Casein kinase II-mediated phosphorylation of NF-κB p65 subunit enhances inducible nitric-oxide synthase gene transcription in vivo
Casein kinase II-mediated phosphorylation of NF-κB p65 subunit enhances inducible nitric-oxide synthase gene transcription in vivo
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DOI：
10.1074/jbc.m313731200
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发表时间：
2004-06-04
影响因子：
4.8
通讯作者：
Reveneau, S
中科院分区：
文献类型：
--
作者：
Chantôme, A;Pance, A;Reveneau, S
诱导型一氧化氮合酶 (NOSII) 产生的一氧化氮 (NO) 主要在转录水平上受到核因子 kappaB (NF-kappaB) 的调节。在本研究中，我们通过比较从EMT-6小鼠乳腺癌细胞系中分离的两个克隆，进一步分析了NF-kappaB在NOSII基因体内转录调控中的作用。响应白细胞介素 (IL)-1beta 或脂多糖 (LPS)，EMT-6 克隆 J (EMT-6J) 细胞产生的 NO 比 EMT-6 克隆 H (EMT-6H) 细胞多 3 倍，这一效应与 EMT-6J 细胞中 NF-kappaB 激活增强相关。响应 IL-1beta，NF-kappaB 抑制剂 IkappaB-alpha 和 IkappaB-beta 的降解动力学、转录因子的核质穿梭及其与特定 DNA 序列的结合在两个克隆中相似。相反，在 EMT-6J 细胞中观察到 IL-1β 诱导的 NF-kappaB p65 亚基丝氨酸残基磷酸化，但在 EMT-6H 细胞中未观察到。用酪蛋白激酶 II 抑制剂 DRB 预处理 EMT-6J 细胞可特异性阻止这种 IL-1β 诱导的 p65 磷酸化。小干扰RNA介导的酪蛋白激酶II-α亚基的消耗也将IL-1β和LPS刺激的EMT-6J细胞中的NF-kappaB转录活性和NOSII基因转录降低至在相同条件下处理的EMT-6H细胞中观察到的水平。总之，这些数据表明酪蛋白激酶 II 介导的 p65 亚基磷酸化可以增强体内 NF-kappaB 的转录活性。转录因子的这种翻译后修饰可能导致暴露于 IL-1beta 或 LPS 的肿瘤细胞中 NOSII 基因转录和 NO 产生增加。
Nitric oxide ( NO) produced by inducible nitric-oxide synthase (NOSII) is mainly regulated at the transcriptional level by the nuclear factor-kappaB (NF-kappaB). In the present study, we further analyzed the role of NF-kappaB in the in vivo transcriptional regulation of NOSII gene by comparing two clones isolated from the EMT-6 mouse mammary cancer cell line. In response to interleukin (IL)-1beta or lipopolysaccharide (LPS), EMT-6 clone J (EMT-6J) cells produce 3-fold more NO than EMT-6 clone H (EMT-6H) cells, an effect correlated with enhanced activation of NF-kappaB in EMT-6J cells. In response to IL-1beta, the kinetics of degradation of NF-kappaB inhibitors IkappaB-alpha and IkappaB-beta, the nucleo-cytoplasmic shuttling of the transcription factor and its binding to a specific DNA sequence were similar in both clones. In contrast, an IL-1beta-induced phosphorylation of serine residues in NF-kappaB p65 subunit was observed in EMT-6J, but not in EMT-6H, cells. This IL-1beta-induced phosphorylation of p65 was specifically prevented by pretreatment of EMT-6J cells with the casein kinase II inhibitor DRB. Small interfering RNA-mediated depletion of casein kinase II-alpha subunit also decreased NF-kappaB transcriptional activity and NOSII gene transcription in IL-1beta and LPS-stimulated EMT-6J cells to the levels observed in EMT-6H cells treated in the same conditions. Altogether, these data indicate that casein kinase II-mediated phosphorylation of p65 subunit can enhance the transcriptional activity of NF-kappaB in vivo. This post-translational modification of the transcription factor can be responsible for increased NOSII gene transcription and NO production in tumor cells exposed to either IL-1beta or LPS.