Repression by C SKI and Melanoma Progression

C SKI 抑制和黑色素瘤进展

基本信息

批准号：
7289699
负责人：
ESTELA E MEDRANO
金额：
$ 26.46万
依托单位：
BAYLOR COLLEGE OF MEDICINE
依托单位国家：
美国
项目类别：
财政年份：
2000
资助国家：
美国
起止时间：
2000-04-01 至 2011-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7289699
关键词：
A Mouse Abbreviations Agar Animals Antisense vector Apoptotic Biological Assay Birds Carcinoma Cell Cycle Cell Nucleus Cell Survival Cells Chickens Chromatin Co-Immunoprecipitations Complex Cyclin-Dependent Kinase Inhibitor Cytoplasm Data Deposition Down-Regulation Electrophoretic Mobility Shift Assay Ensure Environment Event Extracellular Matrix FHL2 gene Fibroblasts Fractionation Genes Genetic Transcription Growth High Pressure Liquid Chromatography Homologous Gene Human In Vitro Inhibition of Apoptosis Invasive Ligand Binding Localized Mass Spectrum Analysis Mediating Melanoma Cell Molecular Mus Mutation Neoplasm Metastasis Nuclear Nuclear Translocation Oncogenes Oncogenic Phenotype Physiological Post-Translational Protein Processing Production Protein Overexpression Proteins RNA Interference Ras Signaling Pathway Reporter Repression Research Personnel Resistance SKI gene Signal Pathway Signal Transduction Skiing Sloan Kettering Virus Smad Proteins Smad protein Small Interfering RNA Stromal Cells Techniques Technology Testing Transforming Growth Factor beta Transforming Growth Factor beta Receptors Translating Tumor Promoters Tumor Suppressor Proteins Tumorigenicity Viral Virus Xenograft Model Xenograft procedure angiogenesis base cell growth cell motility cell stroma chromatin immunoprecipitation human NRCAM protein in vivo knock-down melanocyte melanoma migration mouse model prevent programs promoter protein function receptor response sensor size tumor tumor progression

项目摘要

DESCRIPTION (provided by applicant): Transforming growth factor-beta (TGF-beta) has dual and paradoxical functions as a tumor suppressor and promoter of tumor progression and metastasis. In spite of absence of mutations in TGF-beta receptors, melanoma cells are resistant to its growth inhibitory activity. Furthermore, melanoma cells increase production and secretion of TGF-beta, which upon activation induces matrix deposition, angiogenesis, survival and transition to more aggressive phenotypes. We have demonstrated that SKI functions as a molecular switch, converting the TGF-beta co-regulator proteins Smad2/3 from an activating to a repressing entity on chromatin. SKI is also a potent activator of the Wnt signaling pathway, targeting MITF and NrCAM, two genes involved in survival, migration and invasion. Based on these findings and new preliminary data we hypothesize that SKI functions as a sensor and modifier of environmental signals, protecting melanoma cells from growth inhibition and apoptosis and stimulating proliferation and invasion. To test this hypothesis, we will: Aim 1: Determine the molecular mechanism(s) by which SKI senses and modifies growth inhibitory signals. We will use endogenous and exogenous SKI to determine how SKI complexes respond to inhibitory and/or apoptotic signals mediated by TGF-beta. Analysis and characterization of SKI protein complexes will be performed using new HPLC fractionation/co-immunoprecipitations techniques and Mass Spectrometry. Functional relevance of SKI complexes will be validated by promoter-reporter assays, sequential chromatin Immunoprecipitations, and gel-shift assays. Aim 2: Determine whether knocking down SKI prevents melanoma growth and metastasis in a mouse xenograft model. We will use proven SKI RNAi technology to test melanoma tumor formation using xenografts containing human stroma-derived fibroblasts. Aim 3: Test whether SKI cooperates with the RAS signaling pathway to induce melanoma progression in vivo. We will analyze in detail molecular requirements needed for switching SKI from a tumor suppressor to a tumor promoter using a syngeneic mouse model. Together, the three aims will likely identify and characterize a cascade of molecular and cellular events regulated by SKI that can trigger cell cycle aberrations, increased tumorigenicity and metastatic potential of melanocytes.

描述（由申请人提供）：转化生长因子-β（TGF-β）作为肿瘤抑制因子和肿瘤进展和转移的促进剂具有双重且矛盾的功能。尽管 TGF-β 受体不存在突变，但黑色素瘤细胞对其生长抑制活性具有抵抗力。此外，黑色素瘤细胞增加 TGF-β 的产生和分泌，TGF-β 激活后可诱导基质沉积、血管生成、存活并转变为更具攻击性的表型。我们已经证明，SKI 作为分子开关发挥作用，将染色质上的 TGF-β 辅助调节蛋白 Smad2/3 从激活实体转变为抑制实体。 SKI 也是 Wnt 信号通路的有效激活剂，靶向 MITF 和 NrCAM，这两个基因涉及生存、迁移和入侵。基于这些发现和新的初步数据，我们假设 SKI 充当环境信号的传感器和调节器，保护黑色素瘤细胞免受生长抑制和凋亡并刺激增殖和侵袭。为了检验这一假设，我们将：目标 1：确定 SKI 感知和修改生长抑制信号的分子机制。我们将使用内源性和外源性 SKI 来确定 SKI 复合物如何响应 TGF-β 介导的抑制和/或凋亡信号。 SKI 蛋白复合物的分析和表征将使用新的 HPLC 分级分离/免疫共沉淀技术和质谱法进行。 SKI 复合物的功能相关性将通过启动子-报告基因测定、连续染色质免疫沉淀和凝胶迁移测定进行验证。目标 2：确定在小鼠异种移植模型中敲低 SKI 是否可以阻止黑色素瘤的生长和转移。我们将使用经过验证的 SKI RNAi 技术，使用含有人基质来源的成纤维细胞的异种移植物来测试黑色素瘤的形成。目标 3：测试 SKI 是否与 RAS 信号通路协同诱导体内黑色素瘤进展。我们将使用同基因小鼠模型详细分析将 SKI 从肿瘤抑制因子转变为肿瘤促进因子所需的分子要求。这三个目标共同可能会识别和表征 SKI 调节的一系列分子和细胞事件，这些事件可能引发细胞周期畸变、增加黑素细胞的致瘤性和转移潜力。