A Novel Family of Toxoplasma IMC Proteins Displays a Hierarchical Organization and Functions in Coordinating Parasite Division

Josh R Beck,Vern B Carruthers,Naomi S Morrissette,My-Hang Huynh,Jessica Cruz De Leon,Peter J Bradley,Imilce A Rodriguez-Fernandez,Dominique Soldati-Favre

doi:10.1371/journal.ppat.1001094

Abstract

Apicomplexans employ a peripheral membrane system called the inner membrane complex (IMC) for critical processes such as host cell invasion and daughter cell formation. We have identified a family of proteins that define novel sub-compartments of the Toxoplasma gondii IMC. These IMC Sub-compartment Proteins, ISP1, 2 and 3, are conserved throughout the Apicomplexa, but do not appear to be present outside the phylum. ISP1 localizes to the apical cap portion of the IMC, while ISP2 localizes to a central IMC region and ISP3 localizes to a central plus basal region of the complex. Targeting of all three ISPs is dependent upon N-terminal residues predicted for coordinated myristoylation and palmitoylation. Surprisingly, we show that disruption of ISP1 results in a dramatic relocalization of ISP2 and ISP3 to the apical cap. Although the N-terminal region of ISP1 is necessary and sufficient for apical cap targeting, exclusion of other family members requires the remaining C-terminal region of the protein. This gate-keeping function of ISP1 reveals an unprecedented mechanism of interactive and hierarchical targeting of proteins to establish these unique sub-compartments in the Toxoplasma IMC. Finally, we show that loss of ISP2 results in severe defects in daughter cell formation during endodyogeny, indicating a role for the ISP proteins in coordinating this unique process of Toxoplasma replication.

Highlights

The phylum Apicomplexa contains numerous obligate intracellular pathogens that are the cause of serious disease in humans and animals, greatly influencing global health and causing significant economic loss worldwide
Apicomplexans are the cause of important diseases in humans and animals including malaria (Plasmodium falciparum), which claims over a million human lives each year, and toxoplasmosis (Toxoplasma gondii), which causes birth defects and neurological disorders
Residues predicted for myristoylation and palmitoylation are critical in the membrane targeting of these proteins, suggesting that multiple palmitoyl acyltransferase activities reside within the inner membrane complex (IMC) and dictate its organization

Summary

Introduction

The phylum Apicomplexa contains numerous obligate intracellular pathogens that are the cause of serious disease in humans and animals, greatly influencing global health and causing significant economic loss worldwide. Apicomplexans are grouped with dinoflagellates and ciliates in the alveolata infrakingdom [5]. Molecular phylogenetic data supports this grouping, as does the identification of a conserved family of articulin-like membrane skeleton proteins, the alveolins, which associate with alveoli in all three phyla [6,7]. While the presence of alveoli is conserved, each of these groups has adapted this peripheral membrane structure for different cellular functions to fit their distinct niches. The alveoli sometimes contain cellulose-based plates that function as protective armor [8]. Ciliate alveoli are calcium storage devices thought to play roles in regulation of cilia, exocytosis from cortical organelles known as extrusomes, and control of cytoskeletal elements [9,10,11]

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