Department of Biochemistry and Molecular Biology, Virginia Commonwealth University, Richmond, Virginia 23298, USA.
J Biol Chem. 2011 Dec 16;286(50):42808-17. doi: 10.1074/jbc.M111.310169. Epub 2011 Oct 18.
Tumor necrosis factor α (TNFα) is a well known cytokine involved in systemic and acute inflammation. In this study, we demonstrate that ceramide 1-phosphate (C1P) produced by ceramide kinase (CERK) is a negative regulator of LPS-induced TNFα secretion. Specifically, bone marrow-derived macrophages isolated from CERK knock-out mice (CERK(-/-)) generated higher levels of TNFα than the wild-type mice (CERK(+/+)) in response to LPS. An increase in basal TNFα secretion was also observed in CERK(-/-) murine embryonic fibroblasts, which was rescued by re-expression of wild-type CERK. This effect was due to increased secretion and not transcription. The secretion of TNFα is regulated by TNFα-converting enzyme (TACE also known as ADAM17), and importantly, the activity of TACE was higher in cell extracts from CERK(-/-) as compared with wild type. In vitro analysis also demonstrated that C1P is a potent inhibitor of this enzyme, in stark contrast to ceramide and sphingosine 1-phosphate. Furthermore, TACE specifically bound C1P with high affinity. Finally, several putative C1P-binding sites were identified via homology throughout the protein sequence of TACE. These results indicate that C1P produced by CERK has a negative effect on the processing/secretion of TNFα via modulation of TACE activity.
肿瘤坏死因子-α（TNFα）是一种众所周知的细胞因子，参与全身和急性炎症。在这项研究中，我们证明了由神经酰胺激酶（CERK）产生的神经酰胺 1-磷酸（C1P）是脂多糖诱导的 TNFα分泌的负调节剂。具体来说，与野生型小鼠（CERK(+/+)）相比，来自 CERK 敲除小鼠（CERK(-/-)）的骨髓来源的巨噬细胞在受到 LPS 刺激时产生更高水平的 TNFα。在 CERK(-/-)鼠胚胎成纤维细胞中也观察到基础 TNFα分泌增加，这可以通过重新表达野生型 CERK 来挽救。这种效应是由于分泌增加而不是转录增加所致。TNFα的分泌受 TNFα转化酶（也称为 ADAM17）的调节，重要的是，与野生型相比，CERK(-/-)细胞提取物中的 TACE 活性更高。体外分析还表明，C1P 是该酶的有效抑制剂，与神经酰胺和鞘氨醇 1-磷酸形成鲜明对比。此外，TACE 特异性地与 C1P 结合具有高亲和力。最后，通过 TACE 蛋白序列的同源性鉴定出几个假定的 C1P 结合位点。这些结果表明，CERK 产生的 C1P 通过调节 TACE 活性对 TNFα的加工/分泌产生负面影响。
