Probing a novel signaling complex that sustains AKT activation to support stress survival in cancer

项目摘要 代谢异常是癌症的一个标志，它有助于癌细胞生长，经历恶性转化， 并在营养匮乏等压力条件下生存。癌细胞暴露于许多细胞 在肿瘤发生过程中的压力，这必须克服恶性肿瘤的传播。在癌症中， 许多信号网络的异常激活会切断对生长、代谢和代谢的控制， 存活率，并且最近的努力已经寻求治疗靶向癌症代谢。磷脂酰肌醇3- 激酶（PI 3 K）-AKT（蛋白激酶B）信号通路在人类癌症中是最活化的，并且具有广泛的生物学活性。 对细胞代谢的影响。我们最近发现Cdc 42/Rac鸟嘌呤核苷酸交换 胞质分裂因子7（Dock 7）作为支持持续基础AKT的新型信号传导节点 雷帕霉素（mTOR）活性的激活和机制靶点，如由其下游靶点S6所确定 在应激条件下，S6 K蛋白可被激活，以维持细胞存活和转化所需的信号传导活性。 我们发现，Dock 7是多种癌细胞系抵抗失巢凋亡和表现出锚定非依赖性所必需的。 增长虽然我们观察到与生长因子刺激相比AKT磷酸化水平相对较低， Dock 7依赖性信号传导对于营养剥夺期间癌细胞的存活至关重要。我假设 在细胞应激下，Dock 7作为AKT的支架，维持其磷酸化和组织化， 应激生存所需的mTOR信号传导伙伴。本项目将研究这一作用 在细胞应激下为癌细胞提供存活益处的新型Dock 7/AKT/mTOR信号传导活性。我 建议研究Dock 7依赖性信号传导活性对AKT/mTOR信号传导的影响， 压力，以及恶性进展和攻击的关键特征。在目标1中，我将研究 Dock 7的Dock同源区2（DHR 2）结构域的功能活性，其负责GEF 活性，在基础AKT磷酸化癌细胞应激生存和恶性转化。在目标2中，我 接下来将确定DHR 1结构域在Dock 7依赖性AKT磷酸化、癌细胞应激和细胞凋亡中的新作用。 生存和恶性转化。然后，在目标3中，我将确定这个Dock 7的亚细胞位置 在应激条件下的信号复合物，并确定DHR 1，DHR 2和激活的个体作用， Dock 7中的Cdc 42本地化。本提案中的工作将提供Dock 7的生物化学表征 信号传导活性，这将导致对Dock 7依赖性AKT/mTOR激活的机制的理解， 癌细胞应激存活。这些发现不仅有助于理解癌症的侵略性， 还可以鉴定用于癌症治疗的新的治疗靶点。

PROJECT SUMMARY Abnormal metabolism is a hallmark of cancer that helps cancer cells to grow, undergo malignant transformation, and survive under stressful conditions such as nutrient deprivation. Cancer cells are exposed to many cellular stresses during tumorigenesis, which must be overcome for the propagation of malignancy. In cancer, the abnormal activation of many signaling networks serves to disconnect the control of growth, metabolism, and survival, and recent efforts have sought to therapeutically target cancer metabolism. The phosphatidylinositol 3- kinase (PI3K)-AKT (protein kinase B) signaling pathway is the most activated in human cancer and has a wide range of effects on cellular metabolism. We have recently identified the Cdc42/Rac guanine nucleotide exchange factor (GEF) dedicator of cytokinesis 7 (Dock7) as a novel signaling node that supports sustained basal AKT activation and mechanistic target of rapamycin (mTOR) activity as determined by its downstream target S6 kinase (S6K) during stressful conditions to maintain signaling activity required for cell survival and transformation. We find that Dock7 is required for multiple cancer cell lines to resist anoikis and exhibit anchorage-independent growth. While we observe relatively low levels of AKT phosphorylation compared to stimulation by growth factors, Dock7-dependent signaling is critical for the survival of cancer cells during nutrient deprivation. I hypothesize that under cellular stress Dock7 serves as a scaffold for AKT, sustaining its phosphorylation and organizing signaling partners for mTOR signaling required for stress survival. This project will investigate the role of this novel Dock7/AKT/mTOR signaling activity in providing a survival benefit to cancer cells under cellular stress. I propose to study the impact of Dock7-dependent signaling activity on AKT/mTOR signaling, cell survival under stress, and critical characteristics of malignant progression and aggression. In Aim 1, I will investigate the functional activities of the Dock-homology region 2 (DHR2) domain of Dock7, which is responsible for GEF activity, in basal AKT phosphorylation for cancer cell stress survival and malignant transformation. In Aim 2, I will next identify the novel role of the DHR1 domain in Dock7-dependent AKT phosphorylation, cancer cell stress survival, and malignant transformation. Then, in Aim 3, I will identify the subcellular location of this Dock7 signaling complex under stress conditions and determine the individual roles of DHR1, DHR2, and activated Cdc42 in Dock7 localization. The work in this proposal will provide biochemical characterization of Dock7 signaling activity that will lead to a mechanistic understanding of Dock7-dependent AKT/mTOR activation in cancer cell stress survival. These findings will not only contribute to the understanding of cancer aggression and metabolism but may also identify new therapeutic targets for cancer treatment.