scholarly journals Identification of a Neospora caninum Microneme Protein (NcMIC1) Which Interacts with Sulfated Host Cell Surface Glycosaminoglycans

2002 ◽  
Vol 70 (6) ◽  
pp. 3187-3198 ◽  
Author(s):  
Nadine Keller ◽  
Arunasalam Naguleswaran ◽  
Angela Cannas ◽  
Nathalie Vonlaufen ◽  
Marianne Bienz ◽  
...  
Keyword(s):  
Cell Surface ◽  
Host Cell ◽  
Solid Phase ◽  
Neospora Caninum ◽  
Host Cells ◽  
Sequence Motif ◽  
Host Cell Surface ◽  
Adhesive Proteins ◽  
Microneme Protein ◽  
Microneme Proteins

ABSTRACT The invasive stages of apicomplexan parasites enter their host cells through mechanisms which are largely conserved throughout the phylum. Host cell invasion is divided into two distinct events, namely, adhesion onto the host cell surface and the actual host cell entry process. The former is mediated largely through microneme proteins which are secreted at the onset of establishing contact with the host cell surface. Many of the microneme proteins identified so far contain adhesive domains. We here present the genomic and corresponding cDNA sequences coding for a 460-amino-acid (aa) microneme protein in Neospora caninum tachyzoites which, due to its homology to MIC1 in Toxoplasma gondii (TgMIC1), was named NcMIC1. The deduced NcMIC1 polypeptide sequence contains an N-terminal signal peptide of 20 aa followed by two tandemly internal repeats of 48 and 44 aa, respectively. Integrated into each repeat is a CXXXCG sequence motif reminiscent of the thrombospondin-related family of adhesive proteins. The positioning of this motif is strictly conserved in TgMIC1 and NcMIC1. The C-terminal part, comprised of 278 aa, was expressed in Escherichia coli, and antibodies affinity purified on recombinant NcMIC1 were used to confirm the localization within the micronemes by immunofluorescence and immunogold transmission electron microscopy of tachyzoites. Immunohistochemistry of mouse brains infected with tissue cysts showed that expression of this protein is reduced in the bradyzoite stage. Upon initiation of secretion by elevating the temperature to 37°C, NcMIC1 is released into the medium supernatant. NcMIC1 binds to trypsinized, rounded Vero cells, as well as to Vero cell monolayers. Removal of glycosaminoglycans from the host cell surface and modulation of host cell surface glycosaminoglycan sulfation significantly reduces the binding of NcMIC1 to the host cell surface. Solid-phase binding assays employing defined glycosaminoglycans confirmed that NcMIC1 binds to sulfated glycosaminoglycans.

scholarly journals A Single-Pass Type I Membrane Protein from the Apicomplexan Parasite Cryptosporidium parvum with Nanomolar Binding Affinity to Host Cell Surface

2021 ◽  
Vol 9 (5) ◽  
pp. 1015
Author(s):  
Tianyu Zhang ◽  
Xin Gao ◽  
Dongqiang Wang ◽  
Jixue Zhao ◽  
Nan Zhang ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Cell Surface ◽  
Host Cell ◽  
Binding Affinity ◽  
Cryptosporidium Parvum ◽  
Host Cells ◽  
Watery Diarrhea ◽  
Type I ◽  
Single Pass ◽  
Host Cell Surface

