Nyati Shyam, Schinske-Sebolt Katrina, Pitchiaya Sethuramasundaram, Chekhovskiy Katerina, Chator Areeb, Chaudhry Nauman, Dosch Joseph, Van Dort Marcian E, Varambally Sooryanarayana, Kumar-Sinha Chandan, Nyati Mukesh Kumar, Ray Dipankar, Walter Nils G, Yu Hongtao, Ross Brian Dale, Rehemtulla Alnawaz
Center for Molecular Imaging, University of Michigan, Ann Arbor, MI 48109, USA. Department of Radiation Oncology, University of Michigan, Ann Arbor, MI 48109, USA.
Department of Radiation Oncology, University of Michigan, Ann Arbor, MI 48109, USA.
Sci Signal. 2015 Jan 6;8(358):ra1. doi: 10.1126/scisignal.2005379.
Transforming growth factor-β (TGF-β) signaling regulates cell proliferation and differentiation, which contributes to development and disease. Upon binding TGF-β, the type I receptor (TGFBRI) binds TGFBRII, leading to the activation of the transcription factors SMAD2 and SMAD3. Using an RNA interference screen of the human kinome and a live-cell reporter for TGFBR activity, we identified the kinase BUB1 (budding uninhibited by benzimidazoles-1) as a key mediator of TGF-β signaling. BUB1 interacted with TGFBRI in the presence of TGF-β and promoted the heterodimerization of TGFBRI and TGFBRII. Additionally, BUB1 interacted with TGFBRII, suggesting the formation of a ternary complex. Knocking down BUB1 prevented the recruitment of SMAD3 to the receptor complex, the phosphorylation of SMAD2 and SMAD3 and their interaction with SMAD4, SMAD-dependent transcription, and TGF-β-mediated changes in cellular phenotype including epithelial-mesenchymal transition (EMT), migration, and invasion. Knockdown of BUB1 also impaired noncanonical TGF-β signaling mediated by the kinases AKT and p38 MAPK (mitogen-activated protein kinase). The ability of BUB1 to promote TGF-β signaling depended on the kinase activity of BUB1. A small-molecule inhibitor of the kinase activity of BUB1 (2OH-BNPP1) and a kinase-deficient mutant of BUB1 suppressed TGF-β signaling and formation of the ternary complex in various normal and cancer cell lines. 2OH-BNPP1 administration to mice bearing lung carcinoma xenografts reduced the amount of phosphorylated SMAD2 in tumor tissue. These findings indicated that BUB1 functions as a kinase in the TGF-β pathway in a role beyond its established function in cell cycle regulation and chromosome cohesion.
转化生长因子-β（TGF-β）信号传导调节细胞增殖和分化，这对发育和疾病都有影响。TGF-β结合后，I型受体（TGFBRI）与TGFBRII结合，导致转录因子SMAD2和SMAD3激活。通过对人类激酶组进行RNA干扰筛选以及使用TGFBR活性的活细胞报告基因，我们确定激酶BUB1（苯并咪唑不抑制的芽殖-1）是TGF-β信号传导的关键介质。在TGF-β存在的情况下，BUB1与TGFBRI相互作用并促进TGFBRI和TGFBRII的异二聚化。此外，BUB1与TGFBRII相互作用，提示形成三元复合物。敲低BUB1可阻止SMAD3募集到受体复合物、SMAD2和SMAD3的磷酸化及其与SMAD4的相互作用、SMAD依赖的转录以及TGF-β介导的细胞表型变化，包括上皮-间质转化（EMT）、迁移和侵袭。敲低BUB1也损害了由激酶AKT和p38丝裂原活化蛋白激酶（MAPK）介导的非经典TGF-β信号传导。BUB1促进TGF-β信号传导的能力取决于其激酶活性。BUB1激酶活性的小分子抑制剂（2OH-BNPP1）和BUB1的激酶缺陷突变体在各种正常和癌细胞系中均抑制TGF-β信号传导和三元复合物的形成。给携带肺癌异种移植瘤的小鼠施用2OH-BNPP1可减少肿瘤组织中磷酸化SMAD2的量。这些发现表明，BUB1在TGF-β途径中作为一种激酶发挥作用，其作用超出了其在细胞周期调控和染色体黏附中已确定的功能。
