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.

摘要 混合谱系激酶（MLK）3是促分裂原活化蛋白激酶（MAPK）激酶激酶（MAP 3 K）， 在哺乳动物细胞中调节多种MAPK信号通路。此外，MLK 3已被证明 调节不同类型的人上皮细胞（正常和癌症）的增殖和侵袭，包括 卵巢和乳房。在目前的研究中，我们发现MLK 3在卵巢癌细胞的细胞间粘附中具有新的功能， 上皮TOV 112 D肿瘤细胞。MLK 3 siRNA敲低破坏了TOV 112 D球状体中的细胞-细胞粘附， 减少了细胞连接蛋白，E-钙粘蛋白，并显着增加了鼻涕虫的水平，一个转录 E-钙粘蛋白阻遏物。此外，MLK 3在TOV 112 D细胞中与E-cadherin共沉淀 这表明MLK 3可能与粘附连接蛋白复合物相关。此外，本发明还 MLK 3过表达诱导E-钙粘蛋白的切割，产生80 kDa可溶形式的E-钙粘蛋白， 已发现其激活生长因子受体信号传导。在TOV 112 D细胞中，MLK 3过表达 还强烈诱导了N-钙粘蛋白的表达，这是一种细胞连接蛋白，通常在细胞中表达。 间充质细胞我们的中心假设是MLK 3通过多种途径调节E-cadherin和N-cadherin 这种调节对于卵巢上皮细胞的适当细胞粘附和迁移至关重要。 设计所提出的实验以定义MLK 3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。三个具体 目的是提出来测试的假设。在目的1中，我们将研究MLK 3在E-cadherin多表达中的功能。 蛋白复合物和卵巢球状体细胞-细胞粘附。在目标2中，我们将研究MLK 3依赖 调节Slug和E-cadherin的表达。在目标3中，我们将评估MLK 3依赖的E- 钙粘蛋白裂解、N-钙粘蛋白表达与卵巢球状体结构这项研究将揭示新的 深入了解MLK 3调节E-和N-钙粘蛋白以及调节细胞间粘附的机制， 粘附和迁移，对发育、组织形态发生、迁移和 癌细胞侵袭