scholarly journals Apicomplexan F-actin is required for efficient nuclear entry during host cell invasion

2019 ◽  
Author(s):  
Mario Del Rosario ◽  
Javier Periz ◽  
Georgios Pavlou ◽  
Oliver Lyth ◽  
Fernanda Latorre-Barragan ◽  
...  
Keyword(s):  
Host Cell ◽  
Cell Invasion ◽  
Traction Force ◽  
Super Resolution ◽  
Host Cells ◽  
Host Cell Invasion ◽  
Nuclear Entry ◽  
Apicomplexan Parasites ◽  
Intracellular Parasites ◽  
Pass Through

AbstractThe obligate intracellular parasites Toxoplasma gondii and Plasmodium spp. invade host cells by injecting a protein complex into the membrane of the targeted cell that bridges the two cells through the assembly of a ring-like junction. This circular junction stretches while the parasites applies a traction force to pass through; a step that typically concurs with transient constriction of the parasite body. Here we show that the junction can oppose resistance to the passage of the parasite’s nucleus. Super-resolution microscopy and real time imaging highlighted an F-actin pool at the apex of pre-invading parasite, an F-actin ring at the junction area during invasion but also networks of perinuclear and posteriorly localized F-actin. Mutant parasites with dysfunctional acto-myosin showed significant decrease of junctional and perinuclear F-actin and are coincidently affected in nuclear passage through the junction. We propose that the F-actin machinery eases nuclear passage by stabilising the junction and pushing the nucleus through the constriction, providing first evidence for a dual contribution of actin-forces during host cell invasion by apicomplexan parasites.

scholarly journals Not a Simple Tether: Binding of Toxoplasma gondii AMA1 to RON2 during Invasion Protects AMA1 from Rhomboid-Mediated Cleavage and Leads to Dephosphorylation of Its Cytosolic Tail

mBio ◽  
2016 ◽  
Vol 7 (5) ◽  
Author(s):  
Shruthi Krishnamurthy ◽  
Bin Deng ◽  
Roxana del Rio ◽  
Kerry R. Buchholz ◽  
Moritz Treeck ◽  
...  
Keyword(s):  
Toxoplasma Gondii ◽  
Host Cell ◽  
Cell Invasion ◽  
Transmembrane Domain ◽  
Critical Role ◽  
Host Cells ◽  
Receptor Protein ◽  
Host Cell Invasion ◽  
Cell Binding ◽  
Apicomplexan Parasites

ABSTRACT Apical membrane antigen 1 (AMA1) is a receptor protein on the surface of Toxoplasma gondii that plays a critical role in host cell invasion. The ligand to which T . gondii AMA1 (TgAMA1) binds, TgRON2, is secreted into the host cell membrane by the parasite during the early stages of invasion. The TgAMA1-TgRON2 complex forms the core of the “moving junction,” a ring-shaped zone of tight contact between the parasite and host cell membranes, through which the parasite pushes itself during invasion. Paradoxically, the parasite also expresses rhomboid proteases that constitutively cleave the TgAMA1 transmembrane domain. How can TgAMA1 function effectively in host cell binding if its extracellular domain is constantly shed from the parasite surface? We show here that when TgAMA1 binds the domain 3 (D3) peptide of TgRON2, its susceptibility to cleavage by rhomboid protease(s) is greatly reduced. This likely serves to maintain parasite-host cell binding at the moving junction, a hypothesis supported by data showing that parasites expressing a hypercleavable version of TgAMA1 invade less efficiently than wild-type parasites do. Treatment of parasites with the D3 peptide was also found to reduce phosphorylation of S527 on the cytoplasmic tail of TgAMA1, and parasites expressing a phosphomimetic S527D allele of TgAMA1 showed an invasion defect. Taken together, these data suggest that TgAMA1-TgRON2 interaction at the moving junction protects TgAMA1 molecules that are actively engaged in host cell penetration from rhomboid-mediated cleavage and generates an outside-in signal that leads to dephosphorylation of the TgAMA1 cytosolic tail. Both of these effects are required for maximally efficient host cell invasion. IMPORTANCE Nearly one-third of the world’s population is infected with the protozoan parasite Toxoplasma gondii , which causes life-threatening disease in neonates and immunocompromised individuals. T. gondii is a member of the phylum Apicomplexa, which includes many other parasites of veterinary and medical importance, such as those that cause coccidiosis, babesiosis, and malaria. Apicomplexan parasites grow within their hosts through repeated cycles of host cell invasion, parasite replication, and host cell lysis. Parasites that cannot invade host cells cannot survive or cause disease. AMA1 is a highly conserved protein on the surface of apicomplexan parasites that is known to be important for invasion, and the work presented here reveals new and unexpected insights into AMA1 function. A more complete understanding of the role of AMA1 in invasion may ultimately contribute to the development of new chemotherapeutics designed to disrupt AMA1 function and invasion-related signaling in this important group of human pathogens.

