CD14, TLR4 and TRAM Show Different Trafficking Dynamics During LPS Stimulation
CD14, TLR4 and TRAM Show Different Trafficking Dynamics During LPS Stimulation
复制标题
DOI：
10.1111/tra.12274
复制
发表时间：
2015-07-01
期刊：
影响因子：
4.5
通讯作者：
Husebye, Harald
中科院分区：
文献类型：
--
作者：
Klein, Dionne C. G.;Skjesol, Astrid;Husebye, Harald
toll样受体4 （TLR4）通过招募不同的衔接蛋白对，通过两条主要途径对革兰氏阴性菌和信号作出即时反应。Mal- MYD88 [Mal （MYD88 -adaptor-样）- MYD88（髓样分化初级反应基因（Myeloid differentiation primary response gene, 88））]被招募到质膜上，启动导致促炎细胞因子产生的信号级联反应，而TRAM- trif [TRAM （trf相关适配器分子）- trif（含tir结构域的适配器诱导干扰素-）]被招募到早期核内体中，启动随后的I型干扰素产生。我们研究了TLR4和TRAM在脂多糖（LPS）刺激下的动态。我们发现LPS在质膜中诱导了一个依赖cd14的TLR4不动部分。全内反射荧光显微镜（TIRF）显示，在HEK293细胞和巨噬细胞模型细胞系U373-CD14中，LPS刺激诱导TLR4在含有CD14/LPS和网格蛋白的质膜中聚集成小的点状结构。这些结果表明，横向固定的TLR4受体复合物正在形成并准备内吞作用。RAB11A被发现参与将TRAM定位到内噬循环室（ERC）和早期分选核内体。此外，CD14/LPS固定在rab11a阳性的核内体上，而TRAM没有固定在核内体上，这表明TRAM和CD14/LPS可以独立被募集到核内体上。
Toll-like receptor 4 (TLR4) is responsible for the immediate response to Gram-negative bacteria and signals via two main pathways by recruitment of distinct pairs of adaptor proteins. Mal-MyD88 [Mal (MyD88-adaptor-like) - MYD88 (Myeloid differentiation primary response gene (88))] is recruited to the plasma membrane to initiate the signaling cascade leading to production of pro-inflammatory cytokines while TRAM-TRIF [TRAM (TRIF-related adaptor molecule)-TRIF (TIR-domain-containing adapter-inducing interferon-)] is recruited to early endosomes to initiate the subsequent production of type I interferons. We have investigated the dynamics of TLR4 and TRAM during lipopolysaccharide (LPS) stimulation. We found that LPS induced a CD14-dependent immobile fraction of TLR4 in the plasma membrane. Total internal reflection fluorescence microscopy (TIRF) revealed that LPS stimulation induced clustering of TLR4 into small punctate structures in the plasma membrane containing CD14/LPS and clathrin, both in HEK293 cells and the macrophage model cell line U373-CD14. These results suggest that laterally immobilized TLR4 receptor complexes are being formed and prepared for endocytosis. RAB11A was found to be involved in localizing TRAM to the endocytic recycling compartment (ERC) and to early sorting endosomes. Moreover, CD14/LPS but not TRAM was immobilized on RAB11A-positive endosomes, which indicates that TRAM and CD14/LPS can independently be recruited to endosomes.