Regulation of TGFBeta-induced apoptosis in liver cells

TGFβ诱导的肝细胞凋亡的调节

• 批准号: 7119199
• 负责人: KUNXIN LUO
• 金额: $ 26.73万
• 依托单位: UNIVERSITY OF CALIFORNIA BERKELEY
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-08-21 至 2009-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7119199
• 关键词: apoptosis; biological signal transduction; cell growth regulation; cell line; cell proliferation; genetic regulation; immunoprecipitation; liver cells; phosphorylation; polymerase chain reaction; protein protein interaction; protein sequence; protein structure function; transfection; transforming growth factors

DESCRIPTION (provided by applicant): The tightly regulated homeostatic mechanisms between hepatocyte proliferation and apoptosis that exist in normal liver tissue are disrupted in many liver diseases and during hepatocarcinogenesis. The transforming growth factor beta (TGFbeta) signaling pathway is a central component of the mechanisms by which cell growth and apoptosis are regulated in the liver. The long-term objective of this project is to understand the molecular mechanism by which TGFbetaa regulates hepatocyte proliferation and apoptosis. TGFbeta is a potent inducer of hepatocyte apoptosis both in vivo and in vitro. The apoptotic activity of TGFbeta can be antagonized by the action of insulin and its downstream signaling molecules. This proposal is designed to understand how TGFbeta pathway and insulin pathway cross talk to regulate the sensitivity of hepatocytes to TGFbeta-induced apoptosis. Smad proteins are critical mediators of TGFbeta signaling. In the absence of ligand, Smad2 and Smad3 are located in the cytoplasm. Upon binding of TGFbeta to its receptors and the subsequent activation of the receptor serine/threonine kinases, Smad2 and Smad3 become phosphorylated by the receptor kinases and then translocate to the nucleus, where they can form heteromeric complexes with Smad4 and activate transcription of TGFbeta responsive genes. Among these Smad proteins, Smad3 has been shown to mediate TGFbeta-induced apoptosis. In a search for molecules that interact with Smad3, the P.I. identified Akt, a critical molecule functioning in the cell survival pathway downstream of insulin. Akt can directly interact with Smad3, and this interaction is regulated by TGFbeta and insulin. Because expression of an activated Akt results in protection of cells from TGFbeta-induced apoptosis, we hypothesized that Akt may protect cells from TGFbeta-induced apoptosis by interacting with Smad3 and preventing it from mediating the apoptotic signals of TGFbeta. The following specific aims are designed to further characterize the interaction between Akt and Smad3 and uncover the physiological significance of this interaction. The specific aims are: 1) Mapping the amino acid residues in Akt and Smad3 required for their interaction. 2) Analysis of the role of the Smad3- Akt interaction in protection from TGFbeta-induced apoptosis. 3) Analysis of the molecular mechanism by which Akt inhibits TGFbeta-induced apoptosis. In particular, we will investigate whether Akt inhibits Smad3 through affecting its phosphorylation or through physical sequestration. These studies will allow us to have a better understanding of how TGFa and insulin signaling pathways cross talk to regulate the sensitivity to TGFbeta-induced apoptosis. Since this cross talk plays an important role in the control of liver size and disruption of this highly regulated process can result in liver diseases and hepatocarcinogenesis, these studies may contribute to a better understanding of the maintenance of normal liver homeostasis and the cause of liver diseases.

描述（申请人提供）：正常肝组织中存在的肝细胞增殖和凋亡之间严格调节的稳态机制在许多肝脏疾病和肝癌发生过程中被破坏。转化生长因子β (TGFbeta) 信号通路是肝脏中细胞生长和凋亡调节机制的核心组成部分。该项目的长期目标是了解TGFbetaa调节肝细胞增殖和凋亡的分子机制。 TGFbeta 是体内和体外肝细胞凋亡的有效诱导剂。 TGFbeta的细胞凋亡活性可以被胰岛素及其下游信号分子的作用所拮抗。该提案旨在了解 TGFbeta 通路和胰岛素通路如何相互作用来调节肝细胞对 TGFbeta 诱导的细胞凋亡的敏感性。 Smad 蛋白是 TGFbeta 信号转导的关键介质。在没有配体的情况下，Smad2和Smad3位于细胞质中。当 TGFbeta 与其受体结合并随后激活受体丝氨酸/苏氨酸激酶时​​，Smad2 和 Smad3 被受体激酶磷酸化，然后转位到细胞核，在细胞核中它们可以与 Smad4 形成异聚复合物并激活 TGFbeta 响应基因的转录。在这些 Smad 蛋白中，Smad3 已被证明可以介导 TGFbeta 诱导的细胞凋亡。在寻找与 Smad3 相互作用的分子时，P.I.确定了 Akt，一种在胰岛素下游细胞生存途径中发挥作用的关键分子。 Akt 可以直接与 Smad3 相互作用，并且这种相互作用受到 TGFbeta 和胰岛素的调节。由于激活的 Akt 的表达可保护细胞免受 TGFbeta 诱导的细胞凋亡，因此我们假设 Akt 可能通过与 Smad3 相互作用并阻止其介导 TGFbeta 的细胞凋亡信号来保护细胞免受 TGFbeta 诱导的细胞凋亡。以下具体目标旨在进一步表征 Akt 和 Smad3 之间的相互作用，并揭示这种相互作用的生理意义。具体目标是： 1) 绘制 Akt 和 Smad3 相互作用所需的氨基酸残基。 2)分析Smad3-Akt相互作用在保护TGFbeta诱导的细胞凋亡中的作用。 3)Akt抑制TGFbeta诱导的细胞凋亡的分子机制分析。特别是，我们将研究 Akt 是否通过影响其磷酸化或通过物理隔离来抑制 Smad3。这些研究将使我们更好地了解 TGFα 和胰岛素信号通路如何相互作用来调节对 TGFβ 诱导的细胞凋亡的敏感性。由于这种串扰在控制肝脏大小方面发挥着重要作用，而这种高度调控的过程的破坏可能导致肝脏疾病和肝癌发生，因此这些研究可能有助于更好地了解正常肝脏稳态的维持和肝脏疾病的原因。