Stability and function of a putative microtubule-organizing center in the human parasite<i>Toxoplasma gondii</i>

Leung, Jacqueline M.; He, Yudou; Zhang, Fangliang; Hwang, Yu Chen; Nagayasu, Eiji; Li, Jun; Murray, John M.; Hu, Ke

doi:10.1091/mbc.e17-01-0045

Cited by 63 publications

(97 citation statements)

References 61 publications

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“…In deoxycholate extracted AC9 and AC10 depleted parasites, the SPMTs are formed properly but are not joint together by the APR, which is absent in these mutants. A similar but milder phenotype was also reported in the double knock-out of kinesin A and APR1 with SPMTs partially detached from the parasite apex because of the fragmentation of the APR (26). The strategic position of AC9 and AC10 intercalated between SPTMs suggested a possible implication in maintaining the correct interspacing between the SPMTs.…”

Section: Discussionsupporting

confidence: 63%

“…The conoid markers analyzed so far are incorporated in the daughter cells since the beginning of the division process, shortly after centrosome duplication. Next, we assessed the fate of APR markers such as KinA and APR1, which are incorporated very early (26) and RNG1 a late marker of division (37). In the absence of AC9 or AC10, APR1 and KinA are lost in mature parasites, but present in daughter cells, respectively (Fig 6D and 6E).…”

Section: Resultsmentioning

confidence: 99%

“…DCX was later shown to bundle the conoidal tubulin fibers into comma-shape structures likely conferring the cone-shaped configuration of the conoid (25). Moreover, the double knockout of kinesin A and apical polar ring protein 1 (APR1) caused a fragmentation of the APR with a conoid frequently partially detached resulting in impairment of microneme secretion (26). Lastly, conoid protein hub 1 (CPH1) is a conserved apicomplexan protein essential for parasite motility, invasion and egress that regulates conoid stability in extracellular parasites without impacting on microneme secretion (27).…”

Section: Introductionmentioning

confidence: 99%

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Two alveolin network proteins are essential for the subpellicular microtubules assembly and conoid anchoring to the apical pole of matureToxoplasma gondii

Tosetti

Pacheco

Bertiaux

et al. 2020

Preprint

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Toxoplasma gondii belongs to the coccidian sub-group of Apicomplexa that possess an apical complex harboring a conoid, made of unique tubulin polymer fibers. This enigmatic and dynamic organelle extrudes in extracellular invasive parasites and is associated to the apical polar ring (APR), a microtubule-organizing center for the 22 subpellicular microtubules (SPMTs). The SPMTs are linked to the Inner Membrane Complex (IMC), a patchwork of flattened vesicles, via an intricate network of small filaments composed of alveolins proteins. Here, we capitalize on super-resolution techniques including stimulated emission depletion (STED) microscopy and ultrastructure expansion microscopy (U-ExM) to localize the Apical Cap protein 9 (AC9) and its close partner AC10, identified by BioID, to the alveolin network and intercalated between the SPMTs. Conditional depletion of AC9 or AC10 using the Auxin-induced Degron (AiD) system uncovered a severe loss of fitness. Parasites lacking AC9 or AC10 replicate normally but are defective in microneme secretion and hence fail to invade and egress from infected cells. Remarkably, a series of crucial apical complex proteins (MyoH, AKMT, FRM1, CPH1, ICMAP1 and RNG2) are lost in the mature parasites although they are still present in the forming daughter cells. Electron microscopy on intracellular or deoxycholate-extracted parasites revealed that the mature parasite mutants are conoidless. Closer examination of the SPMTs by U-ExM highlighted the disassembly of the SPMTs in the apical cap region that is presumably at the origin of the catastrophic loss of APR and conoid. AC9 and AC10 are two critical components of the alveolin network that ensure the integrity of the whole apical complex in T. gondii and likely other coccidians.

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Section: Discussionsupporting

confidence: 63%

Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Two alveolin network proteins are essential for the subpellicular microtubules assembly and conoid anchoring to the apical pole of matureToxoplasma gondii

Tosetti

Pacheco

Bertiaux

et al. 2020

Preprint

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“…In apicomplexan parasites, the cilium was likely adapted to form the organizing core of the "apical complex" of cytoskeletal structures and secretory organelles for which the phylum is named (Figure 1; de Leon et al, 2013). The protein components of the apical complex include orthologs of cilium-associated proteins including centrins (Hu et al, 2006;Lentini et al, 2019), a SAS6 cartwheel protein (de Leon et al, 2013;Lévêque et al, 2016), an associated rootlet fiber (Francia et al, 2012), and a distributed microtubule-organizing center called the apical polar ring (Leung et al, 2017). During the asexual stage, Apicomplexa such as Toxoplasma have a specialized cilium-like structure called the conoid, and typical motile flagella grow at this site during the sexual stage (Francia et al, 2015).…”

Section: Introductionmentioning

confidence: 99%

Loss of a conserved MAPK causes catastrophic failure in assembly of a specialized cilium-like structure inToxoplasma gondii

