Lo Y Y, Luo L, McCulloch C A, Cruz T F
Samuel Lunenfeld Research Institute, Mount Sinai Hospital, Toronto, Ontario M5G 1X5, Canada.
J Biol Chem. 1998 Mar 20;273(12):7059-65. doi: 10.1074/jbc.273.12.7059.
Interleukin-1 (IL-1) is an important inflammatory mediator and plays a central role in the destruction of connective tissue matrices in diseases such as arthritis and periodontitis. It is well established that IL-1 activation of the mitogen-activated protein (MAP) kinase pathway and induction of c-fos expression is a required step in the induction of matrix metalloproteinase expression involved in tissue degradation. Previous studies in our laboratory showed that IL-1-induced calcium flux is dependent on focal adhesion formation, suggesting a matrix-dependent restriction system for IL-1 signaling. Therefore, in the present study, we examined the consequences of this restriction on IL-1-mediated activation of the MAP kinase family and on c-fos expression. Treatment of human gingival fibroblasts with IL-1 activated extracellular signal-regulated kinases (ERK), c-Jun N-terminal kinase (JNK), and p38 kinase activity and induced c-fos expression in a dose- and time-dependent fashion. Plating cells on poly-L-lysine prevented focal adhesion formation, eliminated IL-1-induced calcium influx, abolished ERK stimulation, and blocked c-fos expression. Cells in suspension and hence with no suitable substratum for focal adhesion formation also showed no ERK activation or enhanced c-fos expression in response to IL-1. In contrast, eliminating focal adhesion formation or calcium depletion in cells plated on fibronectin had no effect on IL-1 stimulation of JNK and p38 kinases, demonstrating that their activation was mediated through pathways independent of focal adhesions and calcium. Calcium depletion abolished IL-1-induced calcium uptake, ERK activation, and c-fos expression. The focal adhesion dependence of IL-1-induced ERK activation and c-fos expression could be circumvented in cells plated on poly-L-lysine by simultaneous incubation with IL-1 and the calcium ionophore ionomycin. In transfection studies, IL-1 stimulation of serum responsive element (SRE) transcriptional activity was dependent on the presence of extracellular calcium. This is consistent with a requirement for calcium in the activation of ERKs and their involvement in the induction of c-fos expression through the SRE site on the 5' promoter of the c-fos gene. Our results demonstrate that in cells attached to substrates by focal adhesions, IL-1-mediated calcium flux is required for ERK activation and c-fos expression but not for JNK or p38 activation. We conclude that cellular interactions with the extracellular matrix play an important role in restricting ERK and c-fos-dependent processes.
白细胞介素-1（IL-1）是一种重要的炎症介质，在关节炎和牙周炎等疾病中结缔组织基质的破坏过程中起核心作用。众所周知，IL-1激活丝裂原活化蛋白（MAP）激酶途径并诱导c-fos表达是诱导参与组织降解的基质金属蛋白酶表达的必要步骤。我们实验室先前的研究表明，IL-1诱导的钙流依赖于粘着斑的形成，这表明存在一种针对IL-1信号传导的基质依赖性限制系统。因此，在本研究中，我们研究了这种限制对IL-1介导的MAP激酶家族激活和c-fos表达的影响。用IL-1处理人牙龈成纤维细胞可激活细胞外信号调节激酶（ERK）、c-Jun氨基末端激酶（JNK）和p38激酶活性，并以剂量和时间依赖性方式诱导c-fos表达。将细胞接种在聚-L-赖氨酸上可防止粘着斑形成，消除IL-1诱导的钙内流，消除ERK刺激，并阻断c-fos表达。悬浮培养的细胞因此没有适合粘着斑形成的基质，对IL-1也未表现出ERK激活或c-fos表达增强。相反，消除接种在纤连蛋白上的细胞中的粘着斑形成或钙耗竭对IL-1刺激JNK和p38激酶没有影响，表明它们的激活是通过独立于粘着斑和钙的途径介导的。钙耗竭消除了IL-1诱导的钙摄取、ERK激活和c-fos表达。在接种于聚-L-赖氨酸的细胞中，通过同时与IL-1和钙离子载体离子霉素孵育，可以规避IL-1诱导的ERK激活和c-fos表达对粘着斑的依赖性。在转染研究中，IL-1对血清反应元件（SRE）转录活性的刺激依赖于细胞外钙的存在。这与激活ERK需要钙以及它们通过c-fos基因5'启动子上的SRE位点参与诱导c-fos表达是一致的。我们的结果表明，在通过粘着斑附着于底物的细胞中，IL-1介导的钙流是ERK激活和c-fos表达所必需的，但不是JNK或p38激活所必需的。我们得出结论，细胞与细胞外基质的相互作用在限制ERK和c-fos依赖性过程中起重要作用。
