scholarly journals Toxoplasma ferlin1 is a versatile and dynamic mediator of microneme trafficking and secretion

2020 ◽  
Author(s):  
Daniel N.A. Tagoe ◽  
Allison A. Drozda ◽  
Isabelle Coppens ◽  
Bradley I. Coleman ◽  
Marc-Jan Gubbels
Keyword(s):  
Host Cell ◽  
Secretory Pathway ◽  
Retrograde Transport ◽  
Dominant Negative ◽  
C2 Domains ◽  
Apicomplexan Parasites ◽  
Obligate Intracellular ◽  
Calcium Dependent ◽  
Vesicle Pool

AbstractCalcium-dependent exocytosis of the microneme organelles that facilitate host cell invasion is critical for obligate intracellular apicomplexan parasites such as Toxoplasma gondii. Ferlins represent a protein family with roles in exocytosis containing multiple Ca2+-sensing C2 domains. Here we defined the role of T. gondii’s ferlin 1 (FER1) in microneme biology. FER1 localized dynamically to several compartments of the parasite’s secretory pathway as well as to an apical spot near the site of microneme secretion. FER1 function was dissected by overexpression of a variety of N-terminally tagged alleles causing dominant negative phenotypes. This demonstrated FER1 traffics microneme organelles at several discrete steps of their natural trajectories: 1. from ELC to the subpellicular microtubules; 2. along the subpellicular microtubules to the apical end; 3. into the conoid; 4. and inferred from observed retrograde transport from the subpellicular microtubules, recycling of micronemes from mother to daughter parasites. Furthermore, full-length FER1 overexpression results in a squirt of microneme release sufficient for host cell egress. This indicates FER1 facilitates fusion of the most apical, radially organized micronemes with the plasma membrane. Moreover, FER1 acts differentially on the Rab5A/C-dependent and - independent microneme sub-populations. Finally, apical FER1 overlaps with the presence of VP1, a pyrophosphatase proton pump. Integrating all new insights, we propose a model of microneme exocytosis wherein the radial micronemes constitute a readily releasable vesicle pool primed by acidification.

scholarly journals Toxoplasma DJ-1 Regulates Organelle Secretion by a Direct Interaction with Calcium-Dependent Protein Kinase 1

mBio ◽  
2017 ◽  
Vol 8 (1) ◽  
Author(s):  
Matthew A. Child ◽  
Megan Garland ◽  
Ian Foe ◽  
Peter Madzelan ◽  
Moritz Treeck ◽  
...  
Keyword(s):  
Host Cell ◽  
Kinase Activity ◽  
Effector Proteins ◽  
Apicomplexan Parasites ◽  
Obligate Intracellular ◽  
Calcium Dependent ◽  
Intracellular Parasites ◽  
Dependent Protein ◽  
Protective Environment ◽  
Calcium Dependent Protein Kinases

