Role of the Sec61 Translocon in EGF Receptor Trafficking and Signaling

Sec61 易位子在 EGF 受体运输和信号转导中的作用

• 批准号: 7847939
• 负责人: GRAHAM F CARPENTER
• 金额: $ 1.97万
• 依托单位: VANDERBILT UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-04-01 至 2009-10-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7847939
• 关键词: Address; Antibodies; Binding; Biochemistry; Biological; Biology; Cell Cycle Progression; Cell Nucleus; Cells; Cellular biology; Clinical; Cyclin D1; Cytoplasm; Cytosol; Data; Dependence; EGF gene; Endoplasmic Reticulum; Endosomes; Epidermal Growth Factor Receptor; Erbitux; Golgi Apparatus; Grant; Growth; Growth Factor; Heat-Shock Proteins 70; Hormones; Investigation; Ligands; Malignant Neoplasms; Membrane; Monoclonal Antibody C225; Nuclear; Nuclear Localization Signal; Nuclear Receptors; Nuclear Translocation; Pathway interactions; Pharmaceutical Preparations; Proliferating; Proteins; Reagent; Receptor Activation; Role; Route; Signal Transduction; Sorting - Cell Movement; Testing; Transmembrane Domain; Tyrosine; cancer therapy; clinically relevant; factor C; mutant; p97 ATPase; prevent; protein misfolding; receptor; research study; response; trafficking

This application seeks to explore a new route of intracellular trafficking of growth factor- activated EGF receptor to the nucleus. Also, the capacity of a clinically relevant EGF receptor antibody to promote nuclear localization of the EGF receptor will be investigated. In brief, preliminary data show that following the addition of EGF, EGF receptors are slowly trafficked to the endoplasmic reticulum (ER). In the ER these receptors interact with the Sec61 translocon and are retrotranslocated to the cytoplasm as non-membrane bound molecules. HSP70 is required for retrotranslocation to the cytosol and likely functions by associating with transmembrane domains of the soluble EGF receptor to prevent aggregation. Knockdown of Sec61? prevents the EGF-dependent translocation of its receptor to the nucleus and the expression of cyclin D1, indicating that ER translocation is a precursor for nuclear localization and signaling function of the EGF receptor. In the first two aims, mechanistic questions are addressed regarding the cell biology and biochemistry of this pathway. In the last two aims, the pathway is explored in terms of growth control and a clinically used antibody to the EGF receptor. The first aim proposes to investigate the intracellular trafficking route by which the activated EGFR is trafficked to the ER. Experiments are proposed to determine how different cell internalization mechanisms, the Golgi, and endosomes may participate in this pathway The second aim focuses on the mechanism by which the receptor is recognized by the Sec61 translocon in the ER. This will test the possibility that the activated receptor is recognized by the known mechanism in the ER that sorts misfolded proteins for Sec-61-dependent retrotranslocation and cytosolic degradation. Also, this aim will include experiments to identify cytoplasmic and ER factors that participate in retrotranslocation, the fate(s) of translocated receptor, and the identity of EGF receptor fragments found in the ER and nucleus. This includes HSP70 and p97, an ATPase that is tyrosine phosphorylated following the addition of EGF. The third aim will determine the possible role of Sec61 in the EGF-stimulated G1?S transition. The final aim will explore the mechanism by which the clinically employed EGF receptor antibody C225 (Erbitux) is able to induce receptor trafficking to the ER and nucleus. The role of Sec61 in cell responses to C225 will be investigated also. The focus of this grant is the EGF receptor, a protein that is a rational target for several chemotherapeutic drugs that are approved for clinical use for different cancers. The application proposes investigation of this receptor in response to its normal activator, a hormone-like growth factor that stimulates cells to proliferate, as well as an antibody to the receptor that is one of the clinically approved reagents for cancer treatment.

本申请旨在探索生长因子细胞内运输的新途径- 激活细胞核的 EGF 受体。此外，临床相关 EGF 受体的能力 将研究促进EGF受体核定位的抗体。简而言之， 初步数据表明，添加 EGF 后，EGF 受体会缓慢转运至 内质网（ER）。在 ER 中，这些受体与 Sec61 易位子相互作用 并作为非膜结合分子逆向转运至细胞质。 HSP70 是 逆转录至胞质溶胶所需的，并且可能通过与 可溶性 EGF 受体的跨膜结构域以防止聚集。击倒 第 61 节？防止其受体依赖 EGF 易位至细胞核，并且 cyclin D1 的表达，表明 ER 易位是核定位的前体 EGF受体的信号传导功能。在前两个目标中，机械问题是 讨论了该途径的细胞生物学和生物化学。在最后两个目标中， 在生长控制和临床使用的 EGF 抗体方面探索了途径 受体。 第一个目标是研究细胞内运输途径 激活的 EGFR 被转运至 ER。建议进行实验以确定如何不同 细胞内化机制，高尔基体和内体可能参与该途径 第二个目标关注 Sec61 识别受体的机制 急诊室中的易位子。这将测试激活的受体被识别的可能性 ER 中已知的机制，可根据 Sec-61 依赖性对错误折叠的蛋白质进行排序 逆转录易位和胞质降解。此外，这个目标将包括实验来确定 参与逆向易位的细胞质和内质网因子，易位的命运 受体，以及在内质网和细胞核中发现的 EGF 受体片段的身份。这 包括 HSP70 和 p97，一种 ATP 酶，添加后会被酪氨酸磷酸化 表皮生长因子。 第三个目标将确定 Sec61 在 EGF 刺激的 G1?S 中的可能作用 过渡。最终目的是探讨临床上使用的EGF的作用机制。 受体抗体 C225 (Erbitux) 能够诱导受体转运至内质网和细胞核。 Sec61 在细胞对 C225 反应中的作用也将被研究。这项资助的重点是 EGF 受体，这种蛋白质是多种药物的合理靶点。 批准用于临床治疗不同癌症的化疗药物。应用 建议研究这种受体对其正常激活剂（一种激素样生长）的反应 刺激细胞增殖的因子，以及受体之一的抗体 临床批准的用于癌症治疗的试剂。