Grass carp (Ctenopharyngodon idella) PACT induces cell apoptosis and activates NF-кB via PKR.
Grass carp (Ctenopharyngodon idella) PACT induces cell apoptosis and activates NF-кB via PKR.
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DOI：
10.1016/j.fsi.2020.05.036
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发表时间：
2020-05
影响因子：
4.7
通讯作者：
Zhizhen Hu;Hailing Du;Gang Lin;Kun Han;Xining Cheng;Zhiqing Feng;Huiling Mao;Chengyu Hu
中科院分区：
文献类型：
--
作者：
Zhizhen Hu;Hailing Du;Gang Lin;Kun Han;Xining Cheng;Zhiqing Feng;Huiling Mao;Chengyu Hu
PKR是一种依赖于dsrna和干扰素诱导的蛋白激酶，参与多种细胞因子介导的抗病毒免疫应答和细胞凋亡。在哺乳动物细胞中，PKR也可以在缺乏dsRNA的情况下被激活。PKR激活因子PACT （PKR激活蛋白），也称为RAX （PKR相关蛋白X）在其中起着重要作用。近年来，随着对鱼类干扰素系统认识的不断提高，PKR和PACT在鱼类中逐渐被发现。然而，鱼类PACT的功能尚不清楚。在我们之前的工作中，我们认为草鱼（Ctenopharyngodon idella）的PACT一定参与了IRF2和atf4介导的应激反应途径。在本研究中，我们发现c。idellaPACT （CiPACT）和cipkr2在LPS刺激下显著上调。这表明CiPACT和CiPKR可能在LPS刺激反应中起重要作用。另外，cipactto LPS的响应时间要早于cipkr。在CIK细胞中过表达CiPACT可显著提高p-eIF2α水平，诱导细胞凋亡和Cip65从细胞质向细胞核易位。为了进一步了解其机制，我们进行了共免疫沉淀实验。证明CiPACT与CiPKR的相互作用使CiPKR发生磷酸化。过表达CiPACT诱导细胞内bcl-2表达下调，bax表达上调。然而，在CiPKR敲除的细胞中，bcl-2和bax的表达正好相反。因此，鱼类PACT诱导细胞凋亡和激活NF-кB的机制依赖于PKR。
As a dsRNA-dependent and interferon-induced protein kinase, PKR is involved in antiviral immune response and apoptosis mediated by various cytokines. In mammalian cells, PKR can also be activated in the absence of dsRNA. A PKR activator, PACT (PKR activating protein), also referred to as RAX (PKR-associated protein X) plays an important role. In recent years, with the increasing recognition of fish interferon system, PKR and PACT have been gradually revealed in fish. However, the function of fish PACT is unclear. In our previous work, we suggested that grass carp (Ctenopharyngodon idella) PACT must be involved in IRF2 and ATF4-mediated stress response pathways. In the present study, we found that the expression ofC. idellaPACT (CiPACT) andCiPKRwere significantly up-regulated under the stimulation of LPS. It indicated that CiPACT and CiPKR may play an important role in response to LPS stimulation. In addition, the response time ofCiPACTto LPS is earlier than that ofCiPKR. It has also shown that overexpression of CiPACT in CIK cells can significantly enhance the level of p-eIF2α, induces apoptosis and translocation of Cip65 to nucleus from cytoplasm. To further understand the mechanism, we carried out the co-immunoprecipitation assay. It proved that the interaction of CiPACT and CiPKR made the phosphorylation of CiPKR. Overexpression of CiPACT induced the down-regulation of intracellular expression ofbcl-2and up-regulation ofbax.However, in CiPKR knocked-down cells the expression ofbcl-2andbaxwere just the opposite. Therefore, the mechanism of fish PACT induces apoptosis and activates NF-кB is dependent on PKR.