Members of Tumor Necrosis Factor (TNF) Receptor-Associated Factors (TRAFs) family interact with the cytoplasmic tails of TNF receptor family members to mediate signal transduction processes. TRAF3 has a major immunomodulatory function and TRAF3 deficiency has been linked to malignancies, such as multiple myeloma and lymphoid defects. In order to characterize the molecular mechanisms of TRAF3 signaling, the yeast two-hybrid system was used to identify proteins that interact with TRAF3. The yeast two-hybrid screen of a human B cell cDNA library with TRAF3 as bait, identified Glucocorticoid Modulatory Element-Binding Protein 1 (GMEB1) as a TRAF3-interacting protein. Previous studies indicated that GMEB1 functions as a potent inhibitor of caspase activation and apoptosis. The interaction of TRAF3 and GMEB1 proteins was confirmed in mammalian cells lines, using immunoprecipitation assays. The RING and TRAF-C domains of TRAF3 were not essential for this interaction. The overexpression of TRAF3 protein enhanced the anti-apoptotic function of GMEB1 in HeLa cells. On the other hand, downregulation of TRAF3 by RNA interference decreased significantly the ability of GMEB1 to inhibit apoptosis. In addition, LMP1(1–231), a truncated form of the EBV oncoprotein LMP1, that can interact and oligomerize with TRAF3, was also able to cooperate with GMEB1, in order to inhibit apoptosis. Our protein-interaction experiments demonstrated that TRAF3 can interact with GMEB1, which is an inhibitor of apoptosis. In addition, cell viability assays showed that overexpression of TRAF3 enhanced the anti-apoptotic activity of GMEB1, supporting a regulatory role of TRAF3 in GMEB1-mediated inhibition of apoptosis. Better understanding of the molecular mechanism of TRAF3 function will improve diagnostics and targeted therapeutic approaches for TRAF3-associated disorders.

中文翻译：

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TRAF3可与GMEB1相互作用并调节其抗凋亡功能

肿瘤坏死因子（TNF）受体相关因子（TRAF）家族的成员与TNF受体家族成员的胞质尾相互作用，以介导信号转导过程。TRAF3具有主要的免疫调节功能，而TRAF3缺乏与恶性肿瘤有关，例如多发性骨髓瘤和淋巴样缺陷。为了表征TRAF3信号转导的分子机制，酵母双杂交系统被用来识别与TRAF3相互作用的蛋白质。以TRAF3为诱饵的人B细胞cDNA文库的酵母双杂交筛选确定了糖皮质激素调节元件结合蛋白1（GMEB1）为TRAF3相互作用蛋白。先前的研究表明，GMEB1是caspase激活和凋亡的有效抑制剂。使用免疫沉淀测定法在哺乳动物细胞系中证实了TRAF3和GMEB1蛋白的相互作用。TRAF3的RING和TRAF-C结构域对此相互作用不是必需的。TRAF3蛋白的过表达增强了GMEB1在HeLa细胞中的抗凋亡功能。另一方面，RNA干扰对TRAF3的下调显着降低了GMEB1抑制细胞凋亡的能力。此外，可以与TRAF3相互作用和寡聚的EBV癌蛋白LMP1的截短形式LMP1（1-231）也能够与GMEB1协同作用，以抑制细胞凋亡。我们的蛋白质相互作用实验表明TRAF3可以与细胞凋亡抑制剂GMEB1相互作用。此外，细胞活力分析表明TRAF3的过度表达增强了GMEB1的抗凋亡活性，支持TRAF3在GMEB1介导的凋亡抑制中的调节作用。更好地了解TRAF3功能的分子机制将改善TRAF3相关疾病的诊断和靶向治疗方法。