O’Shaughnessy

Beraki

et al. 2020

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TOC: Toxoplasma gondii that lack the kinase ERK7 cannot invade or egress from their host cells, thereby blocking their replicative cycle. These defects are due to the loss of a specialized cilium-like structure called the conoid. Strikingly, the ultrastructural changes are specific to the conoid, and suggest an important role for ERK7 in its biogenesis. AbstractPrimary cilia are important organizing centers that control diverse cellular processes. Apicomplexan parasites like Toxoplasma gondii have a specialized cilium-like structure called the conoid that organizes the secretory and invasion machinery critical for the parasites' lifestyle. The proteins that initiate the biogenesis of this structure are largely unknown. We identified the Toxoplasma ortholog of the conserved kinase ERK7 as essential to conoid assembly. Parasites in which ERK7 has been depleted lose their conoids late during maturation and are immotile and thus unable to invade new host cells. This is the most severe phenotype to conoid biogenesis yet reported, and is made more striking by the fact that ERK7 is not a conoid protein, as it localizes just basal to the structure. ERK7 has been recently implicated in ciliogenesis in metazoan cells, and our data suggest that this kinase has an ancient and central role in regulating ciliogenesis throughout Eukaryota.

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“…Interestingly, such apically polarized localization of Rab11A was also evident during extracellular parasite adhesion and motility. Thus, one may envision that components of the apical complex, a microtubule-rich structure from which emanates the subpellicular microtubules [41], may control Rab11A-dependent recruitment and exocytosis of specific cargos at the apical pole of the parasite. In particular, RING2, a component of the apical polar ring, was shown to function in constitutive and cGMP-stimulated secretion of microneme proteins [42].…”

Section: Discussionmentioning

confidence: 99%

Rab11A regulates the constitutive secretory pathway duringToxoplasma gondiiinvasion of host cells and parasite replication

Venugopal

Chehade

Werkmeister

et al. 2019

Preprint

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22 Summary23 Toxoplasma gondii possesses an armada of secreted virulent factors that enable 24 parasite invasion and survival into host cells. These factors are contained in specific 25 secretory organelles, the rhoptries, micronemes and dense granules that release their 26 content upon host cell recognition. Dense granules are secreted in a constitutive 27 manner during parasite replication and play a crucial role in modulating host 28 metabolic and immune responses. While the molecular mechanisms triggering rhoptry 29 and microneme release upon host cell adhesion have been well studied, constitutive 30 secretion remains a poorly explored aspect of T. gondii vesicular trafficking. Here, we 31 investigated the role of the small GTPase Rab11A, a known regulator of exocytosis in 32 eukaryotic cells. Our data revealed an essential role of Rab11A in promoting the 33 cytoskeleton driven transport of DG and the release of their content into the vacuolar 34 space. Rab11A also regulates transmembrane protein trafficking and localization 2 35 during parasite replication, indicating a broader role of Rab11A in cargo exocytosis at 36 the plasma membrane. Moreover, we found that Rab11A also regulates extracellular 37 parasite motility and adhesion to host cells. In line with these findings, MIC238 secretion was altered in Rab11A-defective parasites, which also exhibited severe 39 morphological defects. Strikingly, by live imaging we observed a polarized 40 accumulation of Rab11A-positive vesicles and dense granules at the apical pole of 41 extracellular motile parasites suggesting that a Rab11A-dependent apically polarized 42 transport of cargo regulates parasite motility. 43 44 45 Introduction 46 Toxoplasma gondii (T. gondii) is an obligatory intracellular parasite that belongs to 47 the phylum Apicomplexa, typified by the presence of specific apical secretory 48 organelles, the rhoptries and the micronemes. Upon contact with the host cell, rhoptry 49 (ROP) and microneme (MIC) proteins are released and promote parasite entry by 50 driving the formation of a tight parasite-host cell adhesive membrane structure (called 51 the circular junction) [1]. ROP proteins also contribute to building the 52 parasitophorous vacuole (PV), within which the parasite rapidly replicates. The 53 molecular mechanisms regulating MIC exocytosis have been well studied leading to 54 the discovery of specific parasite signaling pathways triggering their secretion upon 55 parasite adhesion to host cells [2] [3]. Dense granules (DG) are also parasite secretory 56 organelles essential for parasite survival, which release effectors modulating host 57 immune and metabolic responses [4]. Dense granule proteins (GRA) also promote the 58 formation of an intravacuolar nanotubular network (IVN), which interconnects 59 parasites during intracellular replication, thereby ensuring the synchronicity of the 60 successive divisions [5] [6] [7]. The IVN also connects the parasite to the PV 61 membrane (PVM), presumably enhancing parasite exchanges with its host, not...

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Stability and function of a putative microtubule-organizing center in the human parasiteToxoplasma gondii

Cited by 63 publications

References 61 publications

Two alveolin network proteins are essential for the subpellicular microtubules assembly and conoid anchoring to the apical pole of matureToxoplasma gondii

Two alveolin network proteins are essential for the subpellicular microtubules assembly and conoid anchoring to the apical pole of matureToxoplasma gondii

Loss of a conserved MAPK causes catastrophic failure in assembly of a specialized cilium-like structure inToxoplasma gondii

Rab11A regulates the constitutive secretory pathway duringToxoplasma gondiiinvasion of host cells and parasite replication

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