KRAS-regulated Ago2-mediated sorting of extracellular RNAs into exosomes

KRAS 调节的 Ago2 介导的细胞外 RNA 分选至外泌体

• 批准号: 9327270
• 负责人: Lizandra Jimenez
• 金额: $ 5.67万
• 依托单位: VANDERBILT UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-08-01 至 2020-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9327270
• 关键词: Address; Affect; Area; Binding; Binding Sites; Cancer Cell Growth; Cancer Control; Cell Communication; Cell physiology; Cells; Coculture Techniques; Colon Carcinoma; Colonic Neoplasms; Colorectal Cancer; Colorectal Neoplasms; Complex; Data Set; Endosomes; Extracellular Protein; GTP Binding; Gene Expression; High-Throughput Nucleotide Sequencing; Immunoprecipitation; In Vitro; KRAS2 gene; Large Intestine Carcinoma; Lipids; Location; MEKs; Malignant Neoplasms; Malignant neoplasm of lung; Malignant neoplasm of pancreas; Mediating; Mediator of activation protein; MicroRNAs; Missense Mutation; Modification; Molecular; Mus; Mutation; Neoplasm Metastasis; Oncogenic; Phenotype; Phosphorylation; Play; Proteins; RNA; RNA Binding; RNA Transport; RNA-Induced Silencing Complex; Reporting; Research; Role; Serine; Signal Transduction; Sorting - Cell Movement; System; Therapeutic; Tumor Promotion; Untranslated RNA; angiogenesis; cancer biomarkers; cancer cell; cell type; conditioning; crosslink; exosome; experimental study; extracellular; extracellular vesicles; in vivo; knock-down; late endosome; mRNA Transcript Degradation; malignant breast neoplasm; mutant; novel; prevent; trafficking; transcriptome sequencing; tumor; tumor growth; tumor microenvironment; vesicular release

Project Summary/Abstract Approximately 30-50% of colorectal tumors carry activating missense mutations in KRAS. Late endosome- derived exosomes are often secreted from cancer cells and serve as mediators of cell-cell communication. Exosomes have been reported to be associated with tumor aggressiveness and metastasis. We have previously reported that the mutant KRAS status of colorectal cancer cells greatly affects the extracellular protein and RNA composition of exosomes. We also recently reported that sorting of Argonaute 2 (Ago2) and several Ago2-associated miRNAs to exosomes is controlled by KRAS-MEK-Erk signaling in colon cancer cells. Specifically, phosphorylation of Ago2 on serine 387 prevents Ago2 association with multivesicular endosomes (MVE) and sorting into exosomes. This finding identifies a molecular mechanism that has the potential to control the secretion of multiple miRNAs, miRNA-associated proteins, and potentially additional types of RNAs into exosomes. In this proposed project, I will identify the exact role of Argonaute 2 in sorting RNAs to exosomes, including determining what proportion of miRNAs are secreted in exosomes using this Ago2- controlled mechanism as well as whether there is selectivity in miRNA sorting. I will also determine whether other RNAs rely on Ago2 for their sorting into exosomes. Finally, I will investigate the functional consequence of Ago2-mediated RNA secretion from KRAS-dependent colon cancer cells on recipient cell gene expression and phenotypes both in vitro and in vivo. !

项目概要/摘要 大约 30-50% 的结直肠肿瘤携带 KRAS 激活错义突变。晚期内体- 衍生的外泌体通常由癌细胞分泌，并作为细胞间通讯的介质。 据报道，外泌体与肿瘤侵袭性和转移有关。我们有 此前报道结直肠癌细胞KRAS突变状态极大影响细胞外 外泌体的蛋白质和RNA组成。我们最近还报道了 Argonaute 2 (Ago2) 和 结肠癌细胞中，一些与 Ago2 相关的 miRNA 与外泌体受 KRAS-MEK-Erk 信号控制。 具体而言，Ago2 在丝氨酸 387 上的磷酸化可防止 Ago2 与多囊泡内体结合 (MVE) 并分选为外泌体。这一发现确定了一种有潜力的分子机制 控制多种 miRNA、miRNA 相关蛋白和潜在其他类型 RNA 的分泌 进入外泌体。在这个拟议的项目中，我将确定 Argonaute 2 在分选 RNA 中的确切作用 外泌体，包括使用 Ago2- 确定外泌体中分泌的 miRNA 的比例 控制机制以及 miRNA 分选是否具有选择性。我也会确定是否 其他 RNA 依赖 Ago2 来分选外泌体。最后，我将研究功能后果 KRAS依赖性结肠癌细胞Ago2介导的RNA分泌对受体细胞基因表达的影响 以及体外和体内的表型。 ！