The lectin-specific activity of Toxoplasma gondii microneme proteins 1 and 4 binds Toll-like receptor 2 and 4 N-glycans to regulate innate immune priming

Abstract

Infection of host cells by Toxoplasma gondii is an active process, which is regulated by secretion of microneme (MICs) and rhoptry proteins (ROPs and RONs) from specialized organelles in the apical pole of the parasite. MIC1, MIC4 and MIC6 assemble into an adhesin complex secreted on the parasite surface that functions to promote infection competency. MIC1 and MIC4 are known to bind terminal sialic acid residues and galactose residues, respectively and to induce IL-12 production from splenocytes. Here we show that rMIC1- and rMIC4-stimulated dendritic cells and macrophages produce proinflammatory cytokines, and they do so by engaging TLR2 and TLR4. This process depends on sugar recognition, since point mutations in the carbohydrate-recognition domains (CRD) of rMIC1 and rMIC4 inhibit innate immune cells activation. HEK cells transfected with TLR2 glycomutants were selectively unresponsive to MICs. Following in vitro infection, parasites lacking MIC1 or MIC4, as well as expressing MIC proteins with point mutations in their CRD, failed to induce wild-type (WT) levels of IL-12 secretion by innate immune cells. However, only MIC1 was shown to impact systemic levels of IL-12 and IFN-γ in vivo . Together, our data show that MIC1 and MIC4 interact physically with TLR2 and TLR4 N-glycans to trigger IL-12 responses, and MIC1 is playing a significant role in vivo by altering T . gondii infection competency and murine pathogenesis.

Targeting and Recognition of Toll-Like Receptors by Plant and Pathogen Lectins

“…The lectins’ interactions with macrophages and dendritic cells are established by the recognition of TLR2 and TLR4 N-glycans and result in modulation of the immune response toward the Th1 axis. Sardinha-Silva et al ( 23 ) have recently demonstrated the importance of these lectins for the induction of early IL-12 production in T. gondii infection. Concerning the discrimination of which TLR2 N-glycans are targeted by Tg MIC1 and Tg MIC4, those occupying positions 2, 3, and 4, impact the Tg MIC1–TLR2 interaction.…”

“…Concerning the discrimination of which TLR2 N-glycans are targeted by Tg MIC1 and Tg MIC4, those occupying positions 2, 3, and 4, impact the Tg MIC1–TLR2 interaction. On the other hand, Tg MIC4 requires the TLR2 N-glycans linked to the sites 3 and 4 to interact with the receptor ( 23 ).…”

Interação de proteínas de micronema de >i<Toxoplasma gondii>/i< com glicanas N-ligadas a TLR2 e TLR4: reflexos sobre a ativação celular

“…MIC1 possui massa molecular de 49 kDa, é solúvel e multifuncional, capaz de se ligar a glicoconjugados que apresentam α2-3 sialilactosaminas na região terminal das Nglicanas (Blumenschein et al, 2007;Friedrich et al, 2010;Sardinha-Silva et al, 2017).…”

“…8B). Após o processo de diálise, realizou-se dois ensaios de controle de qualidade das lectinas recombinantes: o primeiro para averiguar a ligação das lectinas ao açúcar específico e outro para verificar a capacidade das lectinas estimularem macrófagos a produzirem IL-12 (Pinzan et al, 2015;Sardinha-Silva et al, 2017).…”

Avaliação funcional de Toll-Like Receptor 4 (TLR4) em células trofoblásticas humanas vilosa/extravilosa e em vilos placentários humanos de terceiro trimestre gestacional infectados por Toxoplasma gondii

“…A invasão da célula é um processo ativo. O parasito invade a célula hospedeira através da expressão e liberação de proteínas de micronemas (TgMICs) e de roptrias (ROPs e ROMs), organelas especializadas presentes na parte apical do parasito (SARDINHA-SILVA et al, 2017). As micronemas são responsáveis pela adesão do parasito à célula hospedeira (SOLDATI-FAVRE, 2008;SOUZA et al, 2010;JIMENEZ-RUIZ et al, 2016), enquanto as roptrias e grânulos densos estão envolvidos na invasão propriamente dita e formação do vacúolo parasitóforo, que é derivado da membrana celular do hospedeiro (SOUZA et al, 2010).…”