Cryptosporidium parvum is a globally recognized zoonotic parasite of medical and veterinary importance. This parasite mainly infects intestinal epithelial cells and causes mild to severe watery diarrhea that could be deadly in patients with weakened or defect immunity. However, its molecular interactions with hosts and pathogenesis, an important part in adaptation of parasitic lifestyle, remain poorly understood. Here we report the identification and characterization of a C. parvum T-cell immunomodulatory protein homolog (CpTIPH). CpTIPH is a 901-aa single-pass type I membrane protein encoded by cgd5_830 gene that also contains a short Vibrio, Colwellia, Bradyrhizobium and Shewanella (VCBS) repeat and relatively long integrin alpha (ITGA) N-terminus domain. Immunofluorescence assay confirmed the location of CpTIPH on the cell surface of C. parvum sporozoites. In congruence with the presence of VCBS repeat and ITGA domain, CpTIPH displayed high, nanomolar binding affinity to host cell surface (i.e., Kd(App) at 16.2 to 44.7 nM on fixed HCT-8 and CHO-K1 cells, respectively). The involvement of CpTIPH in the parasite invasion is partly supported by experiments showing that an anti-CpTIPH antibody could partially block the invasion of C. parvum sporozoites into host cells. These observations provide a strong basis for further investigation of the roles of CpTIPH in parasite-host cell interactions.

scholarly journals Heparan Sulfate Facilitates Spike Protein-Mediated SARS-CoV-2 Host Cell Invasion and Contributes to Increased Infection of SARS-CoV-2 G614 Mutant and in Lung Cancer

2021 ◽  
Vol 8 ◽  
Author(s):  
Jingwen Yue ◽  
Weihua Jin ◽  
Hua Yang ◽  
John Faulkner ◽  
Xuehong Song ◽  
...  
Keyword(s):  
Lung Cancer ◽  
Cell Surface ◽  
Heparan Sulfate ◽  
Host Cell ◽  
Host Cells ◽  
Cell Surface Receptor ◽  
Spike Protein ◽  
Effective Prevention ◽  
Heparin Derivatives ◽  
Host Cell Surface

The severe acute respiratory syndrome (SARS)-like coronavirus disease (COVID-19) is caused by SARS-CoV-2 and has been a serious threat to global public health with limited treatment. Cellular heparan sulfate (HS) has been found to bind SARS-CoV-2 spike protein (SV2-S) and co-operate with cell surface receptor angiotensin-converting enzyme 2 (ACE2) to mediate SARS-CoV-2 infection of host cells. In this study, we determined that host cell surface SV2-S binding depends on and correlates with host cell surface HS expression. This binding is required for SARS-Cov-2 virus to infect host cells and can be blocked by heparin lyase, HS antagonist surfen, heparin, and heparin derivatives. The binding of heparin/HS to SV2-S is mainly determined by its overall sulfation with potential, minor contribution of specific SV2-S binding motifs. The higher binding affinity of SV2-S G614 mutant to heparin and upregulated HS expression may be one of the mechanisms underlying the higher infectivity of the SARS-CoV-2 G614 variant and the high vulnerability of lung cancer patients to SARS-CoV-2 infection, respectively. The higher host cell infection by SARS-CoV-2 G614 variant pseudovirus and the increased infection caused by upregulated HS expression both can be effectively blocked by heparin lyase and heparin, and possibly surfen and heparin derivatives too. Our findings support blocking HS-SV2-S interaction may provide one addition to achieve effective prevention and/treatment of COVID-19.

scholarly journals Mechanism of CD150 (SLAM) Down Regulation from the Host Cell Surface by Measles Virus Hemagglutinin Protein

2004 ◽  
Vol 78 (18) ◽  
pp. 9666-9674 ◽  
Author(s):  
G. Grant Welstead ◽  
Eric C. Hsu ◽  
Caterina Iorio ◽  
Shelly Bolotin ◽  
Christopher D. Richardson
Keyword(s):  
Cell Surface ◽  
Host Cell ◽  
Measles Virus ◽  
Host Cells ◽  
Down Regulation ◽  
Cellular Receptors ◽  
Regulatory Molecule ◽  
Hemagglutinin Protein ◽  
Host Cell Surface ◽  
H Protein

