Structure of Toxoplasma gondii coronin, an actin‐binding protein that relocalizes to the posterior pole of invasive parasites and contributes to invasion and egress

Abstract

Coronins are involved in the regulation of actin dynamics in a multifaceted way, participating in cell migration and vesicular trafficking. Apicomplexan parasites, which exhibit an actin-dependent gliding motility that is essential for traversal through tissues, as well as invasion of and egress from host cells, express only a single coronin, whereas higher eukaryotes possess several isoforms. We set out to characterize the 3-D structure, biochemical function, subcellular localization, and genetic ablation of Toxoplasma gondii coronin (TgCOR), to shed light on its biological role. A combination of X-ray crystallography, small-angle scattering of X-rays, and light scattering revealed the atomic structure of the conserved WD40 domain and the dimeric arrangement of the fulllength protein. TgCOR binds to F-actin and increases the rate and extent of actin polymerization. In vivo, TgCOR relocalizes transiently to the posterior pole of motile and invading parasites, independent of actin dynamics, but concomitant to microneme secretory organelle discharge. TgCOR contributes to, but is not essential for, invasion and egress. Taken together, our data point toward a role for TgCOR in stabilizing newly formed, short filaments and F-actin cross-linking, as well as functions linked to endocytosis and recycling of membranes.-Salamun, J., Kallio, J. P., Daher, W., Soldati-Favre, D., Kursula, I. Structure of Toxoplasma gondii coronin, an actin-binding protein that relocalizes to the posterior pole of invasive parasites and contributes to invasion and egress. FASEB J. 28, 4729 -4747 (2014). www.fasebj.orgKey Words: apicomplexan parasite ⅐ gliding motility ⅐ WD40 domain TOXOPLASMA GONDII is an obligate intracellular parasite of medical and veterinary significance that invades almost any nucleated cell from virtually all warmblooded animals. The invasive stages exhibit a substrate-dependent motility, driven by an actin-and myosin-based motor complex machinery termed the glideosome, which is critical for parasite dissemination (1). Gliding motility depends on an intact parasite actin cytoskeleton and requires actin polymerization (2). However, parasite actin filaments have been difficult to visualize in physiological conditions. Typically, apicomplexan actins appear to be maintained mainly in globular form (3). In vitro polymerization assays performed on purified T. gondii actin indicated that the actin filaments are very short (50 -100 nm; ref. 4). The instability of apicomplexan actin filaments seems to be a prerequisite for efficient gliding motility, since stabilization of actin filaments, either by the known stabilizing agent jasplakinolide (JAS; refs. 2, 5, 6) or by substituting key residues assumed to be involved in the intermolecular contacts between different protomers, disrupt the normal gliding motility of parasites (7).Actin dynamics and its regulation in the Apicomplexa differ significantly from higher eukaryotes. The rapid 1 These authors contributed equally to this work.

Myosin-dependent cell-cell communication controls synchronicity of division in acute and chronic stages of Toxoplasma gondii

“…4b). Furthermore, the F-actin-binding protein coronin (TgCOR), previously shown to accumulate to the posterior pole of extracellular parasites on Ca 2+ -ionophore treatment36 still re-localized at the basal pole in MyoJ-KO, indicating that TgMyoJ is not responsible for TgCOR re-localization in motile parasites (Supplementary Fig. 4c).…”

Toxoplasma gondii F-actin forms an extensive filamentous network required for material exchange and parasite maturation

“…We speculate that this network consists of either short F-actin bundles that are cross-linked via unknown actin-binding proteins (based on the formation of short actin filaments in vitro) (Skillman et al, 2011) or that the presence of actin binding proteins such as formin, profilin, and coronin may coordinate their activities in vivo to produce longer actin filaments than those formed in vitro (Skillman et al, 2011; Olshina et al, 2016;Salamun et al, 2014). While this study focuses on the characterization of the membranous network within the PV, it is worth nothing that highly dynamic actin filaments have been also detected within the cytosol of the parasite, and we show that these dynamics are almost completely abolished upon depletion of ADF in good agreement with previous findings (Haase et al, 2015; Mehta and Sibley, 2011) (Figure 6; Videos 5–7).…”

Gliding motility powers invasion and egress in Apicomplexa

“…berghei sporozoites 46 , whereas in T. gondii coronin relocalizes to the posterior pole of the parasite concomitantly to microneme secretion but does not affect motility 47 . Thus, actin polymerization is clearly important for the process of gliding motility in Apicomplexa.…”