ABSTRACT Human DJ-1 is a highly conserved and yet functionally enigmatic protein associated with a heritable form of Parkinson’s disease. It has been suggested to be a redox-dependent regulatory scaffold, binding to proteins to modulate their function. Here we present the X-ray crystal structure of the Toxoplasma orthologue Toxoplasma gondii DJ-1 (TgDJ-1) at 2.1-Å resolution and show that it directly associates with calcium-dependent protein kinase 1 (CDPK1). The TgDJ-1 structure identifies an orthologously conserved arginine dyad that acts as a phospho-gatekeeper motif to control complex formation. We determined that the binding of TgDJ-1 to CDPK1 is sensitive to oxidation and calcium, and that this interaction potentiates CDPK1 kinase activity. Finally, we show that genetic deletion of TgDJ-1 results in upregulation of CDPK1 expression and that disruption of the CDPK1/TgDJ-1 complex in vivo prevents normal exocytosis of parasite virulence-associated organelles called micronemes. Overall, our data suggest that TgDJ-1 functions as a noncanonical kinase-regulatory scaffold that integrates multiple intracellular signals to tune microneme exocytosis in T. gondii. IMPORTANCE Apicomplexan parasites such as Toxoplasma and Plasmodium are obligate intracellular parasites that require the protective environment of a host cell in order to replicate and survive within a host organism. These parasites secrete effector proteins from specialized apical organelles to select and invade a chosen host cell. The secretion of these organelles is a tightly regulated process coordinated by endogenous small molecules and calcium-dependent protein kinases. We previously identified the Toxoplasma orthologue of the highly conserved protein DJ-1 as a regulator of microneme secretion, but the molecular basis for this was not known. We have now identified the molecular mechanism for how TgDJ-1 regulates microneme secretion. TgDJ-1 interacts with the kinase responsible for the secretion of these organelles (calcium-dependent kinase 1) and synergizes with calcium to potentiate kinase activity. This interaction is direct, phosphodependent, and necessary for the normal secretion of these important organelles. IMPORTANCE Apicomplexan parasites such as Toxoplasma and Plasmodium are obligate intracellular parasites that require the protective environment of a host cell in order to replicate and survive within a host organism. These parasites secrete effector proteins from specialized apical organelles to select and invade a chosen host cell. The secretion of these organelles is a tightly regulated process coordinated by endogenous small molecules and calcium-dependent protein kinases. We previously identified the Toxoplasma orthologue of the highly conserved protein DJ-1 as a regulator of microneme secretion, but the molecular basis for this was not known. We have now identified the molecular mechanism for how TgDJ-1 regulates microneme secretion. TgDJ-1 interacts with the kinase responsible for the secretion of these organelles (calcium-dependent kinase 1) and synergizes with calcium to potentiate kinase activity. This interaction is direct, phosphodependent, and necessary for the normal secretion of these important organelles.

scholarly journals Amino Acid Metabolism in Apicomplexan Parasites

Metabolites ◽  
2021 ◽  
Vol 11 (2) ◽  
pp. 61
Author(s):  
Aarti Krishnan ◽  
Dominique Soldati-Favre
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Current Knowledge ◽  
Acid Biosynthesis ◽  
Apicomplexan Parasites ◽  
Obligate Intracellular ◽  
Metabolic Functions ◽  
Transcriptomics Data

Obligate intracellular pathogens have coevolved with their host, leading to clever strategies to access nutrients, to combat the host’s immune response, and to establish a safe niche for intracellular replication. The host, on the other hand, has also developed ways to restrict the replication of invaders by limiting access to nutrients required for pathogen survival. In this review, we describe the recent advancements in both computational methods and high-throughput –omics techniques that have been used to study and interrogate metabolic functions in the context of intracellular parasitism. Specifically, we cover the current knowledge on the presence of amino acid biosynthesis and uptake within the Apicomplexa phylum, focusing on human-infecting pathogens: Toxoplasma gondii and Plasmodium falciparum. Given the complex multi-host lifecycle of these pathogens, we hypothesize that amino acids are made, rather than acquired, depending on the host niche. We summarize the stage specificities of enzymes revealed through transcriptomics data, the relevance of amino acids for parasite pathogenesis in vivo, and the role of their transporters. Targeting one or more of these pathways may lead to a deeper understanding of the specific contributions of biosynthesis versus acquisition of amino acids and to design better intervention strategies against the apicomplexan parasites.

scholarly journals Cytoskeleton of Apicomplexan Parasites

2002 ◽  
Vol 66 (1) ◽  
pp. 21-38 ◽  
Author(s):  
Naomi S. Morrissette ◽  
L. David Sibley
Keyword(s):  
Cytoskeletal Proteins ◽  
Opportunistic Pathogens ◽  
Protozoan Parasites ◽  
Apicomplexan Parasites ◽  
Obligate Intracellular ◽  
Intracellular Parasites ◽  
Eimeria Spp ◽  
Cytoskeletal Elements ◽  
Theileria Spp

