Structural insights into microneme protein assembly reveal a new mode of EGF domain recognition

Sawmynaden, Kovilen; Saouros, Savvas; Friedrich, Nikolas; Marchant, Jan; Simpson, Peter; Bleijlevens, Boris; Blackman, Michael J.; Soldati‐Favre, Dominique; Matthews, Stephen

doi:10.1038/embor.2008.179

Cited by 32 publications

(40 citation statements)

References 21 publications

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“…Microneme proteins are discharged onto the parasite surface and form tight interactions with host cell receptors. MIC6 forms a multimeric complex with MIC1 and MIC4 and contributes to invasion of host cells by tachyzoites in vitro and to virulence in vivo (4,8,31). Therefore, it appears that infected hosts recognize MIC6 and produce antibodies when active growth of tachyzoites occurs.…”

Section: Discussionmentioning

confidence: 99%

Toxoplasma gondii Antigens Recognized by IgG Antibodies Differ between Mice with and without Active Proliferation of Tachyzoites in the Brain during the Chronic Stage of Infection

Hester

Mullins

et al. 2012

Infect Immun

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We examined whether tachyzoite proliferation in the brains of immunocompetent hosts during the chronic stage of infection with Toxoplasma gondii induces production of IgG antibodies that recognize parasite antigens different from those recognized by the antibodies of infected hosts that do not have tachyzoite growth. For this purpose, two groups of CBA/J mice, which display continuous tachyzoite growth in their brains during the later stage of infection, were infected, and one group received treatment with sulfadiazine to prevent tachyzoite proliferation during the chronic stage of infection. T. gondii antigens recognized by the IgG antibodies from these two groups of mice were compared using immunoblotting following separation of tachyzoite antigens by two-dimensional gel electrophoresis. Several antigens, including the microneme protein MIC2, the cyst matrix protein MAG1, and the dense granule proteins GRA4 and GRA7, were commonly recognized by IgG antibodies from both groups of mice. There were multiple antigens recognized mostly by IgG antibodies of only one group of mice, either with or without cerebral tachyzoite growth. The antigens recognized only by or mostly by the antibodies of mice with cerebral tachyzoite growth include MIC6, the rhoptry protein ROP1, GRA2, one heat shock protein HSP90, one (putative) HSP70, and the myosin heavy chain. These results indicate that levels of IgG antibody to only selected T. gondii antigens increase in association with cerebral tachyzoite proliferation (reactivation of infection) in immunocompetent hosts with chronic infection.

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Section: Discussionmentioning

confidence: 99%

Toxoplasma gondii Antigens Recognized by IgG Antibodies Differ between Mice with and without Active Proliferation of Tachyzoites in the Brain during the Chronic Stage of Infection

Hester

Mullins

et al. 2012

Infect Immun

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“…Thus, the first two methods are suitable for an initial evaluation of parasite invasion and real-time PCR participates as an unequivocal quantitative assay. The incubation of N. caninum or T. gondii at 37°C is a current way to determine the in vitro effects of drugs or sera on the invasion phenomenon (NAGULESWARAN et al, 2003;PEREIRA et al, 2011;SAWMYNADEN et al, 2008). Our results indicate that all these methods show losses of invasive/adhesive tachyzoites incubated at 37°C; therefore, this is an important effect to take into account when designing assays that require long incubation times.…”

Section: Discussionmentioning

confidence: 84%

Comparison of an ELISA assay for the detection of adhesive/invasive Neospora caninum tachyzoites

Pereira

Yatsuda

2014

Rev. Bras. Parasitol. Vet.

