Essential function of the alveolin network in the subpellicular microtubules and conoid assembly in Toxoplasma gondii.

Eloïse Bertiaux,Bohumil Maco,Virginie Hamel,Nicolas Dos Santos Pacheco,Dominique Soldati-Favre,Nicolò Tosetti,Paul Guichard,Lorène Bournonville

doi:10.7554/elife.56635

Abstract

The coccidian subgroup of Apicomplexa possesses an apical complex harboring a conoid, made of unique tubulin polymer fibers. This enigmatic organelle extrudes in extracellular invasive parasites and is associated to the apical polar ring (APR). The APR serves as microtubule-organizing center for the 22 subpellicular microtubules (SPMTs) that are linked to a patchwork of flattened vesicles, via an intricate network composed of alveolins. Here, we capitalize on ultrastructure expansion microscopy (U-ExM) to localize the Toxoplasma gondii Apical Cap protein 9 (AC9) and its partner AC10, identified by BioID, to the alveolin network and intercalated between the SPMTs. Parasites conditionally depleted in AC9 or AC10 replicate normally but are defective in microneme secretion and fail to invade and egress from infected cells. Electron microscopy revealed that the mature parasite mutants are conoidless, while U-ExM highlighted the disorganization of the SPMTs which likely results in the catastrophic loss of APR and conoid.

Highlights

Toxoplasma gondii belongs to the phylum of Apicomplexa that groups numerous parasitic protozoans causing severe diseases in humans and animals
Apical Cap protein 9 (AC9) was modified by CRISPR/Cas9 editing at the endogenous locus to generate a fusion with the mini auxininduced degron and HA-tag at the C-terminus to localize and rapidly downregulate the protein upon addition of auxin (IAA) (Brown et al, 2017; Figure 1—figure supplement 1A)
As previously reported (Chen et al, 2017), AC9 colocalizes with IMC sub-compartment protein 1 (ISP1) at the most apical plate of the inner membrane complex (IMC) termed apical cap, delimited by the peripheral annuli stained by centrin2 (Cen2)

Summary

Introduction

Toxoplasma gondii belongs to the phylum of Apicomplexa that groups numerous parasitic protozoans causing severe diseases in humans and animals. The alveolin network is made of proteins sharing conserved repeat motifs called alveolins, which together with the IMC span the length of the parasite from the apical polar ring (APR) to the basal complex (Mann and Beckers, 2001; Gould et al, 2008; AndersonWhite et al, 2011). The parasite is capped by a single cone-shaped plate called the apical cap. Several proteins have been reported to localize at the apical cap including the IMC sub-compartment protein 1 (ISP1) (Beck et al, 2010) and nine apical cap proteins (called AC1 to AC9) (Chen et al, 2015; Chen et al, 2017).

Methods

Results

Conclusion