Analyzing Adhesion and Signaling Functions for PTPN12 in Invasive Glioma Cells

分析侵袭性胶质瘤细胞中 PTPN12 的粘附和信号传导功能

• 批准号: 10543815
• 负责人: Joseph H McCarty
• 金额: $ 40.1万
• 依托单位: UNIVERSITY OF TX MD ANDERSON CAN CTR
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-01-01 至 2026-12-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10543815
• 关键词: 3-Dimensional; Address; Adhesions; Amino Acid Motifs; Angiogenesis Inhibitors; BCAR1 gene; Biochemical; Brain; Brain Neoplasms; Cell Culture System; Cell Proliferation; Cells; Chemotherapy and/or radiation; Clinical; Clustered Regularly Interspaced Short Palindromic Repeats; Complex; Cullin Proteins; Data; Data Correlations; Equilibrium; Event; Excision; Focal Adhesions; Genes; Genetic; Genetically Engineered Mouse; Glioblastoma; Glioma; Gliomagenesis; Growth; Human; Invaded; Lesion; Ligase; Link; Malignant - descriptor; Malignant neoplasm of brain; Mediating; Modality; Modeling; Molecular; Mutate; Oncogenes; Operative Surgical Procedures; Outcome; PTPN12 gene; Pathway interactions; Patients; Pattern; Pest Control; Phosphoric Monoester Hydrolases; Phosphorylation; Population; Protein Dephosphorylation; Protein Tyrosine Phosphatase; Proteins; Proteomics; Recurrence; Recurrent tumor; Refractory; Reporting; Resected; Resistance; Resolution; Role; Sampling; Scaffolding Protein; Signal Pathway; Signal Transduction; Signaling Protein; Site; System; Tertiary Protein Structure; Testing; Tumor Cell Invasion; Tumor Promotion; Tumor Suppressor Proteins; Tyrosine; Ubiquitin; Ubiquitination; bevacizumab; cancer cell; cell growth; chemoradiation; chemotherapy; effective therapy; experimental study; fluorescence imaging; genetic approach; in vivo Model; inducible gene expression; inhibitor; live cell imaging; mouse model; multicatalytic endopeptidase complex; neoplastic cell; patient response; pharmacologic; pre-clinical; protein complex; protein expression; recruit; temozolomide; treatment response; tumor; tumor growth; tumor microenvironment; tumor progression; valosin-containing protein

Glioblastoma (GBM) is a malignant brain cancer that is resistant to all treatment modalities. This resistance is due, in large part, to a population of high invasive and low proliferative cancer cells that elude surgical resection and are refractory to chemotherapy and radiation. While a great deal is known about oncogenes, tumor suppressors, and other pathways that promote GBM cell proliferation, we understand relatively little about mechanisms that drive GBM cell invasion in the brain microenvironment. Therefore, the PI's group performed genetic and biochemical screens to identify adhesion and signaling factors that regulate invasive cell growth in GBM. These efforts identified the non-receptor protein tyrosine phosphatase PTP-PEST/PTPN12 as a critical signaling effector in invasive GBM cells. Here, we present a significant amount of supporting data showing that PTP-PEST promotes GBM cell invasion by regulating the stability of key focal adhesion signaling proteins, particularly Crk-associated substrate (p130Cas). In particular, we have discovered that PTP-PEST in focal adhesions mediates interactions between p130Cas and valosin containing protein (Vcp), a ubiquitin-dependent segregase and key component of the ubiquitin proteasome system. These findings have led to our working hypothesis that PTP-PEST is essential for GBM cell invasion by regulating the phosphorylation- dependent ubiquitination of focal adhesion protein substrates. To test this hypothesis, we will (1) characterize protein domains and motifs that mediate interactions between PTP-PEST, p130Cas, and Vcp, as well as determine how these interactions modulate focal adhesion protein stability in GBM cells; (2) identify PTP-PEST-generated phosphodegron sequences in p130Cas and determine how they regulate focal adhesion dynamics by recruiting Vcp and facilitating p130Cas degradation by the proteasome; (3) genetically mutate Vcp at specific sites required for the phosphorylation-dependent ubiquitination of p130Cas and analyze cell invasion using three-dimensional culture systems and pre-clinical mouse models of GBM; and (4) quantify levels and spatial patterns of PTP-PEST signaling via p130Cas and Vcp in human GBM samples and primary cancer cell culture systems. We will correlate these data with patient survival as well as response to therapies such as temozolomide and bevacizumab. Collectively, these experiments will not only elucidate signaling pathways that control the GBM cell invasive state, but may lead to new strategies to target invasive cells and block tumor progression.

胶质母细胞瘤（GBM）是一种恶性脑癌，对所有治疗方式具有抗性。这 在很大程度上，抗药性归因于高侵入性和低增生性癌细胞的群体 那种避免的手术切除，对化学疗法和放射线具有难治性。虽然很棒 关于促进GBM细胞的肿瘤基因，肿瘤抑制剂和其他途径已知的交易已知 扩散，我们对驱动GBM细胞入侵的机制相对较少了解 大脑微环境。因此，PI的组进行了遗传和生化 筛选以鉴定调节GBM中侵入性细胞生长的粘附和信号传导因子。 这些努力将非受体蛋白酪氨酸磷酸酶PTP-PEST/PTPN12确定为A 侵入性GBM细胞中的关键信号效应子。在这里，我们提出了大量的 支持数据，表明PTP-PEST通过调节稳定性来促进GBM细胞侵袭 关键局灶性粘附信号蛋白，特别是与CRK相关的底物（P130CAS）。在 特别是，我们发现焦点粘连中的PTP-pest介导了相互作用 在P130CA和含有蛋白质（VCP）的P130CA之间，泛素依赖性分离酶和 泛素蛋白酶体系统的关键组成部分。这些发现导致了我们的工作 假设PTP-PEST通过调节磷酸化 - 局部粘附蛋白底物的依赖泛素化。为了检验这一假设，我们将（1） 表征蛋白质结构域和介导PTP-PEST之间相互作用的基序 P130CA和VCP，并确定这些相互作用如何调节局灶性粘附 GBM细胞中的蛋白质稳定性； （2）识别PTP-PEST生成的磷降射序列 P130CAS并通过招募VCP和 蛋白酶体促进P130CAS降解； （3）在特定部位的基因突变VCP P130CA的磷酸化依赖性泛素化所必需的并分析细胞侵袭 使用三维培养系统和GBM的临床前小鼠模型； （4） 通过P130CA和VCP在人类中量化PTP-pest信号的水平和空间模式 GBM样品和原发性癌细胞培养系统。我们将将这些数据与患者相关联 生存以及对替莫唑酚和贝伐单抗等疗法的反应。共同 这些实验不仅将阐明控制GBM细胞侵入性的信号通路 状态，但可能导致靶向侵入性细胞并阻止肿瘤进展的新策略。