High Resolution Low Voltage Field Emission Scanning Electron Microscopy on De-Embedded Thick Sections Reveals Structural Interactions of the Surface of the Apicomplexan Parasite Toxoplasma Gondii with the Host Cell in the Parasitophorous Vacuole

2000 ◽  
Vol 6 (S2) ◽  
pp. 648-649
Author(s):  
Heide Schatten ◽  
David Sibley ◽  
Hans Ris
Keyword(s):  
Toxoplasma Gondii ◽  
Host Cell ◽  
Cell Invasion ◽  
Biochemical Characterization ◽  
Low Voltage ◽  
Cytoskeletal Proteins ◽  
Host Cells ◽  
Farm Animals ◽  
Host Cell Invasion ◽  
Apicomplexan Parasites

The protozoan parasite Toxoplasma gondii represents a large group of Apicomplexan parasites with a highly unusual motility system that is crucial for cell locomotion and host cell invasion. Studying the motility system and interactions with host cells will increase our knowledge on how to prevent infection. Apicomplexan parasites can cause considerable health problems to animals including sheep, goats, pigs, and chicken. Eimeria is known as a pathogen of coccidiosis in chicken, and Cryptosporidium causes cryptospiridiosis in cattle and other farm animals. Toxoplasma gondii can cause infection that results in abortion, central nervous disorders and death of stressed and immunocompromised farm animals. Common to all Apicomplexan parasites is an unconventional motility system that is thought to be actively involved in host-cell invasion. However, the structural and biochemical characterization of Apicomplexan parasites has proven more difficult than previously thought because of genetic and behavioral differences of the parasites’ cytoskeletal proteins that differ from those in well studied mammalian systems.

scholarly journals Apical Organelle Discharge by Cryptosporidium parvum Is Temperature, Cytoskeleton, and Intracellular Calcium Dependent and Required for Host Cell Invasion

2004 ◽  
Vol 72 (12) ◽  
pp. 6806-6816 ◽  
Author(s):  
Xian-Ming Chen ◽  
Steven P. O'Hara ◽  
Bing Q. Huang ◽  
Jeremy B. Nelson ◽  
Jim Jung-Ching Lin ◽  
...  
Keyword(s):  
Intracellular Calcium ◽  
Host Cell ◽  
Cryptosporidium Parvum ◽  
Cell Invasion ◽  
Cytochalasin D ◽  
Gliding Motility ◽  
Host Cells ◽  
Ionophore A23187 ◽  
Host Cell Invasion ◽  
Apicomplexan Parasites

ABSTRACT The apical organelles in apicomplexan parasites are characteristic secretory vesicles containing complex mixtures of molecules. While apical organelle discharge has been demonstrated to be involved in the cellular invasion of some apicomplexan parasites, including Toxoplasma gondii and Plasmodium spp., the mechanisms of apical organelle discharge by Cryptosporidium parvum sporozoites and its role in host cell invasion are unclear. Here we show that the discharge of C. parvum apical organelles occurs in a temperature-dependent fashion. The inhibition of parasite actin and tubulin polymerization by cytochalasin D and colchicines, respectively, inhibited parasite apical organelle discharge. Chelation of the parasite's intracellular calcium also inhibited apical organelle discharge, and this process was partially reversed by raising the intracellular calcium concentration by use of the ionophore A23187. The inhibition of parasite cytoskeleton polymerization by cytochalasin D and colchicine and the depletion of intracellular calcium also decreased the gliding motility of C. parvum sporozoites. Importantly, the inhibition of apical organelle discharge by C. parvum sporozoites blocked parasite invasion of, but not attachment to, host cells (i.e., cultured human cholangiocytes). Moreover, the translocation of a parasite protein, CP2, to the host cell membrane at the region of the host cell-parasite interface was detected; an antibody to CP2 decreased the C. parvum invasion of cholangiocytes. These data demonstrate that the discharge of C. parvum sporozoite apical organelle contents occurs and that it is temperature, intracellular calcium, and cytoskeleton dependent and required for host cell invasion, confirming that apical organelles play a central role in C. parvum entry into host cells.

