-kinases, but only some of them are predicted to be catalytically active, and none of the latter has been characterized so far. We show here that ROP18, a member of the ROP2 family, is located in the rhoptries and re-localises at the parasitophorous vacuole membrane during invasion. We demonstrate that a recombinant ROP18 catalytic domain (amino acids 243–539) possesses a protein-kinase activity and phosphorylate parasitic substrates, especially a 70-kDa protein of tachyzoites. Furthermore, we show that overexpression of ROP18 in transgenic parasites causes a dramatic increase in intra-vacuolar parasite multiplication rate, which is correlated with kinase activity. Therefore, we demonstrate, to our knowledge for the first time, that rhoptries can discharge active protein-kinases upon host cell invasion, which can exert a long-lasting effect on intracellular parasite development and virulence.

Highlights

Toxoplasma gondii is an obligate intracellular parasite belonging to the protozoan phylum Apicomplexa, which includes a large number of human and animal parasites responsible for diseases such as malaria, toxoplasmosis, coccidiosis, and cryptosporidiosis
We have recently shown that the ROP2 family could be expanded to at least 12 members, some of which show a full set of features compatible with protein-kinase activity, whereas ROP2 and its closest relatives have lost some of these features [12]
We have studied a novel rhoptry protein dubbed ROP18, which is translocated to the parasitophorous vacuole (PV) membrane upon invasion

Summary

Introduction

Toxoplasma gondii is an obligate intracellular parasite belonging to the protozoan phylum Apicomplexa, which includes a large number of human and animal parasites responsible for diseases such as malaria, toxoplasmosis, coccidiosis, and cryptosporidiosis. As for all other members of the phylum, host cell invasion by T. gondii involves specialized apical organelles of the invasive stage, namely micronemes and rhoptries, which discharge their contents successively [1,2]. A complex of microneme and rhoptry neck proteins forms a moving junction with the host cell plasma membrane that propels the parasite within the developing parasitophorous vacuole [7,8]. Proteins of the bulb of the rhoptries (ROP proteins) become associated with the parasitophorous vacuole membrane (PVM) that forms from host plasma membrane and rhoptry components during invasion [9]. Targeted depletion of ROP2 using a ribozyme-modified antisense RNA strategy results in disruption of rhoptry biogenesis and affects cytokinesis, association of host cell mitochondria with the PVM, host cell invasion, and virulence in mice [15]. Several other members of the family have been characterized more recently, and they are targeted to the PVM upon invasion [16,17,18]

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