Pharmazentrum Frankfurt/ZAFES, Institute of Clinical Pharmacology, Goethe-University Frankfurt, Theodor Stern Kai 7, 60590 Frankfurt/Main, Germany.
Pharmazentrum Frankfurt/ZAFES, Institute of Clinical Pharmacology, Goethe-University Frankfurt, Theodor Stern Kai 7, 60590 Frankfurt/Main, Germany.
Cell Signal. 2012 Feb;24(2):460-467. doi: 10.1016/j.cellsig.2011.09.025. Epub 2011 Oct 1.
Dimethylcelecoxib, a non-COX-2 inhibiting derivative of celecoxib, inhibits PGE(2) synthesis by transcriptional inhibition of mPGES-1. Previously we demonstrated that DMC downregulates EGR1 expression and increases nuclear NF-κB in human cervical cancer cells (HeLa). Both transcription factors are important regulators of mPGES-1 expression. Here we show that treatment of HeLa cells with DMC inhibits EGR1 promoter activity by influencing the transactivation activity of NF-κB. Mutation of the NF-κB motif as well as downregulation of NF-κB(p65)RelA using siRNA repealed the inhibitory effect of DMC on the EGR1 promoter. The transactivation activity of NF-κB is regulated by various co-activators or co-repressors. One of these co-repressors is HDAC1. DMC did not influence HDAC1 expression, but the HDAC activity was enhanced under DMC influence. After DMC treatment NF-κB co-immunoprecipitated with HDAC1. Electromobility shift assays depicted an increased interaction between NF-κB-HDAC1 and DNA containing NF-κB binding motives. Performing CHIP-assays we finally demonstrated the interaction of NF-κB and HDAC1 at the EGR1 promoter that was in part reversed by the HDAC1 inhibitor trichostatin A. Using siRNA against HDAC1 we could repeal the inhibitory effect of DMC on the EGR1 promoter. In conclusion we demonstrated that treatment of HeLa cells with DMC leads to an enhanced formation of a complex consisting of NF-κB and HDAC1 that binds to the EGR1 promoter resulting in downregulation of EGR1 expression which plays a major role for transcriptional inhibition of mGPES-1 expression. How these effects of DMC may contribute to a potential therapeutical benefit of various diseases is discussed.
二甲噻嗪，塞来昔布的非 COX-2 抑制衍生物，通过转录抑制 mPGES-1 抑制 PGE（2）的合成。先前我们证明 DMC 通过下调 EGR1 表达和增加人宫颈癌（HeLa）细胞中的核 NF-κB 来下调 DMC。这两种转录因子都是 mPGES-1 表达的重要调节剂。在这里，我们表明，用 DMC 处理 HeLa 细胞通过影响 NF-κB 的反式激活活性来抑制 EGR1 启动子的活性。NF-κB 基序的突变以及使用 siRNA 下调 NF-κB(p65)RelA 逆转了 DMC 对 EGR1 启动子的抑制作用。NF-κB 的反式激活活性受各种共激活因子或共抑制因子的调节。其中一个共抑制因子是 HDAC1。DMC 不影响 HDAC1 的表达，但在 DMC 影响下，HDAC 活性增强。在 DMC 处理后，NF-κB 与 HDAC1 共免疫沉淀。电泳迁移率变动分析显示，NF-κB-HDAC1 与含有 NF-κB 结合基序的 DNA 之间的相互作用增加。进行 CHIP 分析后，我们最终证明了 NF-κB 和 HDAC1 在 EGR1 启动子上的相互作用，该作用部分被 HDAC1 抑制剂曲古抑菌素 A 逆转。使用针对 HDAC1 的 siRNA，我们可以逆转 DMC 对 EGR1 启动子的抑制作用。总之，我们证明了用 DMC 处理 HeLa 细胞会导致 NF-κB 和 HDAC1 形成复合物的能力增强，该复合物与 EGR1 启动子结合，导致 EGR1 表达下调，这对 mGPES-1 表达的转录抑制起主要作用。讨论了 DMC 的这些作用如何有助于各种疾病的潜在治疗益处。