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Neospora caninum belongs to the phylum Apicomplexa, the causative agent of neosporosis, which leads to economic impacts on cattle production. A common feature among apicomplexan parasites is the invasive process driven mostly by the parasite. As a first evaluation of candidate molecules that play a possible role by interfering in this invasive process, the in vitro invasion assay is a fast and direct way to screen future agonists or antagonists. This work involved the development of a new cell culture ELISA and transient β-galactosidase activity applied to the semi-quantitative detection of N. caninum in Vero cell culture. Cell culture ELISA is based on histochemistry and immunology, resulting in a colorimetric reaction. The β-galactosidase activity was obtained by the transient transfection of the lacZ gene under control of RPS13 promoter of N. caninum. These methods were used to evaluate the effects of temperature (37°C and 85°C) on the invasion and adhesion of tachyzoites. The three tested methods (real time PCR, β-galactosidase activity and ELISA) showed a similar pattern, indicating that different methods may be complementary.Keywords: Neospora caninum, cell culture ELISA, LacZ gene, transfection, real-time PCR, invasion assay. ResumoNeospora caninum, parasita do filo Apicomplexa, é causador da neosporose, doença responsável por perdas econômicas importantes na pecuária. Um fator comum entre os apicomplexas é o processo de invasão majoritariamente dirigido pelo parasita. Dentre as primeiras avaliações de moléculas candidatas, que possivelmente interferem no processo de invasão, o ensaio de invasão in vitro é um meio rápido e direto de selecionar futuros agonistas ou antagonistas. Este trabalho desenvolveu um novo ELISA baseado em cultura (Cell-culture ELISA) e um ensaio que mede a atividade transiente de β-galactosidase, aplicados para a detecção semiquantitativa de N. caninum em células Vero. Cell-culture ELISA é baseado em histoquímica e imunologia, resultando em uma reação colorimétrica. A atividade da β-galactosidase foi obtida pela transfecção transiente do gene LacZ sob controle do promotor RPS13 de N. caninum. Esses métodos avaliaram os efeitos da temperatura (37°C e 85°C) sobre a invasão e adesão. Os três métodos testados (real time PCR, atividade de β-galactosidase e ELISA) mostraram um padrão similar, indicando que diferentes métodos podem ser complementares. Adicionalmente, esse ELISA é adequado para aplicação em laboratórios carentes de uma complexa estrutura para métodos de detecção moleculares.Palavras-chave: Neospora caninum, cell-culture ELISA, gene LacZ, transfecção, PCR em tempo real, ensaio de invasão.

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“…In contrast to PfEBA175, TgMIC1 does not dimerise, but is present in the TgMIC1-4-6 complex, with stoichiometry that allows for the presentation of at least two molecules of TgMIC1 in close proximity. Despite this, the presentation of a single TgMIC1 is functional in invasion, but its arrangement within the complex appears to increase the affinity of the complex to host cell receptor(s) or may allow TgMIC1 to target multi-antennary carbohydrate structures including gangliosides which in turn results in enhanced invasion efficiency (Sawmynaden et al, 2008).…”

Section: Discussionmentioning

confidence: 99%

“…The apple domain is another potential carbohydrate-binding fold and several apple-domain-containing proteins are present in the T. gondii genome. Genetic disruption established the non-essential nature of the TgMIC1-MIC4-MIC6 (TgMIC1-4-6) complex (Reiss et al, 2001), however TgMIC1 has a significant role in invasion and contributes to virulence in the mouse model (Cerede et al, 2005;Blumenschein et al, 2007;Sawmynaden et al, 2008). Parasite attachment to host cells is not significantly impaired in the mic1ko strain and therefore cannot explain the invasion defect.…”

Section: Importance Of Sialic Acids In Host Invasion By Coccidiansmentioning

confidence: 99%

Sialic acids: Key determinants for invasion by the Apicomplexa

Friedrich

Matthews

Soldati‐Favre

2010

International Journal for Parasitology

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a b s t r a c tSialic acids are ubiquitously found on the surface of all vertebrate cells at the extremities of glycan chains and widely exploited by viruses and bacteria to enter host cells. Carbohydrate-bearing receptors are equally important for host cell invasion by the obligate intracellular protozoan parasites of the phylum Apicomplexa. Host cell entry is an active process relying crucially on proteins that engage with receptors on the host cell surface and promote adhesion and internalisation. Assembly into complexes, proteolytic processing and oligomerization are important requirements for the functionality of these adhesins. The combination of adhesive proteins with varying stringency in specificity confers some flexibility to the parasite in face of receptor heterogeneity and immune pressure. Sialic acids are now recognised to critically contribute to selective host cell recognition by various species of the phylum. Ó

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Structural insights into microneme protein assembly reveal a new mode of EGF domain recognition

Cited by 32 publications

References 21 publications

Toxoplasma gondii Antigens Recognized by IgG Antibodies Differ between Mice with and without Active Proliferation of Tachyzoites in the Brain during the Chronic Stage of Infection

Toxoplasma gondii Antigens Recognized by IgG Antibodies Differ between Mice with and without Active Proliferation of Tachyzoites in the Brain during the Chronic Stage of Infection

Comparison of an ELISA assay for the detection of adhesive/invasive Neospora caninum tachyzoites

Sialic acids: Key determinants for invasion by the Apicomplexa

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