The N-terminus of PKR is responsible for the activation of the NF-κB signaling pathway by interacting with the IKK complex
The N-terminus of PKR is responsible for the activation of the NF-κB signaling pathway by interacting with the IKK complex
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DOI：
10.1016/j.cellsig.2006.02.010
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发表时间：
2006-11-01
影响因子：
4.8
通讯作者：
Meurs, Eliane F.
中科院分区：
文献类型：
--
作者：
Bonnet, Marion C.;Daurat, Caroline;Meurs, Eliane F.
干扰素诱导的双链RNA（dsRNA）激活的蛋白激酶（PKR）在与IKK复合物相互作用后，已显示独立于其激酶功能激活NF-κ B。为了研究PKR激活NF-κ B的机制，我们使用NF-κ B依赖性报告基因分析鉴定了PKR缺陷成纤维细胞中负责刺激NF-κ B通路的PKR结构域。PKR的N-末端1-265 AA激活NF-κ B，而限制于两个dsRNA结合结构域（DRBD）的1 - 180 AA N-末端、单独的第三基本结构域（AA 181-265）或PKR的C-末端（AA 266-550）不能刺激NF-κ B依赖性报告基因的表达。使用共聚焦显微镜，我们证实PKR全长以及PKR N-末端与IKK β共定位。通过GST-下拉分析，使用不同的PKR结构域，我们揭示了PKR N-末端1-265结合并激活IKK的特异性能力，并表明这种激活需要IKK复合物的完整性。这种激活不仅是由于DRBD，因为DRBD片段1 - 180不能抑制PKR 1-265诱导的NF-κ B激活。因此，我们的研究结果表明，PKR介导NF-κ B激活的能力存在于其完整的N-末端，并需要DRBDs和第三个基本结构域。(c)2006年爱思唯尔公司All rights reserved.
The interferon-induced double-stranded RNA (dsRNA)-activated protein kinase (PKR) has been shown to activate NF-kappa B independently of its kinase function after interaction with the IKK complex. In order to investigate the mechanism of NF-kappa B activation by PKR, we identified the domain of PKR responsible for stimulating the NF-kappa B pathway in PKR-deficient fibroblasts using an NF-kappa B dependent reporter assay. The N-terminal 1-265 AA of PKR activates NF-kappa B, whereas the 1 - 180 AA N-terminus restricted to the two dsRNA Binding Domains (DRBD), the third basic domain alone (AA 181-265), or the C-terminus of PKR (AA 266-550) were unable to stimulate the expression of the NF-kappa B dependent reporter gene. Using confocal microscopy, we confirmed that PKR full length as well as PKR N-terminus colocalized with IKK beta. By GST-pulldown analysis, using different PKR domains, we then revealed the specific ability of the PKR N-terminus 1-265 to bind to and activate IKK and showed that this activation requires the integrity of the IKK complex. This activation is not only due to DRBDs since the DRBD fragment 1 - 180 failed to inhibit PKR 1-265 induced NF-kappa B activation. Our results therefore demonstrate that the ability of PKR to mediate NF-kappa B activation resides in its full N-terminus, and requires both DRBDs and the third basic domain. (c) 2006 Elsevier Inc. All rights reserved.