SUMMARY The Apicomplexa are a phylum of diverse obligate intracellular parasites including Plasmodium spp., the cause of malaria; Toxoplasma gondii and Cryptosporidium parvum, opportunistic pathogens of immunocompromised individuals; and Eimeria spp. and Theileria spp., parasites of considerable agricultural importance. These protozoan parasites share distinctive morphological features, cytoskeletal organization, and modes of replication, motility, and invasion. This review summarizes our current understanding of the cytoskeletal elements, the properties of cytoskeletal proteins, and the role of the cytoskeleton in polarity, motility, invasion, and replication. We discuss the unusual properties of actin and myosin in the Apicomplexa, the highly stereotyped microtubule populations in apicomplexans, and a network of recently discovered novel intermediate filament-like elements in these parasites.

scholarly journals Structural insights into an atypical secretory pathway kinase crucial for Toxoplasma gondii invasion

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Gaëlle Lentini ◽  
Rouaa Ben Chaabene ◽  
Oscar Vadas ◽  
Chandra Ramakrishnan ◽  
Budhaditya Mukherjee ◽  
...  
Keyword(s):  
Toxoplasma Gondii ◽  
Host Cell ◽  
Cell Invasion ◽  
Plasma Membranes ◽  
Secretory Pathway ◽  
Host Cell Invasion ◽  
Host Cell Membrane ◽  
Apicomplexan Parasites ◽  
Cell Plasma ◽  
Structural Insights

AbstractActive host cell invasion by the obligate intracellular apicomplexan parasites relies on the formation of a moving junction, which connects parasite and host cell plasma membranes during entry. Invading Toxoplasma gondii tachyzoites secrete their rhoptry content and insert a complex of RON proteins on the cytoplasmic side of the host cell membrane providing an anchor to which the parasite tethers. Here we show that a rhoptry-resident kinase RON13 is a key virulence factor that plays a crucial role in host cell entry. Cryo-EM, kinase assays, phosphoproteomics and cellular analyses reveal that RON13 is a secretory pathway kinase of atypical structure that phosphorylates rhoptry proteins including the components of the RON complex. Ultimately, RON13 kinase activity controls host cell invasion by anchoring the moving junction at the parasite-host cell interface.

scholarly journals The transmembrane protein p23 contributes to the organization of the Golgi apparatus

2000 ◽  
Vol 113 (6) ◽  
pp. 1043-1057 ◽  
Author(s):  
M. Rojo ◽  
G. Emery ◽  
V. Marjomaki ◽  
A.W. McDowall ◽  
R.G. Parton ◽  
...  
Keyword(s):  
Golgi Apparatus ◽  
Secretory Pathway ◽  
Transmembrane Protein ◽  
Ectopic Expression ◽  
Retrograde Transport ◽  
Early Secretory Pathway ◽  
Golgi Membranes ◽  
Constant Diameter ◽  
Golgi Network

In previous studies we have shown that p23, a member of the p24-family of small transmembrane proteins, is highly abundant in membranes of the cis-Golgi network (CGN), and is involved in sorting/trafficking in the early secretory pathway. In the present study, we have further investigated the role of p23 after ectopic expression. We found that ectopically expressed p23 folded and oligomerized properly, even after overexpression. However, in contrast to endogenous p23, exogenous p23 molecules did not localize to the CGN, but induced a significant expansion of characteristic smooth ER membranes, where they accumulated in high amounts. This ER-derived, p23-rich subdomain displayed a highly regular morphology, consisting of tubules and/or cisternae of constant diameter, which were reminiscent of the CGN membranes containing p23 in control cells. The expression of exogenous p23 also led to the specific relocalization of endogenous p23, but not of other proteins, to these specialized ER-derived membranes. Relocalization of p23 modified the ultrastructure of the CGN and Golgi membranes, but did not affect anterograde and retrograde transport reactions to any significant extent. We conclude (i) that p23 has a morphogenic activity that contributes to the morphology of CGN-membranes; and (ii) that the presence of p23 in the CGN is necessary for the proper organization of the Golgi apparatus.

