Abstract：
TNF receptor-associated factor 2 (TRAF2) is involved in different cellular processes including signal transduction and transcription regulation. We here provide evidence of a direct interaction between the TRAF domain of TRAF2 and the monosialotetrahexosylganglioside (GM1). Previously, we showed that the TRAF domain occurs mainly in a trimeric form in solution, but it can also exist as a stable monomer when in the nanomolar concentration range. Here, we report that the quaternary structure of the TRAF domain is also affected by pH changes, since a weakly acidic pH (5.5) favors the dissociation of the trimeric TRAF domain into stable monomers, as previously observed at neutral pH (7.6) with the diluted protein. The TRAF domain-GM1 binding was similar at pH 5.5 and 7.6, suggesting that GM1 interacts with both the trimeric and monomeric forms of the protein. However, only the monomeric protein appeared to cause membrane deformation and inward vesiculation in GM1-containing giant unilamellar vesicles (GUVs). The formation of complexes between GM1 and TRAF2, or its TRAF domain, was also observed in cultured human leukemic HAP1 cells expressing either the truncated TRAF domain or the endogenous full length TRAF2. The GM1-protein complexes were observed after treatment with tunicamycin and were more concentrated in cells undergoing apoptosis, a condition which is known to cause cytoplasm acidification. These findings open the avenue for future studies aimed at deciphering the physiopathological relevance of the TRAF domain-GM1 interaction.
摘要：
TNF受体相关因子2（TRAF2）参与不同的细胞过程，包括信号转导和转录调节。我们在此提供TRAF2的TRAF结构域与单唾液酸四己糖神经节苷脂（GM1）之间直接相互作用的证据。以前,我们发现TRAF结构域在溶液中主要以三聚体形式出现，但当在纳摩尔浓度范围内，它也可以作为稳定的单体存在。这里,我们报道TRAF结构域的四级结构也受pH变化的影响，由于弱酸性pH（5.5）有利于三聚体TRAF结构域解离成稳定的单体，如先前用稀释的蛋白质在中性pH(7.6)下观察到的。TRAF结构域-GM1结合在pH5.5和7.6时相似,表明GM1与蛋白质的三聚体和单体形式相互作用。然而,在含有GM1的巨大单层囊泡（GUV）中，只有单体蛋白似乎会引起膜变形和向内的囊泡形成。GM1和TRAF2或其TRAF结构域之间的复合物的形成，在表达截短的TRAF结构域或内源性全长TRAF2的培养的人白血病HAP1细胞中也观察到。衣霉素处理后观察到GM1-蛋白复合物，并且在经历凋亡的细胞中更集中，已知引起细胞质酸化的条件。这些发现为未来研究开辟了道路，旨在破译TRAF域-GM1相互作用的生理病理学相关性。