scholarly journals A member of the ferlin calcium sensor family is essential for Toxoplasma gondii rhoptry secretion

2018 ◽  
Author(s):  
Bradley I. Coleman ◽  
Sudeshna Saha ◽  
Seiko Sato ◽  
Klemens Engelberg ◽  
David J. P. Ferguson ◽  
...  
Keyword(s):  
Toxoplasma Gondii ◽  
Host Cell ◽  
Cell Invasion ◽  
Membrane Skeleton ◽  
Host Cells ◽  
Model Organisms ◽  
Host Cell Invasion ◽  
Apicomplexan Parasites ◽  
Intracellular Ca ◽  
Related Proteins

AbstractInvasion of host cells by apicomplexan parasites such as Toxoplasma gondii is critical for their infectivity and pathogenesis. In Toxoplasma, secretion of essential egress, motility and invasion-related proteins from microneme organelles is regulated by oscillations of intracellular Ca2+. Later stages of invasion are considered Ca2+-independent, including the secretion of proteins required for host cell entry and remodeling from the parasite’s rhoptries. We identified a family of three Toxoplasma proteins with homology to the ferlin family of double C2 domain-containing Ca2+ sensors. In humans and model organisms such Ca2+ sensors orchestrate Ca2+-dependent exocytic membrane fusion with the plasma membrane. One ferlin that is conserved across the Apicomplexa, TgFER2, localizes to the parasite’s cortical membrane skeleton, apical end, and rhoptries. Unexpectedly, conditionally TgFER2-depleted parasites secreted their micronemes normally and were completely motile. However, these parasites were unable to invade host cells and were therefore not viable. Specifically, knockdown of TgFER2 prevented rhoptry secretion and these parasites failed to form the moving junction on the parasite-host interface necessary for host cell invasion. Collectively, these data demonstrate that the putative Ca2+ sensor TgFER2 is required for the secretion of rhoptries. These findings provide the first regulatory and mechanistic insights into this critical yet poorly understood aspect of apicomplexan host cell invasion.Graphical abstract

scholarly journals 1,3,4-Thiadiazoles Effectively Inhibit Proliferation of Toxoplasma gondii

Cells ◽  
2021 ◽  
Vol 10 (5) ◽  
pp. 1053
Author(s):  
Lidia Węglińska ◽  
Adrian Bekier ◽  
Katarzyna Dzitko ◽  
Barbara Pacholczyk-Sienicka ◽  
Łukasz Albrecht ◽  
...  
Keyword(s):  
Toxoplasma Gondii ◽  
Host Cell ◽  
Cell Invasion ◽  
Direct Action ◽  
Human Fibroblasts ◽  
Imidazole Ring ◽  
Host Cells ◽  
In Vitro Assays ◽  
Host Cell Invasion

Congenital and acquired toxoplasmosis caused by the food- and water-born parasite Toxoplasma gondii (T. gondii) is one of the most prevalent zoonotic infection of global importance. T. gondii is an obligate intracellular parasite with limited capacity for extracellular survival, thus a successful, efficient and robust host cell invasion process is crucial for its survival, proliferation and transmission. In this study, we screened a series of novel 1,3,4-thiadiazole-2-halophenylamines functionalized at the C5 position with the imidazole ring (1b–12b) for their effects on T. gondii host cell invasion and proliferation. To achieve this goal, these compounds were initially subjected to in vitro assays to assess their cytotoxicity on human fibroblasts and then antiparasitic efficacy. Results showed that all of them compare favorably to control drugs sulfadiazine and trimethoprim in terms of T. gondii growth inhibition (IC50) and selectivity toward the parasite, expressed as selectivity index (SI). Subsequently, the most potent of them with meta-fluoro 2b, meta-chloro 5b, meta-bromo 8b, meta-iodo 11b and para-iodo 12b substitution were tested for their efficacy in inhibition of tachyzoites invasion and subsequent proliferation by direct action on established intracellular infection. All the compounds significantly inhibited the parasite invasion and intracellular proliferation via direct action on both tachyzoites and parasitophorous vacuoles formation. The most effective was para-iodo derivative 12b that caused reduction in the percentage of infected host cells by 44% and number of tachyzoites per vacuole by 93% compared to non-treated host cells. Collectively, these studies indicate that 1,3,4-thiadiazoles 1b–12b, especially 12b with IC50 of 4.70 µg/mL and SI of 20.89, could be considered as early hit compounds for future design and synthesis of anti-Toxoplasma agents that effectively and selectively block the invasion and subsequent proliferation of T. gondii into host cells.

