Structural/functional studies of Trio provide insights into its configuration and show that conserved linker elements enhance its activity for Rac1.
Structural/functional studies of Trio provide insights into its configuration and show that conserved linker elements enhance its activity for Rac1.
复制标题
DOI：
10.1016/j.jbc.2022.102209
复制
发表时间：
2022-08
影响因子：
4.8
通讯作者：
Tesmer, John J. G.
中科院分区：
文献类型：
--
作者：
Bandekar, Sumit J.;Chen, Chun-Liang;Ravala, Sandeep K.;Cash, Jennifer N.;V. Avramova, Larisa;V. Zhalnina, Mariya;Gutkind, J. Silvio;Li, Sheng;Tesmer, John J. G.
Trio是一种大型且高度保守的后生动物信号传导支架，其含有两个Dbl家族鸟嘌呤核苷酸交换因子（GEF）模块TrioN和TrioC，分别对Rac和RhoA GTP酶具有选择性。TrioN和TrioC的GEF活性与几种癌症，特别是葡萄膜黑色素瘤有关。然而，很少有人知道这些模块如何在更大的Trio片段的上下文中运行。在这里，我们通过负染电子显微镜显示，Trio的N-末端区域被延长，因此可以作为N-末端推定的脂质结合结构域和TrioN之间的刚性间隔区，而Trio的C-末端的一半似乎是球状的。我们发现TrioN的C端区域增强了其Rac 1 GEF活性，因此可以发挥调节作用。我们继续使用低温电子显微镜和氢-氘交换质谱法表征具有增强活性的最小的、表现良好的Trio片段Trio 1284 -1959与Rac 1复合，并发现赋予增强活性的区域是无序的。删除两个不同的强烈保守的图案在这个区域消除了这种增强，这表明它们形成短暂的分子内相互作用，促进GEF活性。由于Dbl家族RhoGEF模块直接靶向小分子一直具有挑战性，因此诸如这些的辅助Trio结构域的表征可以为开发抑制人类癌症中Trio活性的治疗剂提供替代途径。
Trio is a large and highly conserved metazoan signaling scaffold that contains two Dbl family guanine nucleotide exchange factor (GEF) modules, TrioN and TrioC, selective for Rac and RhoA GTPases, respectively. The GEF activities of TrioN and TrioC are implicated in several cancers, especially uveal melanoma. However, little is known about how these modules operate in the context of larger fragments of Trio. Here we show via negative stain electron microscopy that the N-terminal region of Trio is extended and could thus serve as a rigid spacer between the N-terminal putative lipid-binding domain and TrioN, whereas the C-terminal half of Trio seems globular. We found that regions C-terminal to TrioN enhance its Rac1 GEF activity and thus could play a regulatory role. We went on to characterize a minimal, well-behaved Trio fragment with enhanced activity, Trio1284–1959, in complex with Rac1 using cryo-electron microscopy and hydrogen-deuterium exchange mass spectrometry and found that the region conferring enhanced activity is disordered. Deletion of two different strongly conserved motifs in this region eliminated this enhancement, suggesting that they form transient intramolecular interactions that promote GEF activity. Because Dbl family RhoGEF modules have been challenging to directly target with small molecules, characterization of accessory Trio domains such as these may provide alternate routes for the development of therapeutics that inhibit Trio activity in human cancer.
登录
查看更多内容
影响因子：
5.2
作者：
Mallone F;Sacchetti M;Lambiase A;Moramarco A
通讯作者：
Moramarco A
DOI：
10.1107/s0907444904019158
发表时间：
2004-12-01
影响因子：
2.2
作者：
Emsley, P;Cowtan, K
通讯作者：
Cowtan, K
影响因子：
9.2
作者：
Estrach, S;Schmidt, S;Debant, A
通讯作者：
Debant, A
影响因子：
56.9
作者：
Lutz, Susanne;Shankaranarayanan, Aruna;Tesmer, John J. G.
通讯作者：
Tesmer, John J. G.
影响因子：
16.8
作者：
Mitin, Natalia;Betts, Laurie;Rossman, Kent L.
通讯作者：
Rossman, Kent L.