Wells R G, Gilboa L, Sun Y, Liu X, Henis Y I, Lodish H F
Department of Medicine, Brigham and Women's Hospital and Harvard Medical School, Boston, Massachusetts 02115, USA.
J Biol Chem. 1999 Feb 26;274(9):5716-22. doi: 10.1074/jbc.274.9.5716.
Transforming growth factor-beta (TGF-beta) binds to and signals via two serine-threonine kinase receptors, the type I (TbetaRI) and type II (TbetaRII) receptors. We have used different and complementary techniques to study the physical nature and ligand dependence of the complex formed by TbetaRI and TbetaRII. Velocity centrifugation of endogenous receptors suggests that ligand-bound TbetaRI and TbetaRII form a heteromeric complex that is most likely a heterotetramer. Antibody-mediated immunofluorescence co-patching of epitope-tagged receptors provides the first evidence in live cells that TbetaRI. TbetaRII complex formation occurs at a low but measurable degree in the absence of ligand, increasing significantly after TGF-beta binding. In addition, we demonstrate that pretreatment of cells with dithiothreitol, which inhibits the binding of TGF-beta to TbetaRI, does not prevent formation of the TbetaRI.TbetaRII complex, but increases its sensitivity to detergent and prevents TGF-beta-activated TbetaRI from phosphorylating Smad3 in vitro. This indicates that either a specific conformation of the TbetaRI. TbetaRII complex, disrupted by dithiothreitol, or direct binding of TGF-beta to TbetaRI is required for signaling.
转化生长因子-β（TGF-β）通过两种丝氨酸-苏氨酸激酶受体，即I型（TβRI）和II型（TβRII）受体结合并发出信号。我们使用了不同且互补的技术来研究由TβRI和TβRII形成的复合物的物理性质和配体依赖性。内源性受体的速度离心表明，与配体结合的TβRI和TβRII形成了一个异源复合物，很可能是一个异源四聚体。抗体介导的表位标记受体的免疫荧光共定位提供了活细胞中的首个证据，表明在没有配体的情况下，TβRI和TβRII复合物的形成程度较低但可测量，在TGF-β结合后显著增加。此外，我们证明用二硫苏糖醇预处理细胞，该物质可抑制TGF-β与TβRI的结合，并不阻止TβRI-TβRII复合物的形成，但会增加其对去污剂的敏感性，并阻止TGF-β激活的TβRI在体外磷酸化Smad3。这表明信号传导需要TβRI-TβRII复合物的特定构象（被二硫苏糖醇破坏）或TGF-β与TβRI的直接结合。
