Receptor Heterodimerization and Co-Receptor Engagement in TLR2 Activation Induced by MIC1 and MIC4 from Toxoplasma gondii.

Flávia Costa Mendonça-Natividade,Rafael Ricci-Azevedo,Ademilson Panunto-Castelo,Carla Duque Lopes,Aline Sardinha-Silva,Camila Figueiredo Pinzan,Maria Roque-Barreira,Igor C Almeida,Alan B Carneiro,Lilian L Nohara,Ana Paiva Alegre-Maller

doi:10.3390/ijms20205001

Abstract

The microneme organelles of Toxoplasma gondii tachyzoites release protein complexes (MICs), including one composed of the transmembrane protein MIC6 plus MIC1 and MIC4. In this complex, carbohydrate recognition domains of MIC1 and MIC4 are exposed and interact with terminal sialic acid and galactose residues, respectively, of host cell glycans. Recently, we demonstrated that MIC1 and MIC4 binding to the N-glycans of Toll-like receptor (TLR) 2 and TLR4 on phagocytes triggers cell activation and pro-inflammatory cytokine production. Herein, we investigated the requirement for TLR2 heterodimerization and co-receptors in MIC-induced responses, as well as the signaling molecules involved. We used MICs to stimulate macrophages and HEK293T cells transfected with TLR2 and TLR1 or TLR6, both with or without the co-receptors CD14 and CD36. Then, the cell responses were analyzed, including nuclear factor-kappa B (NF-κB) activation and cytokine production, which showed that (1) only TLR2, among the studied factors, is crucial for MIC-induced cell activation; (2) TLR2 heterodimerization augments, but is not critical for, activation; (3) CD14 and CD36 enhance the response to MIC stimulus; and (4) MICs activate cells through a transforming growth factor beta-activated kinase 1 (TAK1)-, mammalian p38 mitogen-activated protein kinase (p38)-, and NF-κB-dependent pathway. Remarkably, among the studied factors, the interaction of MIC1 and MIC4 with TLR2 N-glycans is sufficient to induce cell activation, which promotes host protection against T. gondii infection.

Highlights

Toxoplasma gondii, a ubiquitous intracellular protozoan parasite of the phylum Apicomplexa, has received considerable scientific and medical attention due to its high transmissibility and endemicity [1]
The preparations that were assayed for their ability to stimulate cell activation were the recombinant microneme proteins rMIC1 and rMIC4 and the Lac+ fraction, which is a tachyzoite fraction containing soluble antigens, including native MIC1 and MIC4, that was obtained by affinity binding to immobilized lactose
The IL-12 levels produced by untreated bone marrow-derived macrophages (BMDMs) that were stimulated with microneme proteins were close to those in LPS-stimulated BMDMs

Summary

Introduction

Toxoplasma gondii, a ubiquitous intracellular protozoan parasite of the phylum Apicomplexa, has received considerable scientific and medical attention due to its high transmissibility and endemicity [1]. It has been estimated that approximately 30% of the human population is infected by T. gondii [2,3]. Host cell invasion by T. gondii is an active process that relies on the motility of the tachyzoite, which requires its actomyosin system, and protein secretion from two apical organelles, micronemes and rhoptries [5]. MIC1 and MIC4 are exposed on the tachyzoite surface and bind to host cell surface receptors, and MIC6 is a transmembrane protein that binds the complex to the parasite surface. Together, these proteins promote tachyzoite adhesion and subsequent host cell invasion [6,7,8]

Methods

Results

Discussion

Conclusion