Hambleton J, Weinstein S L, Lem L, DeFranco A L
Cancer Research Insitute, Department of Medicine, G.W. Hooper Foundation, San Francisco, CA 94143, USA.
Proc Natl Acad Sci U S A. 1996 Apr 2;93(7):2774-8. doi: 10.1073/pnas.93.7.2774.
Activation of macrophages by bacterial lipopolysaccharide (LPS) induces transcription of genes that encode for proinflammatory regulators of the immune response. Previous work has suggested that activation of the transcription factor activator protein 1 (AP-1) is one LPS-induced event that mediates this response. Consistent with this notion, we found that LPS stimulated AP-1-mediated transcription of a transfected reporter gene in the murine macrophage cell line RAW 264.7. As AP-1 activity is regulated in part by activation of the c-Jun N-terminal kinase (JNK), which phosphorylates and subsequently increases the transcriptional activity of c-Jun, we examined whether LPS treatment of macrophages resulted in activation of this kinase. LPS treatment of RAW 264.7 cells, murine bone marrow-derived macrophages, and the human monocyte cell line THP-1 resulted in rapid activation of the p46 and p54 isoforms of JNK. Treatment with wild-type and rough mutant forms of LPS and synthetic lipid A resulted in JNK activation, while pretreatment with the tyrosine kinase inhibitor herbimycin A inhibited this response. Binding of LPS-LPS binding protein (LBP) complexes to CD14, a surface receptor that mediates many LPS responses, was found to be crucial, as pretreatment of THP-1 cells with the monoclonal antibody 60b, which blocks this binding, inhibited JNK activation. These results suggest that LPS activation of JNK in monocyte/macrophage cells is a CD14- and protein tyrosine phosphorylation-dependent event that may mediate the early activation of AP-1 in regulating LPS-triggered gene induction.
细菌脂多糖（LPS）激活巨噬细胞可诱导编码免疫反应促炎调节因子的基因转录。先前的研究表明，转录因子激活蛋白1（AP-1）的激活是介导这种反应的LPS诱导事件之一。与此观点一致，我们发现LPS刺激了小鼠巨噬细胞系RAW 264.7中转染的报告基因的AP-1介导的转录。由于AP-1活性部分受c-Jun N端激酶（JNK）激活的调节，JNK使c-Jun磷酸化并随后增加其转录活性，因此我们研究了LPS处理巨噬细胞是否会导致该激酶的激活。用LPS处理RAW 264.7细胞、小鼠骨髓来源的巨噬细胞和人单核细胞系THP-1会导致JNK的p46和p54亚型快速激活。用野生型和粗糙突变型LPS以及合成脂质A处理会导致JNK激活，而用酪氨酸激酶抑制剂赫伯霉素A预处理会抑制这种反应。发现LPS-LPS结合蛋白（LBP）复合物与CD14（介导许多LPS反应的表面受体）的结合至关重要，因为用阻断这种结合的单克隆抗体60b预处理THP-1细胞会抑制JNK激活。这些结果表明，单核细胞/巨噬细胞中LPS对JNK的激活是一个依赖CD14和蛋白酪氨酸磷酸化的事件，可能在调节LPS触发的基因诱导中介导AP-1的早期激活。
