CNRS, UMR 5203, Institut de Génomique Fonctionnelle, Département de Pharmacologie Moléculaire, Montpellier, France.
EMBO J. 2011 Jan 5;30(1):32-42. doi: 10.1038/emboj.2010.270. Epub 2010 Nov 9.
Seven-transmembrane domain (7TM) receptors have important functions in cell-cell communication and can assemble into dimers or oligomers. Such complexes may allow specific functional cross-talk through trans-activation of interacting 7TMs, but this hypothesis requires further validation. Herein, we used the GABAB receptor, which is composed of two distinct subunits, GABAB1, which binds the agonist, and GABAB2, which activates G proteins, as a model system. By using a novel orthogonal-labelling approach compatible with time-resolved FRET and based on ACP- and SNAP-tag technologies to verify the heterodimerization of wild-type and mutated GABAB subunits, we demonstrate the existence of a direct allosteric coupling between the 7TMs of GABAB heterodimers. Indeed, a GABAB receptor, in which the GABAB2 extracellular domain was deleted, was still capable of activating G proteins. Furthermore, synthetic ligands for the GABAB2 7TM could increase agonist affinity at the GABAB1 subunit in this mutated receptor. In addition to bringing new information on GABAB receptor activation, these data clearly demonstrate the existence of direct trans-activation between the 7TM of two interacting proteins.
七跨膜域（7TM）受体在细胞间通讯中具有重要功能，可形成二聚体或寡聚体。这种复合物可能允许通过相互作用的 7TM 的跨激活来进行特定的功能串扰，但这一假设需要进一步验证。在此，我们使用 GABAB 受体作为模型系统，该受体由两个不同的亚基组成，即结合激动剂的 GABAB1 和激活 G 蛋白的 GABAB2。通过使用一种新颖的正交标记方法，该方法与时间分辨荧光共振能量转移（FRET）兼容，并基于 ACP 和 SNAP 标签技术来验证野生型和突变的 GABAB 亚基的异二聚化，我们证明了 GABAB 异二聚体的 7TM 之间存在直接的变构偶联。事实上，即使删除了 GABAB2 细胞外结构域，GABAB 受体仍能够激活 G 蛋白。此外，GABAB2 7TM 的合成配体可以增加在这种突变受体中 GABAB1 亚基的激动剂亲和力。除了提供有关 GABAB 受体激活的新信息外，这些数据还清楚地证明了两种相互作用的蛋白质的 7TM 之间存在直接的跨激活。