scholarly journals Evolution of Invasion in a Diverse Set of Fusobacterium Species

mBio ◽  
2014 ◽  
Vol 5 (6) ◽  
Author(s):  
Abigail Manson McGuire ◽  
Kyla Cochrane ◽  
Allison D. Griggs ◽  
Brian J. Haas ◽  
Thomas Abeel ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Preterm Birth ◽  
Host Cell ◽  
Cell Invasion ◽  
Unknown Function ◽  
Host Cells ◽  
Host Cell Invasion ◽  
Whole Genome ◽  
Large Expansion ◽  
Invading Species

ABSTRACTThe diverseFusobacteriumgenus contains species implicated in multiple clinical pathologies, including periodontal disease, preterm birth, and colorectal cancer. The lack of genetic tools for manipulating these organisms leaves us with little understanding of the genes responsible for adherence to and invasion of host cells. Actively invadingFusobacteriumspecies can enter host cells independently, whereas passively invading species need additional factors, such as compromise of mucosal integrity or coinfection with other microbes. We applied whole-genome sequencing and comparative analysis to study the evolution of active and passive invasion strategies and to infer factors associated with active forms of host cell invasion. The evolution of active invasion appears to have followed an adaptive radiation in which two of the three fusobacterial lineages acquired new genes and underwent expansions of ancestral genes that enable active forms of host cell invasion. Compared to passive invaders, active invaders have much larger genomes, encode FadA-related adhesins, and possess twice as many genes encoding membrane-related proteins, including a large expansion of surface-associated proteins containing the MORN2 domain of unknown function. We predict a role for proteins containing MORN2 domains in adhesion and active invasion. In the largest and most comprehensive comparison of sequencedFusobacteriumspecies to date, we have generated a testable model for the molecular pathogenesis ofFusobacteriuminfection and illuminate new therapeutic or diagnostic strategies.IMPORTANCEFusobacteriumspecies have recently been implicated in a broad spectrum of human pathologies, including Crohn’s disease, ulcerative colitis, preterm birth, and colorectal cancer. Largely due to the genetic intractability of member species, the mechanisms by whichFusobacteriumcauses these pathologies are not well understood, although adherence to and active invasion of host cells appear important. We examined whole-genome sequence data from a diverse set ofFusobacteriumspecies to identify genetic determinants of active forms of host cell invasion. Our analyses revealed that actively invadingFusobacteriumspecies have larger genomes than passively invading species and possess a specific complement of genes—including a class of genes of unknown function that we predict evolved to enable host cell adherence and invasion. This study provides an important framework for future studies on the role ofFusobacteriumin pathologies such as colorectal cancer.

scholarly journals A Toxoplasma gondii Ortholog of Plasmodium GAMA Contributes to Parasite Attachment and Cell Invasion

mSphere ◽  
2016 ◽  
Vol 1 (1) ◽  
Author(s):  
My-Hang Huynh ◽  
Vern B. Carruthers
Keyword(s):  
Toxoplasma Gondii ◽  
Host Cell ◽  
Cell Invasion ◽  
Genetic Evidence ◽  
Specific Role ◽  
Host Cells ◽  
Human Pathogen ◽  
Content Type ◽  
Apicomplexan Parasites ◽  
Adhesive Proteins

