Synthetic Human TLR9-LRR11 Peptide Attenuates TLR9 Signaling by Binding to and thus Decreasing Internalization of CpG Oligodeoxynucleotides
Synthetic Human TLR9-LRR11 Peptide Attenuates TLR9 Signaling by Binding to and thus Decreasing Internalization of CpG Oligodeoxynucleotides
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DOI：
10.3390/ijms17020242
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发表时间：
2016-02-01
影响因子：
5.6
通讯作者：
Zhou, Hong
中科院分区：
文献类型：
--
作者：
Pan, Xichun;Li, Bin;Zhou, Hong
Toll样受体（TLR）9是一种识别细菌DNA/CpG ODN的内体受体。阻断CpG ODN/TLR 9活性代表了治疗性预防免疫系统过度激活的策略。在此，我们报告了一种代表人TLR 9胞外区富含亮氨酸重复序列11亚区的合成肽（SP）可以通过与CpG ODN结合并降低其内化来减弱CpG ODN/TLR 9在RAW 264.7细胞中的活性。我们的研究结果表明，与SP预孵育特异性抑制CpG ODN-而不是脂多糖（LPS）-和脂肽（PAM 3CSK 4）-刺激的TNF-和IL-6的释放。SP与CpG ODN的预孵育剂量依赖性地降低TLR 9驱动的IB和ERK磷酸化以及NF-B/p65的活化。此外，SP剂量依赖性地降低FAM标记的CpG ODN的内化，而未标记的CpG ODN逆转抑制。SP-CpG ODN结合的K-D值在微摩尔范围内。我们的研究结果表明，SP是一种特异性的CpG ODN/TLR 9活性的抑制剂，通过结合CpG ODN，导致减少ODN的内化和减少细胞内后续途径的激活。因此，SP可作为一种潜在的CpG ODN拮抗剂来阻断TLR 9信号转导。
Toll-like receptor (TLR) 9 is an endosomal receptor recognizing bacterial DNA/CpG-containing oligodeoxynucleotides (CpG ODN). Blocking CpG ODN/TLR9 activity represents a strategy for therapeutic prevention of immune system overactivation. Herein, we report that a synthetic peptide (SP) representing the leucine-rich repeat 11 subdomain of the human TLR9 extracellular domain could attenuate CpG ODN/TLR9 activity in RAW264.7 cells by binding to CpG ODN and decreasing its internalization. Our results demonstrate that preincubation with SP specifically inhibited CpG ODN- but not lipopolysaccharide (LPS)- and lipopeptide (PAM3CSK4)-stimulated TNF- and IL-6 release. Preincubation of SP with CpG ODN dose-dependently decreased TLR9-driven phosphorylation of IB and ERK and activation of NF-B/p65. Moreover, SP dose-dependently decreased FAM-labeled CpG ODN internalization, whereas non-labeled CpG ODN reversed the inhibition. The K-D value of SP-CpG ODN binding was within the micromolar range. Our results demonstrated that SP was a specific inhibitor of CpG ODN/TLR9 activity via binding to CpG ODN, leading to reduced ODN internalization and decreased activation of subsequent pathways within cells. Thus, SP could be used as a potential CpG ODN antagonist to block TLR9 signaling.