ABSTRACT Measles virus has been reported to enter host cells via either of two cellular receptors, CD46 and CD150 (SLAM). CD46 is found on most cells of higher primates, while SLAM is expressed on activated B, T, and dendritic cells and is an important regulatory molecule of the immune system. Previous reports have shown that measles virus can down regulate expression of its two cellular receptors on the host cell surface during infection. In this study, the process of down regulation of SLAM by measles virus was investigated. We demonstrated that expression of the hemagglutinin (H) protein of measles virus was sufficient for down regulation. Our studies provided evidence that interactions between H and SLAM in the endoplasmic reticulum (ER) can promote the down regulation of SLAM but not CD46. In addition, we demonstrated that interactions between H and SLAM at the host cell surface can also contribute to SLAM down regulation. These results indicate that two mechanisms involving either intracellular interactions between H and SLAM in the ER or receptor-mediated binding to H at the surfaces of host cells can lead to the down regulation of SLAM during measles virus infection.

scholarly journals EnP1, a Microsporidian Spore Wall Protein That Enables Spores To Adhere to and Infect Host Cells In Vitro

2007 ◽  
Vol 6 (8) ◽  
pp. 1354-1362 ◽  
Author(s):  
Timothy R. Southern ◽  
Carrie E. Jolly ◽  
Melissa E. Lester ◽  
J. Russell Hayman
Keyword(s):  
Cell Surface ◽  
Host Cell ◽  
Spore Wall ◽  
Site Directed Mutagenesis ◽  
Host Cells ◽  
Binding Motif ◽  
Heparin Binding ◽  
Cell Infection ◽  
Host Cell Surface

ABSTRACT Microsporidia are spore-forming fungal pathogens that require the intracellular environment of host cells for propagation. We have shown that spores of the genus Encephalitozoon adhere to host cell surface glycosaminoglycans (GAGs) in vitro and that this adherence serves to modulate the infection process. In this study, a spore wall protein (EnP1; Encephalitozoon cuniculi ECU01_0820) from E. cuniculi and Encephalitozoon intestinalis is found to interact with the host cell surface. Analysis of the amino acid sequence reveals multiple heparin-binding motifs, which are known to interact with extracellular matrices. Both recombinant EnP1 protein and purified EnP1 antibody inhibit spore adherence, resulting in decreased host cell infection. Furthermore, when the N-terminal heparin-binding motif is deleted by site-directed mutagenesis, inhibition of adherence is ablated. Our transmission immunoelectron microscopy reveals that EnP1 is embedded in the microsporidial endospore and exospore and is found in high abundance in the polar sac/anchoring disk region, an area from which the everting polar tube is released. Finally, by using a host cell binding assay, EnP1 is shown to bind host cell surfaces but not to those that lack surface GAGs. Collectively, these data show that given its expression in both the endospore and the exospore, EnP1 is a microsporidian cell wall protein that may function both in a structural capacity and in modulating in vitro host cell adherence and infection.

scholarly journals Identification and Characterization of a Neospora caninum Microneme-Associated Protein (NcMIC4) That Exhibits Unique Lactose-Binding Properties

2004 ◽  
Vol 72 (8) ◽  
pp. 4791-4800 ◽  
Author(s):  
Nadine Keller ◽  
Michèle Riesen ◽  
Arunasalam Naguleswaran ◽  
Nathalie Vonlaufen ◽  
Rebecca Stettler ◽  
...  
Keyword(s):  
Host Cell ◽  
Gel Electrophoresis ◽  
Neospora Caninum ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Vero Cells ◽  
Host Cells ◽  
Immunogold Electron Microscopy ◽  
Binding Properties ◽  
Microneme Protein