ABSTRACT Toxoplasma gondii is a successful human pathogen in the same phylum as malaria-causing Plasmodium parasites. Invasion of a host cell is an essential process that begins with secretion of adhesive proteins onto the parasite surface for attachment and subsequent penetration of the host cell. Conserved invasion proteins likely play roles that were maintained through the divergence of these parasites. Here, we identify a new conserved invasion protein called glycosylphosphatidylinositol-anchored micronemal antigen (GAMA). Tachyzoites lacking TgGAMA were partially impaired in parasite attachment and invasion of host cells, yielding the first genetic evidence of a specific role in parasite entry into host cells. These findings widen our appreciation of the repertoire of conserved proteins that apicomplexan parasites employ for cell invasion. Toxoplasma gondii and its Plasmodium kin share a well-conserved invasion process, including sequential secretion of adhesive molecules for host cell attachment and invasion. However, only a few orthologs have been shown to be important for efficient invasion by both genera. Bioinformatic screening to uncover potential new players in invasion identified a previously unrecognized T. gondii ortholog of Plasmodium glycosylphosphatidylinositol-anchored micronemal antigen (TgGAMA). We show that TgGAMA localizes to the micronemes and is processed into several proteolytic products within the parasite prior to secretion onto the parasite surface during invasion. TgGAMA from parasite lysate bound to several different host cell types in vitro, suggesting a role in parasite attachment. Consistent with this function, tetracycline-regulatable TgGAMA and TgGAMA knockout strains showed significant reductions in host cell invasion at the attachment step, with no defects in any of the other stages of the parasite lytic cycle. Together, the results of this work reveal a new conserved component of the adhesive repertoire of apicomplexan parasites. IMPORTANCE Toxoplasma gondii is a successful human pathogen in the same phylum as malaria-causing Plasmodium parasites. Invasion of a host cell is an essential process that begins with secretion of adhesive proteins onto the parasite surface for attachment and subsequent penetration of the host cell. Conserved invasion proteins likely play roles that were maintained through the divergence of these parasites. Here, we identify a new conserved invasion protein called glycosylphosphatidylinositol-anchored micronemal antigen (GAMA). Tachyzoites lacking TgGAMA were partially impaired in parasite attachment and invasion of host cells, yielding the first genetic evidence of a specific role in parasite entry into host cells. These findings widen our appreciation of the repertoire of conserved proteins that apicomplexan parasites employ for cell invasion.

scholarly journals Reassessing the mechanics of parasite motility and host-cell invasion

2016 ◽  
Vol 214 (5) ◽  
pp. 507-515 ◽  
Author(s):  
Isabelle Tardieux ◽  
Jake Baum
Keyword(s):  
Plasma Membrane ◽  
Host Cell ◽  
Cell Invasion ◽  
Motor System ◽  
Current Model ◽  
Cell Movement ◽  
Host Cell Invasion ◽  
Motor Functions ◽  
Apicomplexan Parasites ◽  
Parasite Motility

The capacity to migrate is fundamental to multicellular and single-celled life. Apicomplexan parasites, an ancient protozoan clade that includes malaria parasites (Plasmodium) and Toxoplasma, achieve remarkable speeds of directional cell movement. This rapidity is achieved via a divergent actomyosin motor system, housed within a narrow compartment that lies underneath the length of the parasite plasma membrane. How this motor functions at a mechanistic level during motility and host cell invasion is a matter of debate. Here, we integrate old and new insights toward refining the current model for the function of this motor with the aim of revitalizing interest in the mechanics of how these deadly pathogens move.

scholarly journals Modulation of the host cell transcriptome and epigenome by Fusobacterium nucleatum

2021 ◽  
Author(s):  
Cody A Despins ◽  
Scott D Brown ◽  
Avery V Robinson ◽  
Andrew J Mungall ◽  
Emma Allen-Vercoe ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Host Cell ◽  
Cell Invasion ◽  
Vascular Endothelial Cells ◽  
Fusobacterium Nucleatum ◽  
Opportunistic Pathogen ◽  
Host Cells ◽  
Host Cell Invasion ◽  
Primary Host ◽  
Cell Responses

