Fibronectin inhibits cytokine production induced by CpG DNA in macrophages without direct binding to DNA
Fibronectin inhibits cytokine production induced by CpG DNA in macrophages without direct binding to DNA
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纤连蛋白抑制巨噬细胞中 CpG DNA 诱导的细胞因子产生，但不直接与 DNA 结合
DOI：
10.1016/j.cyto.2012.06.237
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发表时间：
2012
期刊：
影响因子：
3.8
通讯作者：
Takakura Y
中科院分区：
文献类型：
--
作者：
Yoshida H;Nishikawa M;Yasuda S;Toyota H;Kiyota T;Takahashi Y;Takakura Y
纤连蛋白（FN）已知具有四个DNA结合结构域，尽管它们的生理意义尚不清楚。与小鼠巨噬细胞样细胞系相比，原代小鼠腹腔巨噬细胞对CpG基序填充质粒DNA（pDNA）、Toll样受体-9（TLR 9）配体的反应性显著降低。本研究进行检查是否FN具有DNA结合域参与这种现象。FN在原代巨噬细胞中的表达显著高于巨噬细胞样细胞系RAW264.7，表明丰富的FN可能抑制原代巨噬细胞的反应性。然而，电泳分析表明，FN不结合pDNA的存在下，生理浓度的二价阳离子。令人惊讶的是，在CpG DNA刺激下，外源性FN以浓度依赖性方式显著降低了小鼠巨噬细胞产生的显著肿瘤坏死因子-（TNF-）α，但BSA、层粘连蛋白或胶原蛋白则没有。FN不影响细胞的表观pDNA摄取。此外，FN减少了由polyI：C（TLR 3配体）和咪喹莫特（TLR 7配体）诱导的TNF-α产生，但不减少LPS（TLR 4配体）或非CpG pDNA/阳离子脂质体复合物诱导的TNF-α产生。共聚焦显微镜研究显示，pDNA与FN共定位于RAW264.7的同一细胞内区室中，表明FN抑制内体/溶酶体区室中的细胞因子信号转导。总之，本研究的结果首次揭示了FN的一种新效应，即糖蛋白通过巨噬细胞中的CpG-DNA/TLR 9相互作用调节细胞因子信号转导，而不通过其推定的DNA结合结构域直接结合DNA。
Fibronectin (FN) is known to have four DNA-binding domains although their physiological significance is unknown. Primary murine peritoneal macrophages have been shown to exhibit markedly lower responsiveness to CpG motif-replete plasmid DNA (pDNA), Toll-like receptor-9 (TLR9) ligand, compared with murine macrophage-like cell lines. The present study was conducted to examine whether FN having DNA-binding domains is involved in this phenomenon. The expression of FN was significantly higher in primary macrophages than in a macrophage-like cell line, RAW264.7, suggesting that abundant FN might suppress the responsiveness in the primary macrophages. However, electrophoretic analysis revealed that FN did not bind to pDNA in the presence of a physiological concentration of divalent cations. Surprisingly, marked tumor necrosis factor – (TNF-)α production from murine macrophages upon CpG DNA stimulation was significantly reduced by exogenously added FN in a concentration-dependent manner but not by BSA, laminin or collagen. FN did not affect apparent pDNA uptake by the cells. Moreover, FN reduced TNF-α production induced by polyI:C (TLR3 ligand), and imiquimod (TLR7 ligand), but not by LPS (TLR4 ligand), or a non-CpG pDNA/cationic liposome complex. The confocal microscopic study showed that pDNA was co-localized with FN in the same intracellular compartment in RAW264.7, suggesting that FN inhibits cytokine signal transduction in the endosomal/lysosomal compartment. Taken together, the results of the present study has revealed, for the first time, a novel effect of FN whereby the glycoprotein modulates cytokine signal transduction via CpG-DNA/TLR9 interaction in macrophages without direct binding to DNA through its putative DNA-binding domains.