ABSTRACT Microneme proteins have been shown to play an important role in the early phase of host cell adhesion, by mediating the contact between the parasite and host cell surface receptors. In this study we have identified and characterized a lectin-like protein of Neospora caninum tachyzoites which was purified by α-lactose-agarose affinity chromatography. Upon separation by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, this lactose-binding protein migrated at 70 and 55 kDa under reducing and nonreducing conditions, respectively. Immunofluorescence and immunogold electron microscopy with affinity-purified antibodies showed that the protein was associated with the tachyzoite micronemes. Mass spectrometry analyses and expressed sequence tag database mining revealed that this protein is a member of the Neospora microneme protein family; the protein was named NcMIC4 (N. caninum microneme protein 4). Upon two-dimensional gel electrophoresis, NcMIC4 separated into seven distinct isoforms. Incubation of extracellular parasites at 37°C resulted in the secretion of NcMIC4 into the medium as a soluble protein, and the secreted protein exhibited a slightly reduced M r but retained its lactose-binding properties. Immunofluorescence was used to investigate the temporal and spatial distribution of NcMIC4 in tachyzoites entering their host cells and showed that reexpression of NcMIC4 took place 30 min after entry into the host cell. Incubation of secreted fractions and purified NcMIC4 with Vero cells demonstrated binding of NcMIC4 to Vero cells as well as binding to chondroitin sulfate A glycosaminoglycans.

scholarly journals Role of Sulfated Glycans in Adherence of the Microsporidian Encephalitozoon intestinalis to Host Cells In Vitro

2005 ◽  
Vol 73 (2) ◽  
pp. 841-848 ◽  
Author(s):  
J. Russell Hayman ◽  
Timothy R. Southern ◽  
Theodore E. Nash
Keyword(s):  
Cell Surface ◽  
Host Cell ◽  
Mutant Cell ◽  
Host Cells ◽  
Target Cells ◽  
Cell Infection ◽  
Encephalitozoon Intestinalis ◽  
Successful Infection ◽  
Host Cell Surface ◽  
Spore Adherence

ABSTRACT Microsporidia are obligate intracellular opportunistic protists that infect a wide variety of animals, including humans, via environmentally resistant spores. Infection requires that spores be in close proximity to host cells so that the hollow polar tube can pierce the cell membrane and inject the spore contents into the cell cytoplasm. Like other eukaryotic microbes, microsporidia may use specific mechanisms for adherence in order to achieve target cell proximity and increase the likelihood of successful infection. Our data show that Encephalitozoon intestinalis exploits sulfated glycans such as the cell surface glycosaminoglycans (GAGs) in selection of and attachment to host cells. When exogenous sulfated glycans are used as inhibitors in spore adherence assays, E. intestinalis spore adherence is reduced by as much as 88%. However, there is no inhibition when nonsulfated glycans are used, suggesting that E. intestinalis spores utilize sulfated host cell glycans in adherence. These studies were confirmed by exposure of host cells to xylopyranoside, which limits host cell surface GAGs, and sodium chlorate, which decreases surface sulfation. Spore adherence studies with CHO mutant cell lines that are deficient in either surface GAGs or surface heparan sulfate also confirmed the necessity of sulfated glycans. Furthermore, when spore adherence is inhibited, host cell infection is reduced, indicating a direct association between spore adherence and infectivity. These data show that E. intestinalis specifically adheres to target cells by way of sulfated host cell surface GAGs and that this mechanism serves to enhance infectivity.

scholarly journals Microneme Protein 6 Is Involved in Invasion and Egress by Neospora caninum

Pathogens ◽  
2021 ◽  
Vol 10 (2) ◽  
pp. 201
Author(s):  
Xianmei Wang ◽  
Di Tang ◽  
Fei Wang ◽  
Gaowei Jin ◽  
Lifang Wang ◽  
...  
Keyword(s):  
Neospora Caninum ◽  
Parasite Load ◽  
Host Cells ◽  
Host Cell Invasion ◽  
Caninum Infection ◽  
Apicomplexan Parasites ◽  
Key Steps ◽  
Microneme Protein ◽  
Adhesion And Invasion ◽  
Microneme Proteins

