Molecular mechanisms of TGF-beta signaling pathway

TGF-β信号通路的分子机制

• 批准号: 7053953
• 负责人: YING E Zhang
• 金额: --
• 依托单位: DIVISION OF BASIC SCIENCES - NCI
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7053953
• 关键词: RNA interference; apoptosis; biological signal transduction; bone morphogenetic proteins; cell differentiation; cell proliferation; enzyme activity; extracellular matrix; gene expression; genetic transcription; genetically modified animals; growth factor receptors; laboratory mouse; mitogen activated protein kinase; neoplastic growth; proteasome; protein degradation; protein kinase; protein protein interaction; protein structure function; transforming growth factors; ubiquitin

Members of the transforming growth factor-beta (TGF-beta) family of peptide growth factors, which include TGF-beta, bone morphogenetic proteins (BMPs) and activins, regulate a broad range of cellular processes from cell growth and differentiation to apoptosis. The signaling responses to TGF-beta and other family members are mediated by a heteromeric complex of two types of transmembrane serine/threonine kinase receptors at the cell surface, and their intracellular substrates, the Smad proteins. Proper TGF-beta superfamily signaling requires precise control of Smad functions. One of the important mechanisms that control Smad activity is ubiquitin-proteasome-mediated degradation. Previously, we and others identified Smurf1 and Smurf2, two new members of the HECT family of E3 ubiquitin ligases, as interacting partners for Smads, but the in vivo function of Smurfs and the selectivity of Smurfs towards Smad in specific signaling pathways remain to be determined. We continued our research interest in this area and began to address these important biological issues. We examined the role of Smurf1 in myogenic and osteogenic differentiation by taking advantage of the in vitro differentiation process of mouse C2C12 myoblast cells, which is subject to control by both TGF-beta and bone morphogenetic protein (BMP). We found that increased expression of Smurf1 promotes myogenic differentiation of C2C12 cells and blocks the BMP-induced osteogenic conversion but has no effect on the TGF-beta-induced differentiation arrest. Consistent with an inhibitory role in the BMP signaling pathway, the elevated Smurf1 markedly reduces the level of endogenous Smad5 while it leaves unaltered the levels of Smad2, Smad3 and Smad7, which are components of the TGF-beta pathway. Adding back Smad5 from a different source to the Smurf1-overexpressing cells restores the BMP-mediated osteoblast conversion. Finally, by depletion of endogenous Smurf1 through small interfering RNA-mediated RNA interference, we demonstrated that Smurf1 is required for the myogenic differentiation of C2C12 cells and plays an important regulatory role in the BMP-2-mediated osteoblast conversion. Although Smads are involved in most actions of the TGF-beta superfamily, many reports have suggested that TGF-beta may signal through alternative pathways. In order to characterize the mechanism of TGF-beta signaling through Smad-independent pathways and to understand the function of Smad-independent TGF-beta receptor signaling, we have generated a mutant TGF-beta type I receptor that is unable to activate Smads but retains kinase activity. We found that this mutant TGF-beta type I receptor is able to activate p38 kinase, and the p38 activation is required for TGF-beta induced apoptosis and epithelial to mesenchymal transition. These results indicate that the TGF-beta receptor exerts its signals through multiple intracellular pathways and provide first hand biochemical evidence to support the existence of Smad-independent TGF-beta receptor signaling. Currently we are working to identify downstream mediators that are responsible for Smad-independent TGF-beta receptor signaling. These studies could uncover novel molecular mechanisms that account for a number of Smad-independent TGF-beta signaling responses. In addition, we are also interested in how TGF-beta signaling converges with other pathways in response to growth factors and consequent activation of mitogen-activated protein kinase(MAPK) pathways. We would like to understand the role of this cross-talk in controlling TGF-beta-regulated gene transcription, cell proliferation, extracellular matrix production,apoptosis and tumor progression. In the meantime, a long term research program using mouse genetics to address the physiological and pathological roles of TGF-beta/Smad signaling in tumorigenesis has also been initiated.

肽生长因子的转化生长因子-β（TGF-β）家族的成员，包括TGF-β、骨形态发生蛋白（BMP）和激活素，调节从细胞生长和分化到凋亡的广泛的细胞过程。对TGF-β和其他家族成员的信号传导应答由细胞表面的两种类型的跨膜丝氨酸/苏氨酸激酶受体及其细胞内底物Smad蛋白的异聚复合物介导。 适当的TGF-β超家族信号传导需要精确控制Smad功能。泛素-蛋白酶体介导的降解是控制Smad活性的重要机制之一。以前，我们和其他人确定了Smurf 1和Smurf 2，E3泛素连接酶的HECT家族的两个新成员，作为Smads的相互作用伴侣，但Smurfs的体内功能和Smurfs在特定信号通路中对Smad的选择性仍有待确定。我们继续对这一领域的研究兴趣，并开始解决这些重要的生物学问题。我们利用小鼠C2 C12成肌细胞的体外分化过程研究了Smurf 1在成肌和成骨分化中的作用，该分化过程受TGF-β和骨形态发生蛋白（BMP）的控制。我们发现Smurf 1的表达增加促进C2 C12细胞的成肌分化，并阻断BMP诱导的成骨转化，但对TGF-β诱导的分化停滞没有影响。与BMP信号通路中的抑制作用一致，升高的Smurf 1显著降低内源性Smad 5的水平，而其保持TGF-β通路的组分Smad 2、Smad 3和Smad 7的水平不变。将来自不同来源的Smad 5重新添加到Smurf 1过表达细胞中可以恢复BMP介导的成骨细胞转化。最后，通过小干扰RNA介导的RNA干扰消除内源性Smurf 1，我们证明了Smurf 1是C2 C12细胞肌源性分化所需的，并在BMP-2介导的成骨细胞转化中起着重要的调节作用。 虽然Smads参与了TGF-β超家族的大多数活动，但许多报告表明TGF-β可能通过替代途径发出信号。为了表征TGF-β信号通过Smad非依赖性途径的机制，并了解Smad非依赖性TGF-β受体信号的功能，我们已经产生了一种突变的TGF-β I型受体，其不能激活Smads，但保留激酶活性。我们发现这种突变的TGF-β I型受体能够激活p38激酶，而p38激活是TGF-β诱导的细胞凋亡和上皮向间充质转化所需的。这些结果表明，TGF-β受体通过多种细胞内途径发挥其信号，并提供第一手的生化证据，支持存在Smad非依赖性TGF-β受体信号转导。目前，我们正在努力确定负责Smad非依赖性TGF-β受体信号传导的下游介质。这些研究可能揭示新的分子机制，解释了许多Smad非依赖性TGF-β信号转导反应。 此外，我们还对TGF-β信号传导如何与其他途径融合以响应生长因子以及随后的促分裂原活化蛋白激酶（MAPK）途径的激活感兴趣。我们希望了解这种串扰在控制TGF-β调节的基因转录、细胞增殖、细胞外基质产生、细胞凋亡和肿瘤进展中的作用。与此同时，一个长期的研究计划，使用小鼠遗传学来解决TGF-β/Smad信号在肿瘤发生中的生理和病理作用也已启动。