scholarly journals Role of the early secretory pathway in SARS-CoV-2 infection

2020 ◽  
Vol 219 (9) ◽  
Author(s):  
Daria Sicari ◽  
Aristotelis Chatziioannou ◽  
Theodoros Koutsandreas ◽  
Roberto Sitia ◽  
Eric Chevet
Keyword(s):  
Host Cell ◽  
Secretory Pathway ◽  
Tissue Specificity ◽  
Molecular Machines ◽  
Transport Proteins ◽  
Host Cells ◽  
Early Secretory Pathway ◽  
Antiviral Targets ◽  
Exocytic Pathway

Similar to other RNA viruses, SARS-CoV-2 must (1) enter a target/host cell, (2) reprogram it to ensure its replication, (3) exit the host cell, and (4) repeat this cycle for exponential growth. During the exit step, the virus hijacks the sophisticated machineries that host cells employ to correctly fold, assemble, and transport proteins along the exocytic pathway. Therefore, secretory pathway–mediated assemblage and excretion of infective particles represent appealing targets to reduce the efficacy of virus biogenesis, if not to block it completely. Here, we analyze and discuss the contribution of the molecular machines operating in the early secretory pathway in the biogenesis of SARS-CoV-2 and their relevance for potential antiviral targeting. The fact that these molecular machines are conserved throughout evolution, together with the redundancy and tissue specificity of their components, provides opportunities in the search for unique proteins essential for SARS-CoV-2 biology that could also be targeted with therapeutic objectives. Finally, we provide an overview of recent evidence implicating proteins of the early secretory pathway as potential antiviral targets with effective therapeutic applications.

Redox diversity in ERAD-mediated protein retrotranslocation from the endoplasmic reticulum: a complex puzzle

2015 ◽  
Vol 396 (5) ◽  
pp. 539-554 ◽  
Author(s):  
Yutaka Suzuki ◽  
Manfred J. Schmitt
Keyword(s):  
Endoplasmic Reticulum ◽  
Host Cell ◽  
Secretory Pathway ◽  
Redox Status ◽  
Current View ◽  
Cell Penetration ◽  
Protein Toxins ◽  
Underlying Mechanisms ◽  
Disulfide Isomerization

Abstract Misfolded and incorrectly assembled proteins in the secretory pathway are eliminated by ubiquitylation and proteasomal degradation in a process known as ER-associated degradation (ERAD). Retrotranslocation of diverse substrates including misfolded proteins and viruses occurs through channels in the ER membrane, which are also utilized for host cell penetration by A/B class protein toxins such as cholera toxin, ricin or K28. According to the current view, disulfide-bonded proteins must either be reduced or rearranged to ensure translocation competence and entry into the cytosol from the ER. As the underlying mechanisms are still largely mysterious, we here focus on the redox status and disulfide isomerization of ERAD substrates and the role of oxidoreductases in the essential process of ER-to-cytosol retrotranslocation.

scholarly journals δ-COP modulates Aβ peptide formation via retrograde trafficking of APP

2016 ◽  
Vol 113 (19) ◽  
pp. 5412-5417 ◽  
Author(s):  
Karima Bettayeb ◽  
Jerry C. Chang ◽  
Wenjie Luo ◽  
Suvekshya Aryal ◽  
Dante Varotsis ◽  
...  
Keyword(s):  
Secretory Pathway ◽  
Amyloid Β ◽  
Retrograde Transport ◽  
Surface Expression ◽  
Cell Surface Expression ◽  
Aβ Peptide ◽  
Early Secretory Pathway ◽  
Peptide Formation ◽  
App Metabolism

