Cytohesin dependent ARF to Rac signaling in HGF mediated motility

HGF 介导的运动中细胞粘附素依赖性 ARF 至 Rac 信号传导

• 批准号: 9132250
• 负责人: Lorraine C Santy
• 金额: $ 29.05万
• 依托单位: PENNSYLVANIA STATE UNIVERSITY, THE
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-09-20 至 2018-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9132250
• 关键词: Acute; Acute Renal Failure with Renal Papillary Necrosis; Affect; Area; Binding; Cell Shape; Cell Survival; Cells; Cessation of life; Complication; Cytosol; Data; Development; Epithelial; Epithelial Cells; Event; Fibrosis; Goals; Growth; Guanosine Triphosphate Phosphohydrolases; HGF gene; Hospitals; Immigration; Injury; Kidney; Knowledge; Link; Lipids; Mediating; Membrane; Modeling; Modification; Molecular; Molecular Conformation; Monomeric GTP-Binding Proteins; Movement; Mutagenesis; Neoplasm Metastasis; Normal Cell; Pathway interactions; Patients; Phosphatidylinositols; Phosphorylation; Process; Production; Publishing; Recovery; Recruitment Activity; Renal function; Reperfusion Injury; Research; Role; Scaffolding Protein; Signal Pathway; Signal Transduction; Small Interfering RNA; Staging; Surface; Testing; Work; Wound Healing; base; cell motility; fighting; kidney repair; knock-down; migration; mutant; prevent; protein protein interaction; renal ischemia; repaired; research study; response; scaffold; therapeutic target; therapy design; therapy development; tool development; tumor

DESCRIPTION (provided by applicant): Acute kidney injury is a common hospital complication that leads death or long-term kidney damage if recovery is not complete. Hepatocyte growth factor (HGF) promotes the recovery from kidney damage at multiple steps. While many of the proximal signals activated by HGF and the downstream targets that remodel cell shape in response to HGF are known, the details of the circuits that link them together are unclear. This limits the possibility of selectively modulating HGF signaling without impacting the actions of other growth factors. Work in the applicant's lab has recently demonstrated that protein- protein interactions are critical for building a pro-migratory circuit that links activatin of Arf6 by cytohesin-2 to the downstream activation of Rac by Dock180. Preliminary data suggest that this signaling circuit is required for HGF-stimulated Rac activation and migration. The overall objective of this application is to determine the molecular details of this cytohesin-dependent Arf-to-Rac signaling module and to test the degree to which it mediates HGF-induced motility and the recovery from acute kidney injury. This is the next step toward the long-term goal of understanding the initiation of motility in epithelial cells by small GTPase cascades. This application will test a hypothesized model for the cytohesin-dependent Arf-to-Rac signaling module. This module will be tested by pursuing 4 specific aims: 1) Determine how cytohesin-2 activity is controlled; 2) Determine the mechanism of interaction of cytohesin-2 and Dock180 and test the degree to which this interaction mediates HGF dependent Rac activation; 3) Determine the role of active Arf6 in cytohesin-dependent Rac activation; 4) Determine the extent to which cytohesin-dependent Rac activation regulates HGF-stimulated wound healing and recovery of the kidney from ischemia reperfusion injury. Aim 1 will test the hypothesis that cytohesin-2 intramolecular interactions prevent its interaction with membranes until these interactions are disrupted by phosphorylation. Aim 2 will use mutagenesis to identify the regions necessary for the interaction of the cytohesin-binding scaffold GRASP with Dock180. The importance of this interaction for HGF- induced Rac activation will be tested using knockdown of GRASP and expression of GRASP mutants. Aim 3 will test the hypothesis that Arf6-dependent lipid modifications promote activation of Rac by Dock180. Aim 4 will use siRNA, mutant and pharmacologic modulation of cytohesin-dependent Arf-to- Rac signaling to test its involvement in HGF-stimulated wound healing and recovery of the kidney from ischemia reperfusion injury. A proven model of this pro-migratory circuit will allow the design of interventions to modulate this circuit without impacting other Arf6 or Rac-dependent functions. Such treatments could promote or inhibit HGF-dependent migration without disrupting the actions of a broad range of growth factors.

