Identification and Immune Functional Characterization of Pigeon TLR7

Shizhong Geng,Xinan Jiao,Dan Xiong,Zhiming Pan,Xiang Chen,Li Song

doi:10.3390/ijms16048364

Shizhong Geng, Xinan Jiao + Show 4 more

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https://doi.org/10.3390/ijms16048364

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Abstract

Toll-like receptor 7 (TLR7) is activated by single-stranded RNA and synthetic imidazoquinoline components, and induces interferon production. In this study, we cloned the TLR7 gene from King pigeon (Columba livia). The TLR7 open reading frame is 3144 bp and encodes a 1047-amino acid protein, consisting of a canonical TLR composition with 15 leucine-rich repeats (LRRs). Amino acid-inserting modifications were found at position 15 of LRR2, LRR11, LRR13, and LRR14 and position 10 of LRR10. The tissue distribution of pigeon TLR7 suggests that immune-associated tissues, especially the spleen and liver, have high TLR7 expression. HEK293T cells transfected with pigeon TLR7 plasmid responded to the agonist R848, indicating a functional TLR7 homolog. Following R848 stimulation of pigeon peripheral blood mononuclear cells, the levels of IFN-γ, IL-6, IL-8, CCL5, and IL-10 mRNA, assessed using quantitative real-time PCR, were significantly up-regulated. After Newcastle disease virus vaccine strain LaSota inoculation and agonist R848 injection, the level of TLR7 mRNA in the spleen of pigeons increased significantly in the R848-injected group, but decreased in the LaSota-inoculated group at three day post-infection (d.p.i.). The mRNA levels of inflammatory cytokines and chemokines were significantly upregulated in both LaSota-inoculated and R848-injected groups. Triggering pigeon TLR7 leads to robust up-regulation of inflammatory cytokines and chemokines, suggesting an important role in the innate immune response.

Highlights

The innate immune system acts as the front line of defense against invading microorganisms in most multicellular organisms by recognizing pathogen-associated molecular patterns (PAMPs) through pattern recognition receptors (PRRs) [1]
Sequencing results show the pigeon Toll-like receptor 7 (TLR7) gene contains an open reading frame (ORF) of 3144 bp encoding a protein of 1047 amino acids (Figure 1)
The deduced amino acid sequence of pigeon TLR7 was aligned with reported sequences from human (GenBank ID: BAG55056), mouse (GenBank ID: AAK62676), chicken (GenBank ID: CAF28878), and duck (GenBank ID: ABK51522) using ClustalW

Summary

Introduction

The innate immune system acts as the front line of defense against invading microorganisms in most multicellular organisms by recognizing pathogen-associated molecular patterns (PAMPs) through pattern recognition receptors (PRRs) [1]. Toll-like receptors (TLRs) form part of an ancient receptor family of PRRs and are the major cell-surface initiators of inflammatory responses to invading pathogens [2]. TLRs bind a wide variety of pathogen-associated substances through their ectodomains [6]. The basic framework of TLRs is a horseshoe-shaped solenoid, or “m”-shaped architecture that contains an extensive β-sheet on its concave surface, and numerous ligand-binding insertions [7]. These surfaces are responsible for ligand binding and ligand-induced TLR dimerization [8]

Results

Discussion

Conclusion