Myristoylated rhinovirus VP4 protein activates TLR2-dependent proinflammatory gene expression
| Autor: | Suraj Jaipalli, Marc B. Hershenson, Adam M. Goldsmith, J. Kelley Bentley, Tomoko Ishikawa, Jing Lei, Charu Rajput, Mingyuan Han, Joanna L. Hinde |
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| Rok vydání: | 2019 |
| Předmět: |
Male
0301 basic medicine Rhinovirus Physiology viruses medicine.medical_treatment Virus Replication medicine.disease_cause Mice 0302 clinical medicine Gene expression Macrophage Child Receptor Toll-like receptor Chemistry Cytokine Host-Pathogen Interactions Female Myristic Acids Protein Binding Signal Transduction Research Article Pulmonary and Respiratory Medicine Adolescent Proinflammatory cytokine Microbiology Viral Proteins 03 medical and health sciences Physiology (medical) Eosinophilia medicine Animals Humans Amino Acid Sequence Picornaviridae Infections Macrophages Epithelial Cells Cell Biology Asthma Toll-Like Receptor 2 Mice Inbred C57BL TLR2 HEK293 Cells 030104 developmental biology 030228 respiratory system Capsid Proteins Protein Processing Post-Translational HeLa Cells |
| Zdroj: | Am J Physiol Lung Cell Mol Physiol |
| ISSN: | 1522-1504 1040-0605 |
| Popis: | Asthma exacerbations are often caused by rhinovirus (RV). We and others have shown that Toll-like receptor 2 (TLR2), a membrane surface receptor that recognizes bacterial lipopeptides and lipoteichoic acid, is required and sufficient for RV-induced proinflammatory responses in vitro and in vivo. We hypothesized that viral protein-4 (VP4), an internal capsid protein that is myristoylated upon viral replication and externalized upon viral binding, is a ligand for TLR2. Recombinant VP4 and myristoylated VP4 (MyrVP4) were purified by Ni-affinity chromatography. MyrVP4 was also purified from RV-A1B-infected HeLa cells by urea solubilization and anti-VP4 affinity chromatography. Finally, synthetic MyrVP4 was produced by chemical peptide synthesis. MyrVP4-TLR2 interactions were assessed by confocal fluorescence microscopy, fluorescence resonance energy transfer (FRET), and monitoring VP4-induced cytokine mRNA expression in the presence of anti-TLR2 and anti-VP4. MyrVP4 and TLR2 colocalized in TLR2-expressing HEK-293 cells, mouse bone marrow-derived macrophages, human bronchoalveolar macrophages, and human airway epithelial cells. Colocalization was absent in TLR2-null HEK-293 cells and blocked by anti-TLR2 and anti-VP4. Cy3-labeled MyrVP4 and Cy5-labeled anti-TLR2 showed an average fractional FRET efficiency of 0.24 ± 0.05, and Cy5-labeled anti-TLR2 increased and unlabeled MyrVP4 decreased FRET efficiency. MyrVP4-induced chemokine mRNA expression was higher than that elicited by VP4 alone and was attenuated by anti-TLR2 and anti-VP4. Cytokine expression was similarly increased by MyrVP4 purified from RV-infected HeLa cells and synthetic MyrVP4. We conclude that, during RV infection, MyrVP4 and TLR2 interact to generate a proinflammatory response. |
| Databáze: | OpenAIRE |
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