Fusobacterium nucleatum (Fn) is a ubiquitous opportunistic pathogen with an emerging role as an oncomicrobe in colorectal and other cancer types. Fn can adhere to and invade host cells in a manner that varies across Fn strains and host cell phenotypes. Here we performed pairwise co-cultures between three Fn strains and two immortalized primary host cell types (colonic epithelial cells and vascular endothelial cells) followed by RNA-seq and ChIP-seq to investigate transcriptional and epigenetic host cell responses. We observed that Fn-induced host cell transcriptional modulation involves strong upregulation of genes related to immune migration and inflammatory processes, such as TNF, CXCL8, CXCL1, and CCL20. Further, we identified genes strongly upregulated specifically in conditions of host cell invasion, including overexpression of both EFNA1 and LIF, two genes commonly upregulated in colorectal cancer and associated with poor patient outcomes, and PTGS2 (COX2), a gene associated with the protective effect of aspirin in the colorectal cancer setting. Interestingly, we also observed downregulation of numerous histone modification genes upon Fn exposure. To further explore this relationship, we used the ChIP-seq data to annotate chromatin states genome-wide. We found significant chromatin remodeling following Fn exposure in conditions of host cell invasion, with substantial increases in the frequency of states corresponding to active enhancers as well as low signal or quiescent states. Thus, our results highlight increased inflammation and chemokine gene expression as conserved host cell responses to Fn exposure, and extensive host cell epigenomic changes associated with Fn host cell invasion. These results extend our understanding of Fn as an emerging pathogen and highlight the importance of considering strain heterogeneity and host cell phenotypic variation when exploring pathogenic mechanisms of Fn.

scholarly journals A Member of the Ferlin Calcium Sensor Family Is Essential for Toxoplasma gondii Rhoptry Secretion

mBio ◽  
2018 ◽  
Vol 9 (5) ◽  
Author(s):  
Bradley I. Coleman ◽  
Sudeshna Saha ◽  
Seiko Sato ◽  
Klemens Engelberg ◽  
David J. P. Ferguson ◽  
...  
Keyword(s):  
Toxoplasma Gondii ◽  
Host Cell ◽  
Cell Invasion ◽  
Host Cells ◽  
Model Organisms ◽  
Host Cell Invasion ◽  
Content Type ◽  
Calcium Dependent ◽  
Calcium Sensing ◽  
Intracellular Ca

ABSTRACT Invasion of host cells by apicomplexan parasites such as Toxoplasma gondii is critical for their infectivity and pathogenesis. In Toxoplasma, secretion of essential egress, motility, and invasion-related proteins from microneme organelles is regulated by oscillations of intracellular Ca2+. Later stages of invasion are considered Ca2+ independent, including the secretion of proteins required for host cell entry and remodeling from the parasite’s rhoptries. We identified a family of three Toxoplasma proteins with homology to the ferlin family of double C2 domain-containing Ca2+ sensors. In humans and model organisms, such Ca2+ sensors orchestrate Ca2+-dependent exocytic membrane fusion with the plasma membrane. Here we focus on one ferlin that is conserved across the Apicomplexa, T. gondii FER2 (TgFER2). Unexpectedly, conditionally TgFER2-depleted parasites secreted their micronemes normally and were completely motile. However, these parasites were unable to invade host cells and were therefore not viable. Knockdown of TgFER2 prevented rhoptry secretion, and these parasites failed to form the moving junction at the parasite-host interface necessary for host cell invasion. Collectively, these data demonstrate the requirement of TgFER2 for rhoptry secretion in Toxoplasma tachyzoites and suggest a possible Ca2+ dependence of rhoptry secretion. These findings provide the first mechanistic insights into this critical yet poorly understood aspect of apicomplexan host cell invasion. IMPORTANCE Apicomplexan protozoan parasites, such as those causing malaria and toxoplasmosis, must invade the cells of their hosts in order to establish a pathogenic infection. Timely release of proteins from a series of apical organelles is required for invasion. Neither the vesicular fusion events that underlie secretion nor the observed reliance of the various processes on changes in intracellular calcium concentrations is completely understood. We identified a group of three proteins with strong homology to the calcium-sensing ferlin family, which are known to be involved in protein secretion in other organisms. Surprisingly, decreasing the amounts of one of these proteins (TgFER2) did not have any effect on the typically calcium-dependent steps in invasion. Instead, TgFER2 was essential for the release of proteins from organelles called rhoptries. These data provide a tantalizing first look at the mechanisms controlling the very poorly understood process of rhoptry secretion, which is essential for the parasite’s infection cycle.

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