Identifying endosomal Phosphatidylinositol 4-phosphate 5-kinase isoforms regulating growth factor stimulated integrin recycling and migration

鉴定调节生长因子刺激的整合素回收和迁移的内体磷脂酰肌醇 4-磷酸 5-激酶亚型

• 批准号: 10217317
• 负责人: Lorraine C Santy
• 金额: $ 16.05万
• 依托单位: PENNSYLVANIA STATE UNIVERSITY, THE
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-05-01 至 2022-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10217317
• 关键词: Actins; Adhesions; Affect; Automobile Driving; Binding; Biosensor; Carcinoma; Cell Adhesion; Cell membrane; Cell surface; Cells; Chemicals; Consumption; Core Protein; Cytoskeleton; Development; Dimerization; Disease; Disease Progression; EGF gene; Endosomes; Enzymes; Epithelial; Epithelial Cells; Extracellular Matrix; Fibrosis; Focal Adhesions; Functional disorder; Generations; Genes; Growth Factor; Homeostasis; Hyperactivity; Individual; Integrin Binding; Integrins; Intervention; Knowledge; Lead; Lipid Binding; Lipids; Location; Malignant Epithelial Cell; Membrane; Membrane Proteins; Metastatic to; Monomeric GTP-Binding Proteins; Mutation; Neoplasm Metastasis; Normal tissue morphology; Oncogenic; Pathway interactions; Phosphatidylinositols; Phosphorylation; Phosphotransferases; Play; Process; Production; Protein Isoforms; Protein Kinase; Proteins; RNA Splicing; Recycling; Role; Serum; Side; Signal Pathway; Signal Transduction; Specificity; Spliced Genes; Stromal Cells; Structure; System; Tail; Testing; Tissues; Traction; Variant; Work; base; cancer cell; cancer invasiveness; cell motility; epithelial repair; extracellular; fighting; genetic regulatory protein; inorganic phosphate; knock-down; migration; migration stimulating factor; phosphatidylinositol 4-phosphate; phosphoinositide-3,4,5-triphosphate; recruit; repaired; response; restoration; therapy development; tissue repair; trafficking; tumor microenvironment; tumor progression; tumorigenesis; wound healing

PROJECT SUMMARY Epithelial cells are normally non-motile but can become migratory in response to growth factor signaling. Growth factor stimulated migration plays important roles in development and is an early step in the repair process after tissue damage. During cancer progression however, growth factor signaling can become aberrantly activated through oncogenic mutation or the actions of stromal cells in the tumor microenvironment. Growth factor stimulated migration of carcinoma cells promotes metastasis and further disease progression. A thorough understanding of the signals that initiate migration in response to growth factor signaling is essential for developing interventions to either enhance migration during wound healing or inhibit adhesion and migration of cancer cells. Focal adhesions attach cells to the extracellular matrix and allow the generation of traction forces that cells use to migrate. By necessity, growth factor stimulated migration requires increased assembly of new focal adhesions, which are organized by integrins. Growth factor stimulation leads to the rapid recycling of previously internalized integrins to the plasma membrane. Previous work in the Santy lab demonstrated that cytohesin activity is required for stimulated integrin recycling and that only those cytohesin splice variants that bind phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2) can initiate this trafficking process. PI(4,5)P2 is produced locally where needed from PI4P by phosphatidylinositol 4-phosphate 5-kinases (PIP5Ks), implicating PIP5Ks in the stimulated recycling of integrins. Previous work on these understudied kinases has predominately focused on their actions at the plasma membrane during signaling and internalization of membrane proteins. The proposed studies will determine which PIP5K isoforms are required for stimulated integrin recycling and determine where within the endosomal system they produce PI(4,5)P2 to initiate stimulated integrin recycling. A knockdown and rescue strategy will be used to test the impact of PIP5K splice isoforms on integrin recycling. The location where these isoforms act will be determined by localization of both the isoforms and their product, PI(4,5)P2. In an orthogonal approach, chemically induced dimerization will be used to recruit PI(4,5)P2 producing and consuming enzymes to individual endosomal compartments to determine where local production of this lipid is required to stimulate the return of integrins to the cell surface. Understanding the steps that lead to growth factor stimulated integrin recycling will allow development of interventions to modulate this process without affecting growth factor signaling as a whole.

项目概要 上皮细胞通常是不活动的，但可以响应生长因子信号而迁移。 生长因子刺激的迁移在发育中发挥重要作用，是修复的早期步骤 组织损伤后的过程。然而，在癌症进展过程中，生长因子信号传导可能会变得 通过致癌突变或肿瘤微环境中基质细胞的作用异常激活。 生长因子刺激癌细胞的迁移促进转移和进一步的疾病进展。一个 彻底了解响应生长因子信号传导启动迁移的信号至关重要 开发干预措施以增强伤口愈合过程中的迁移或抑制粘附和 癌细胞的迁移。粘着斑将细胞附着在细胞外基质上，并允许产生 细胞用于迁移的牵引力。必然地，生长因子刺激的迁移需要增加 由整合素组织的新粘着斑的组装。生长因子刺激导致 先前内化的整合素快速回收至质膜。之前在 Santy 实验室的工作 证明细胞粘附素活性是刺激整合素再循环所必需的，并且只有那些细胞粘附素 结合磷脂酰肌醇 4,5-二磷酸 (PI(4,5)P2) 的剪接变体可以启动此运输过程。 PI(4,5)P2 在需要时通过磷脂酰肌醇 4-磷酸 5-激酶从 PI4P 局部产生 (PIP5Ks)，表明 PIP5Ks 参与了整合素的刺激再循环。之前关于这些被研究对象的工作 激酶主要集中在信号传导过程中在质膜上的作用 膜蛋白的内化。拟议的研究将确定需要哪些 PIP5K 同工型 用于刺激整合素回收并确定在内体系统中它们产生 PI(4,5)P2 的位置 启动刺激整合素回收。将采用击倒和救援策略来测试 PIP5K 的影响 整合素回收中的剪接亚型。这些亚型作用的位置将由定位决定 两种异构体及其产物 PI(4,5)P2。在正交方法中，化学诱导二聚化 将用于将 PI(4,5)P2 产生和消耗酶募集到各个内体区室 确定需要在何处局部产生这种脂质以刺激整合素返回细胞表面。 了解导致生长因子刺激的整合素回收的步骤将有助于开发 在不影响整个生长因子信号传导的情况下调节这一过程的干预措施。