Regulation of Cadherins by MLK3

MLK3 对钙粘蛋白的调节

• 批准号: 10793060
• 负责人: Deborah N Chadee
• 金额: $ 45.15万
• 依托单位: UNIVERSITY OF TOLEDO
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-22 至 2026-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10793060
• 关键词: Adherens Junction; Adhesions; Biological Process; Breast; Cadherins; Carcinoma; Cell Adhesion; Cell-Cell Adhesion; Cells; Co-Immunoprecipitations; Complex; Development; E-Cadherin; Epithelial Cells; Epithelium; Experimental Designs; Gene Expression; Growth Factor Receptors; Human; Intercellular Junctions; Invaded; MAP Kinase Kinase Kinase; Malignant Neoplasms; Mammalian Cell; Matrix Metalloproteinases; Mediating; Mesenchymal; Mitogen-Activated Protein Kinases; Morphogenesis; Multiprotein Complexes; N-Cadherin; Neurofibromin 2; Ovarian; Phosphorylation; Phosphotransferases; Process; Proliferating; Protein Kinase; Proteins; Receptor Signaling; Regulation; Research; Role; Signal Pathway; Small Interfering RNA; Structure; Testing; Tissues; Transcription Repressor; beta catenin; cancer cell; cell motility; insight; knock-down; migration; mixed lineage kinase 3; neoplastic cell; novel; ovarian neoplasm; overexpression; protein complex; slug

ABSTRACT Mixed lineage kinase (MLK) 3 is a mitogen-activated protein kinase (MAPK) kinase kinase (MAP3K) that regulates multiple MAPK signaling pathways in mammalian cells. In addition, MLK3 has been demonstrated to regulate proliferation and invasion of different types of human epithelial cells (normal and cancer) including ovarian and breast. In the current study, we identified a novel function for MLK3 in cell-cell adhesion of ovarian epithelial TOV112D tumor cells. MLK3 siRNA knockdown disrupted cell-cell adhesion in TOV112D spheroids, reduced the cell junction protein, E-cadherin, and dramatically increased the level of Slug, a transcriptional repressor of E-cadherin. Furthermore, MLK3 was co-immunoprecipitated with E-cadherin in TOV112D cell lysates, which indicates that MLK3 may be associated with adherens junction protein complexes. Additionally, MLK3 overexpression induced the cleavage of E-cadherin resulting in an 80 kDa soluble form of E-cadherin, which has been found to activate growth factor receptor signaling. In TOV112D cells, MLK3 overexpression also strongly induced the expression of N-cadherin, a cell junction protein that is normally expressed in mesenchymal cells. Our central hypothesis is that MLK3 regulates E-cadherin and N-cadherin through multiple mechanisms, and this regulation is essential for proper cell adhesion and migration in ovarian epithelial cells. The proposed experiments are designed to define the mechanism(s) by which MLK3 regulates E- and N- cadherin, and the impact of this regulation on ovarian cell spheroid adhesion and migration. Three specific aims are proposed to test the hypothesis. In Aim 1, we will investigate MLK3 functions in E-cadherin multi- protein complexes and ovarian spheroid cell-cell adhesion. In aim 2, we will examine MLK3-dependent regulation of Slug and E-cadherin expression. In Aim 3, we will evaluate MLK3-dependent regulation of E- Cadherin cleavage, N-Cadherin expression and ovarian spheroid structure. This research will reveal novel insight into the mechanisms by which MLK3 functions to regulate E- and N-cadherin and modulate cell-cell adhesion and migration, processes which are essential for development, tissue morphogenesis, migration and cancer cell invasion.

抽象的 混合谱系激酶 (MLK​​) 3 是一种丝裂原激活蛋白激酶 (MAPK) 激酶 (MAP3K)， 调节哺乳动物细胞中的多个 MAPK 信号通路。此外，MLK3 已被证明可以 调节不同类型人类上皮细胞（正常和癌症）的增殖和侵袭，包括 卵巢和乳房。在当前的研究中，我们发现了 MLK3 在卵巢细胞间粘附中的新功能。 上皮TOV112D肿瘤细胞。 MLK3 siRNA 敲低破坏了 TOV112D 球体中的细胞间粘附， 减少细胞连接蛋白 E-钙粘蛋白，并显着增加转录因子 Slug 的水平 E-钙粘蛋白的阻遏物。此外，MLK3 与 E-cadherin 在 TOV112D 细胞中进行免疫共沉淀 裂解物，这表明 MLK3 可能与粘附连接蛋白复合物相关。此外， MLK3 过表达诱导 E-钙粘蛋白裂解，产生 80 kDa 可溶形式的 E-钙粘蛋白， 已发现它可以激活生长因子受体信号传导。在 TOV112D 细胞中，MLK3 过度表达 还强烈诱导 N-钙粘蛋白的表达，这是一种细胞连接蛋白，通常在 间充质细胞。我们的中心假设是 MLK3 通过多种方式调节 E-钙粘蛋白和 N-钙粘蛋白 机制，这种调节对于卵巢上皮细胞的正确细胞粘附和迁移至关重要。 所提出的实验旨在定义 MLK3 调节 E- 和 N- 的机制 钙粘蛋白，以及这种调节对卵巢细胞球体粘附和迁移的影响。三具体 提出了测试假设的目标。在目标 1 中，我们将研究 MLK3 在 E-cadherin multi- 蛋白质复合物和卵巢球体细胞间粘附。在目标 2 中，我们将检查 MLK3 依赖性 Slug 和 E-钙粘蛋白表达的调节。在目标 3 中，我们将评估 MLK3 依赖性 E- 钙粘蛋白裂解、N-钙粘蛋白表达和卵巢球体结构。这项研究将揭示新颖的 深入了解 MLK3 调节 E-和 N-钙粘蛋白以及调节细胞间的机制 粘附和迁移，对于发育、组织形态发生、迁移和发育至关重要的过程 癌细胞侵袭。