Identification and characterization of receptor-interacting protein 2 as a TNFR-associated factor 3 binding partner
Identification and characterization of receptor-interacting protein 2 as a TNFR-associated factor 3 binding partner
复制标题
作为 TNFR 相关因子 3 结合伴侣的受体相互作用蛋白 2 的鉴定和表征
DOI：
10.1016/j.gene.2012.12.026
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发表时间：
2013-04-01
期刊：
影响因子：
3.5
通讯作者：
Xu, Donggang
中科院分区：
文献类型：
--
作者：
Cai, Xin;Du, Jianfang;Xu, Donggang
肿瘤坏死因子受体相关因子3（TRAF 3）是一种高度通用的免疫调节剂，其积极控制I型干扰素的产生，但消极调节丝裂原活化蛋白激酶和替代核因子-κ B信号传导的活化。TRAF 3在不同信号通路中的确切功能尚不清楚。因此，在酵母双杂交试验中，TRAF 3被用作诱饵来筛选人脾cDNA文库中可能潜在介导TRAF 3调节功能的TRAF 3相互作用物。受体相互作用蛋白2（RIP 2）被鉴定为TRAF 3结合伴侣。TRAF 3和RIP 2之间的相互作用通过哺乳动物双杂交、免疫共沉淀和GST pull-down试验进一步证实，并且这种相互作用也通过人B淋巴瘤细胞系拉莫斯细胞中内源性蛋白的免疫沉淀来验证。RIP 2是NF-κ B的激活剂。因此，我们研究了TRAF 3在RIP 2诱导的NF-κ B活化中的作用。结果表明TRAF 3可抑制RIP 2诱导的NF-κ B活化。鉴于RIP 2在B淋巴瘤细胞系中的高表达和TRAF 3与RIP 2在拉莫斯细胞中的内源性相互作用，进一步研究了RIP 2的作用。结果表明，RIP 2基因敲低后，拉莫斯细胞TRAF 3表达增加，NF-κ B B通路激活受到抑制。这些发现表明，RIP 2和TRAF 3之间的功能相互作用可能提供一些线索的机制，TRAF 3参与的积极和消极的调节功能。(C)2012爱思唯尔有限公司版权所有。
Tumor necrosis factor receptor-associated factor 3 (TRAF3) is a highly versatile immune regulator that positively controls type I interferon production, but negatively regulates the activation of mitogen-activated protein kinase and alternative nuclear factor-kappa B signaling. The precise function of TRAF3 in different signaling pathways remains unclear. Thus, in a yeast two-hybrid assay, TRAF3 was used as the bait to screen a human spleen cDNA library for TRAF3 interactors that may potentially mediate TRAF3-regulated functions. Receptor-interacting protein 2 (RIP2) was identified as a TRAF3 binding partner. The interaction between TRAF3 and RIP2 was further confirmed by mammalian two-hybrid, co-immunoprecipitation and GST pull-down assays, and this interaction was also verified by immunoprecipitation of endogenous proteins in Ramos cells, a human B lymphoma cell line. RIP2 is an activator of NF-kappa B. We therefore examined the effect of TRAF3 in RIP2-induced NF-kappa B activation. The result showed that TRAF3 could inhibit RIP2-induced NF-kappa B activation. Given the high expression of RIP2 in the B lymphoma cell line and endogenous interaction between TRAF3 and RIP2 in Ramos cells, the role of RIP2 was further studied. The result demonstrated that RIP2 knockdown was capable of increasing the expression of TRAF3 and suppressing the activation of alternative NF-kappa B pathway in Ramos cells. These findings suggest that functional interactions between RIP2 and TRAF3 may provide some clues to the mechanisms of TRAF3-involvement in both positive and negative regulatory functions. (C) 2012 Elsevier B.V. All rights reserved.