Takada Y, Ichikawa H, Pataer A, Swisher S, Aggarwal B B
Cytokine Research Laboratory, Department of Experimental Therapeutics, The University of Texas MD Anderson Cancer Center, Houston, TX 77030, USA.
Oncogene. 2007 Feb 22;26(8):1201-12. doi: 10.1038/sj.onc.1209906. Epub 2006 Aug 21.
Double-stranded RNA-dependent protein kinase (PKR), a ubiquitously expressed serine/threonine kinase, has been implicated in the regulation or modulation of cell growth through multiple signaling pathways, but how PKR regulates tumor necrosis factor (TNF)-induced signaling pathways is poorly understood. In the present study, we used fibroblasts derived from PKR gene-deleted mice to investigate the role of PKR in TNF-induced activation of nuclear factor-kappaB (NF-kappaB), mitogen-activated protein kinases (MAPKs) and growth modulation. We found that in wild-type mouse embryonic fibroblast (MEF), TNF induced NF-kappaB activation as measured by DNA binding but deletion of PKR abolished this activation. This inhibition was associated with suppression of inhibitory subunit of NF-kappaB (IkappaB)alpha kinase (IKK) activation, IkappaBalpha phosphorylation and degradation, p65 phosphorylation and nuclear translocation, and NF-kappaB-dependent reporter gene transcription. TNF-induced Akt activation needed for IKK activation was also abolished by deletion of PKR. NF-kappaB activation was diminished in PKR-deleted cells transfected with TNF receptor (TNFR) 1, TNFR-associated death domain and TRAF2 plasmids; NF-kappaB activated by NF-kappaB-inducing kinase, IKK or p65, however, was minimally affected. Among the MAPKs, it was interesting that whereas TNF-induced c-Jun N-terminal kinase (JNK) activation was abolished, activation of p44/p42 MAPK and p38 MAPK was potentiated in PKR-deleted cells. TNF induced the expression of NF-kappaB-regulated gene products cyclin D1, c-Myc, matrix metalloproteinase-9, survivin, X-linked inhibitor-of-apoptosis protein (IAP), IAP1, Bcl-x(L), A1/Bfl-1 and Fas-associated death domain protein-like IL-1beta-converting enzyme-inhibitory protein in wild-type MEF but not in PKR-/- cells. Similarly, TNF induced the proliferation of wild-type cells, but this proliferation was completely suppressed in PKR-deleted cells. Overall, our results indicate that PKR differentially regulates TNF signaling; IKK, Akt and JNK were positively regulated, whereas p44/p42 MAPK and p38 MAPK were negatively regulated.
双链RNA依赖性蛋白激酶（PKR）是一种广泛表达的丝氨酸/苏氨酸激酶，已被证明通过多种信号通路参与细胞生长的调节或调控，但PKR如何调节肿瘤坏死因子（TNF）诱导的信号通路却知之甚少。在本研究中，我们使用来自PKR基因敲除小鼠的成纤维细胞来研究PKR在TNF诱导的核因子-κB（NF-κB）激活、丝裂原活化蛋白激酶（MAPK）激活及生长调节中的作用。我们发现，在野生型小鼠胚胎成纤维细胞（MEF）中，TNF通过DNA结合检测可诱导NF-κB激活，但PKR的缺失消除了这种激活。这种抑制与NF-κB抑制亚基（IkappaB）α激酶（IKK）激活的抑制、IkappaBα磷酸化和降解、p65磷酸化和核转位以及NF-κB依赖性报告基因转录的抑制有关。PKR的缺失也消除了IKK激活所需的TNF诱导的Akt激活。在用TNF受体（TNFR）1、TNFR相关死亡结构域和TRAF2质粒转染的PKR缺失细胞中，NF-κB激活减弱；然而，由NF-κB诱导激酶、IKK或p65激活的NF-κB受影响最小。在MAPK中，有趣的是，虽然TNF诱导的c-Jun氨基末端激酶（JNK）激活被消除，但在PKR缺失细胞中，p44/p42 MAPK和p38 MAPK的激活增强。TNF诱导野生型MEF中NF-κB调节的基因产物细胞周期蛋白D1、c-Myc、基质金属蛋白酶-9、生存素、X连锁凋亡抑制蛋白（IAP）、IAP1、Bcl-x（L）、A1/Bfl-1和Fas相关死亡结构域蛋白样白细胞介素-1β转化酶抑制蛋白的表达，但在PKR -/- 细胞中不诱导。同样，TNF诱导野生型细胞增殖，但这种增殖在PKR缺失细胞中被完全抑制。总体而言，我们的结果表明PKR对TNF信号传导有不同的调节作用；IKK、Akt和JNK被正向调节，而p44/p42 MAPK和p38 MAPK被负向调节。
