Double Plant Homeodomain (PHD) Finger Proteins DPF3a and-3b Are Required as Transcriptional Co-activators in SWI/SNF Complex-dependent Activation of NF-κB RelA/p50 Heterodimer
Double Plant Homeodomain (PHD) Finger Proteins DPF3a and-3b Are Required as Transcriptional Co-activators in SWI/SNF Complex-dependent Activation of NF-κB RelA/p50 Heterodimer
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DOI：
10.1074/jbc.m111.322792
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发表时间：
2012-04-06
影响因子：
4.8
通讯作者：
Iba, Hideo
中科院分区：
文献类型：
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作者：
Ishizaka, Aya;Mizutani, Taketoshi;Iba, Hideo
我们以前已经表明，DPF 2（安魂曲/安魂曲）作为SWI/SNF复合物和RelB/p52 NF-κ B异源二聚体之间的连接蛋白发挥作用，并通过其非经典途径在NF-κ B反式激活中发挥重要作用。利用整合了SWI/SNF依赖性NF-κ B报告基因的敏感293 FT报告细胞克隆，我们在这项研究中发现，DPF 1、DPF 2、DPF 3a、DPF 3 B和PHF 10的过表达显著增强了典型NF-κ B二聚体的反式激活活性。使用低水平内源性表达这五种蛋白质的293 FT报告细胞的敲低分析清楚地表明，通过选择性剪接从DPF 3基因产生的DPF 3 a和DPF 3 B b对于TNF-α刺激诱导的RelA/p50 NF-κ B异二聚体反式激活是最关键的。我们的数据进一步表明，这种反式激活需要SWI/SNF复合物。DPF 3a和DPF 3b还显示在体外与RelA、p50和SWI/SNF复合物的几个亚基直接相互作用，并与RelA/p50和来自用TNF-α处理的细胞的核部分的SWI/SNF复合物共免疫沉淀。在ChIP实验中，我们进一步发现内源性DPF 3a/B和SWI/SNF复合物连续存在于HIV-1 LTR上，而TNF-α处理后RelA/p50募集的动力学与病毒转录激活水平良好相关。此外，再ChIP实验显示，TNF-α处理后，DPF 3a/B和SWI/SNF复合物与内源性IL-6启动子上的RelA缔合。总之，我们目前的数据表明，通过将RelA/p50连接到SWI/SNF复合物，DPF 3a/B诱导NF-κ B靶基因启动子在相对不活跃的染色质环境中的反式激活。
We have previously shown that DPF2 (requiem/REQ) functions as a linker protein between the SWI/SNF complex and RelB/p52 NF-kappa B heterodimer and plays important roles in NF-kappa B transactivation via its noncanonical pathway. Using sensitive 293FT reporter cell clones that had integrated a SWI/SNF-dependent NF-kappa B reporter gene, we find in this study that the overexpression of DPF1, DPF2, DPF3a, DPF3b, and PHF10 significantly potentiates the transactivating activity of typical NF-kappa B dimers. Knockdown analysis using 293FT reporter cells that endogenously express these five proteins at low levels clearly showed that DPF3a and DPF3b, which are produced from the DPF3 gene by alternative splicing, are the most critical for the RelA/p50 NF-kappa B heterodimer transactivation induced by TNF-alpha stimulation. Our data further show that this transactivation requires the SWI/SNF complex. DPF3a and DPF3b are additionally shown to interact directly with RelA, p50, and several subunits of the SWI/SNF complex in vitro and to be co-immunoprecipitated with RelA/p50 and the SWI/SNF complex from the nuclear fractions of cells treated with TNF-alpha. In ChIP experiments, we further found that endogenous DPF3a/b and the SWI/SNF complex are continuously present on HIV-1 LTR, whereas the kinetics of RelA/p50 recruitment after TNF-alpha treatment correlate well with the viral transcriptional activation levels. Additionally, re-ChIP experiments showed DPF3a/b and the SWI/SNF complex associate with RelA on the endogenous IL-6 promoter after TNF-alpha treatment. In conclusion, our present data indicate that by linking RelA/p50 to the SWI/SNF complex, DPF3a/b induces the transactivation of NF-kappa B target gene promoters in relatively inactive chromatin contexts.