Background: Neospora caninum, is the etiological agent of neosporosis, an infection that causes abortions in cattle and nervous system dysfunction in dogs. Invasion and egress are the key steps of the pathogenesis of N. caninum infection. Microneme proteins (MICs) play important roles in the recognition, adhesion, and invasion of host cells in other apicomplexan parasites. However, some MICs and their functions in N. caninum infection have rarely been reported. Methods: The homologous recombination strategy was used to investigate the function of MIC6 in N. caninum infection. Results: ΔNcMIC6 showed a smaller plaque size and weakened capacities of invasion and egress than Nc1. Transcription levels of the egress-related genes CDPK1, PLP1, and AMA1 of ΔNcMIC6 were downregulated. Due to the lack of NcMIC6, virulence of the pathogen in the infected mouse was weakened. The subcellular localization of NcMIC1 and NcMIC4 in ΔNcMIC6, however, did not change. Nevertheless, the transcription levels of MIC1 and MIC4 in ΔNcMIC6 were downregulated, and the expression and secretion of MIC1 and MIC4 in ΔNcMIC6 were reduced compared with that in Nc1. Furthermore, the absence of NcMIC6 weakened the virulence in mice and lower parasite load detected in mice brains. Conclusions: NcMIC6 is involved in host cell invasion and egress in N. caninum and may work synergistically with other MICs to regulate the virulence of the pathogen. These data lay a foundation for further research into the function and application of NcMIC6.

scholarly journals hFUT1-based live cell assay to profile α1-2-fucosides enhanced influenza A virus infection

2019 ◽  
Author(s):  
Senlian Hong ◽  
Geramie Grande ◽  
Chenhua Yu ◽  
Digantkumar G. Chapla ◽  
Natalie Reigh ◽  
...  
Keyword(s):  
Cell Surface ◽  
Host Cell ◽  
Influenza A Virus ◽  
Influenza A ◽  
Host Cells ◽  
Viral Pathogen ◽  
Host Tropism ◽  
Host Cell Surface ◽  
Different Strains

AbstractHost cell-surface glycans play critical roles in influenza A virus (IAV) infection ranging from modulation of IAV attachment to membrane fusion and host tropism. Approaches for quick and sensitive profiling of the viral avidity towards a specific type of host-cell glycan can contribute to the understanding of tropism switching among different strains of IAV. In this study, we developed a method based on chemoenzymatic glycan engineering to investigate the possible involvement of α1-2-fucosides in IAV infections. Using a truncated human fucosyltransferase 1 (hFuT1), we were able to create α1-2-linked fucosides in situ on the host cell surface to assess their influence on the host cell binding to IAV hemagglutinin and the susceptibility of host cells toward IAV induced killing. We discovered that the newly added α1-2-fucosides on host cells enhanced the infection of several human pandemic IVA subtypes. These findings suggest that glycan epitopes other than sialic aicds should be taken into consideration for assessing the human pandemic risk of this viral pathogen.

scholarly journals Mathematical Modelling of Actin treadmill in Apicomplexans

2018 ◽  
Author(s):  
Mahendra Prajapat ◽  
Samridhi Pathak ◽  
Ricka Gauba ◽  
Avinash Kale ◽  
Supreet Saini
Keyword(s):  
Cell Surface ◽  
Host Cell ◽  
Host Cells ◽  
Tight Control ◽  
Nucleation Time ◽  
Host Cell Membrane ◽  
Host Cell Surface ◽  
Average Size ◽  
Fine Tune ◽  
Representative Member

AbstractPlasmodium parasite, a representative member of phylum Apicomplexa is a causative agent of malaria in human as well as other animals. To infect host cells, Plasmodium first finds receptors on the host cell surface, then binds specifically, and finally penetrates host cell membrane to acquire the host cellular resources. The motility for moving on the cell surface is equipped by the precise and tight control of actin treadmill. Several regulators are required to achieve precision and robustness in the control of actin treadmill. However, the mechanistic detail of the treadmill regulatory network and the cross-talk among regulators are not well understood. We developed a stochastic model of treadmill regulation and explored the dynamics of filament growth, nucleation time, and elongation time. Our study mainly highlighted on how and what helps cells to maintain an average size of the actin filaments within a species. This is particularly important, since, excessive growth of filament can lead to cell lysis. Moreover, we also explore how the regulators interact to fine-tune the control elements in the actin treadmill.

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