描述（由申请人提供）：急性肾损伤是一种常见的医院并发症，如果恢复不完全，会导致死亡或长期肾脏损害。肝细胞生长因子 (HGF) 通过多个步骤促进肾脏损伤的恢复。虽然许多由 HGF 激活的近端信号和响应 HGF 重塑细胞形状的下游靶标是已知的，但将它们连接在一起的电路的细节尚不清楚。这限制了选择性调节 HGF 信号而不影响其他生长因子作用的可能性。申请人实验室的工作最近表明，蛋白质-蛋白质相互作用对于构建促迁移回路至关重要，该回路将细胞粘连蛋白-2对Arf6的激活与Dock180对Rac的下游激活连接起来。初步数据表明，该信号通路是 HGF 刺激的 Rac 激活和迁移所必需的。本申请的总体目标是确定这种细胞粘附素依赖性 Arf-to-Rac 信号传导模块的分子细节，并测试其介导 HGF 诱导的运动和急性肾损伤恢复的程度。这是实现了解小 GTP 酶级联启动上皮细胞运动的长期目标的下一步。 该应用将测试细胞粘附素依赖性 Arf-to-Rac 信号传导模块的假设模型。该模块将通过追求 4 个具体目标进行测试：1) 确定如何控制 cytohesin-2 活性； 2）确定cytohesin-2和Dock180相互作用的机制，并测试这种相互作用介导HGF依赖性Rac激活的程度； 3）确定活性Arf6在细胞粘附素依赖性Rac激活中的作用； 4) 确定细胞粘附素依赖性 Rac 激活在多大程度上调节 HGF 刺激的伤口愈合和肾脏从缺血再灌注损伤中的恢复。目标 1 将检验以下假设：细胞粘附素 2 分子内相互作用阻止其与膜的相互作用，直到这些相互作用被磷酸化破坏。目标 2 将使用诱变来鉴定细胞粘连蛋白结合支架 GRASP 与 Dock180 相互作用所需的区域。这种相互作用对于 HGF 诱导的 Rac 激活的重要性将通过 GRASP 的敲低和 GRASP 突变体的表达来测试。目标 3 将检验 Arf6 依赖性脂质修饰促进 Dock180 激活 Rac 的假设。目标 4 将使用 siRNA、细胞粘附素依赖性 Arf-to-Rac 信号传导的突变体和药理学调节来测试其在 HGF 刺激的伤口愈合和肾脏从缺血再灌注损伤中恢复的作用。这种促迁移回路的经过验证的模型将允许设计干预措施来调节该回路，而不影响其他 Arf6 或 Rac 相关功能。此类治疗可以促进或抑制 HGF 依赖性迁移，而不破坏多种生长因子的作用。

项目成果

The cytohesin coiled-coil domain interacts with threonine 276 to control membrane association.

• DOI: 10.1371/journal.pone.0082084
• 发表时间: 2013
• 期刊: PloS one
• 影响因子: 3.7
• 作者: Hiester KG;Santy LC
• 通讯作者: Santy LC

The scaffolding protein GRASP/Tamalin directly binds to Dock180 as well as to cytohesins facilitating GTPase crosstalk in epithelial cell migration.

• DOI: 10.1186/1471-2121-14-9
• 发表时间: 2013-02-26
• 期刊: BMC cell biology
• 作者: Attar MA;Santy LC
• 通讯作者: Santy LC

ARF-GEF cytohesin-2/ARNO regulates R-Ras and α5-integrin recycling through an EHD1-positive compartment.

• DOI: 10.1091/mbc.e15-05-0278
• 发表时间: 2015-11-15
• 期刊: Molecular biology of the cell
• 影响因子: 3.3
• 作者: Salem JC;Reviriego-Mendoza MM;Santy LC
• 通讯作者: Santy LC

A traction force threshold signifies metastatic phenotypic change in multicellular epithelia.

• DOI: 10.1039/c9sm00733d
• 发表时间: 2019-09
• 期刊: Soft matter
• 影响因子: 3.4
• 作者: Yao Zhang;Xuechen Shi;Tiankai Zhao;Changjin Huang;Qiong Wei;Xin Tang;Lorraine C. Santy;M. Taher A. Saif;Sulin Zhang

The cytohesin guanosine exchange factors (GEFs) are required to promote HGF-mediated renal recovery after acute kidney injury (AKI) in mice.

• DOI: 10.14814/phy2.12442
• 发表时间: 2015-06
• 期刊: Physiological reports
• 影响因子: 2.5
• 作者: Reviriego-Mendoza MM;Santy LC
• 通讯作者: Santy LC