The components involved in cellular trafficking and protein recycling machinery that have been associated with increased Alzheimer’s disease (AD) risk belong to the late secretory compartments for the most part. Here, we hypothesize that these late unavoidable events might be the consequence of earlier complications occurring while amyloid precursor protein (APP) is trafficking through the early secretory pathway. We investigated the relevance to AD of coat protein complex I (COPI)-dependent trafficking, an early step in Golgi-to-endoplasmic reticulum (ER) retrograde transport and one of the very first trafficking steps. Using a complex set of imaging technologies, including inverse fluorescence recovery after photobleaching (iFRAP) and photoactivatable probes, coupled to biochemical experiments, we show that COPI subunit δ (δ-COP) affects the biology of APP, including its subcellular localization and cell surface expression, its trafficking, and its metabolism. These findings demonstrate the crucial role of δ-COP in APP metabolism and, consequently, the generation of amyloid-β (Aβ) peptide, providing previously nondescribed mechanistic explanations of the underlying events.

scholarly journals ToxoplasmaRhoptries: Unique Secretory Organelles and Source of Promising Vaccine Proteins for Immunoprevention of Toxoplasmosis

2008 ◽  
Vol 2008 ◽  
pp. 1-7 ◽  
Author(s):  
Henryka Dlugonska
Keyword(s):  
Neospora Caninum ◽  
Parasitophorous Vacuole ◽  
Protozoan Parasite ◽  
Current Data ◽  
Intracellular Killing ◽  
Apicomplexan Parasites ◽  
Obligate Intracellular ◽  
Animal Pathogens ◽  
Intracellular Protozoan Parasite

Toxoplasma gondiiis an obligate intracellular protozoan parasite classified in the phylum Apicomplexa, which includes numerous notable human and animal pathogens (Plasmodiumspecies,Cryptosporidiumspecies,Neospora caninum, etc.). The invasive stages of apicomplexans are characterized by the presence of an apical complex composed of specialized cytoskeletal and secretory organelles, including rhoptries. Rhoptries, unique apical secretory organelles shared exclusively by all apicomplexan parasites, are known to be involved in an active parasite's penetration into the host cell associated with the biogenesis of specific intracellular compartment, parasitophorous vacuole in which the parasite multiplies intensively, avoiding intracellular killing. Due to the key biological role of rhoptries, rhoptry proteins have recently become vaccine candidates for the prevention of several parasitoses, toxoplasmosis among them. The article presents current data onT. gondiirhoptries biology and new approaches to the development of effective vaccines against toxoplasmosis using rhoptry antigens.

scholarly journals A central role of glutamine in Chlamydia infection

2019 ◽  
Author(s):  
Karthika Rajeeve ◽  
Nadine Vollmuth ◽  
Sudha Janaki-Raman ◽  
Thomas Wulff ◽  
Maximilian Schmalhofer ◽  
...  
Keyword(s):  
Host Cell ◽  
Metabolic Reprogramming ◽  
Glutamine Metabolism ◽  
Chlamydia Infection ◽  
Developmental Cycle ◽  
Dependent Manner ◽  
Obligate Intracellular ◽  
A Cell ◽  
Obligate Intracellular Bacteria

AbstractObligate intracellular bacteria like Chlamydia trachomatis undergo a complex developmental cycle between infectious non-replicative (EBs) and non-infectious replicative (RBs) forms. EBs shortly after entering a host cell transform to RBs, a crucial process in infection, initiating chlamydial replication. As Chlamydia fail to replicate outside the host cell it is currently unknown how the transition from EBs to RBs is initiated. Here we show in a cell-free approach in axenic media that uptake of glutamine by the bacteria is critical to initiate EB-RB transition. These bacteria utilize glutamine to synthesize cell wall peptidoglycan which has recently been detected in the septa of replicating intracellular Chlamydia. The increased requirement for glutamine in infected cells is achieved by reprogramming the glutamine metabolism in a c-Myc-dependent manner. Glutamine was effectively taken up by the glutamine transporter SLC1A5 and metabolized via glutaminase. Interference with this metabolic reprogramming limited growth of Chlamydia. Intriguingly, Chlamydia failed to produce progeny in SLC1A5 knockout mice. Thus, we report on the central role of glutamine for the development of an obligate intracellular pathogenic bacterium and the reprogramming of host glutamine metabolism, which may provide a basis for innovative anti-infective strategies.

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