Tissue-Regulated Lineage-Specific Splicing in Glioblastoma Pathogenesis

胶质母细胞瘤发病机制中组织调节的谱系特异性剪接

• 批准号: 10409836
• 负责人: Markus Bredel
• 金额: $ 34.9万
• 依托单位: UNIVERSITY OF ALABAMA AT BIRMINGHAM
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-01 至 2024-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10409836
• 关键词: Address; Adult; Affect; Alternative Splicing; Amino Acids; Annexins; Attenuated; Binding; Brain; Brain Neoplasms; Cell Lineage; Cells; Clinical; Complex; Data; Development; Disease; Endocytosis; Epidermal Growth Factor Receptor; Exclusion; Exocytosis; Exons; Foundations; Future; Genetic; Glioblastoma; Glioma; Knowledge; Length; Malignant - descriptor; Malignant Glioma; Mediating; Membrane; Mission; Mitotic; Modeling; N-terminal; Neoplasms; Neurons; Nuclear Pore Complex; Oncogenic; PDGFRA gene; Pathogenesis; Pathway interactions; Patients; Pattern; Polypyrimidine Tract-Binding Protein; Prevalence; Process; Protein Isoforms; Protein Tyrosine Kinase; Public Health; RNA Binding; RNA Splicing; Receptor Protein-Tyrosine Kinases; Recycling; Regulation; Research; Role; Signal Transduction; Sorting - Cell Movement; Testing; Tissues; Tumor Suppressor Genes; Tumor Suppressor Proteins; Tumorigenicity; United States National Institutes of Health; Variant; Work; Xenograft procedure; base; cell growth; epidermal growth factor receptor VIII; exon skipping; extracellular; extracellular vesicles; gain of function; in vivo; knock-down; mRNA Precursor; neoplastic cell; novel; novel therapeutics; precursor cell; prevent; progenitor; prototype; receptor; receptor recycling; relating to nervous system; response; stem cells; targeted treatment; therapeutic target; trafficking; trait; treatment response; tumor; tumorigenesis; tumorigenic; vesicle transport; virtual

PROJECT SUMMARY Lineage-specific alternative splicing, where splicing occurs in a tissue-regulated manner involving evolutionary conserved alternative exons, has a determinative role in brain development. The role of tissue-specific alternative exons in malignant transformation and tumor development is undefined. The impact of lineage- specific splicing on glioblastoma (GBM) pathogenesis remains unclear but is conceptually attractive given the prevalence of this process in determining the fate of ancestral cells of potential GBM origin. We found that brain-enriched splice factor PTBP1 mediates lineage-specific alternative splicing of the ANXA7 tumor suppressor gene. PTBP1 is expressed in neural and glial precursor cells (NPCs/GPCs) and binds ANXA7 pre-mRNA to skip exon 6; this produces spliced ANXA7 isoform 2 (I2). During lineage specification, PTBP1 is downregulated and this allows the expression of unspliced ANXA7 I1 (I1), which includes exon 6. We determined that the patterned expression of the I2 splice variant in the brain is restricted to lineages that represent potential GBM cells of origin but that I1 is virtually absent in these cells. Our preliminary data illustrate that lineage-specific splicing can augment genetic mechanisms to deregulate oncogenic pathways. Specifically, we showed ANXA7 I1, but not I2, targets oncogenic receptor tyrosine kinases (RTKs) such as EGFR, MET and PDGFRA for endosomal degradation. In the adult brain, PTBP1 is nearly absent; in GBM, PTBP1 aberrantly persists and RTK levels are elevated. However, we demonstrate that upon PTBP1 knockdown and/or, ANXA7 I1 re-expression, RTK signaling is reduced, and GBM tumorigenicity is diminished. Therefore, we hypothesize that GBMs benefit from persistent PTBP1 expression as this suppresses ANXA7 I1, prevents RTK endosomal degradation, and sustains RTK signaling in GBM. Our Specific Aims are: 1. Define the Mechanism by which ANXA7 I1 Regulates Endosomal and Cellular Trafficking of EGFR. Herein, we will determine the detailed mechanism by which ANXA7 I1 interacts with and regulates endosomal and extracellular vesicle transport of EGFR as a prototype RTK. 2. Demonstrate ANXA7 I1 is a Signaling Modulator for Multiple RTKs and RTK-Targeted Therapies by Perpetuating Endosomal Degradation. Herein, we will demonstrate that ANXA7 I1 is competent to attenuate signaling through multiple pro-tumorigenic RTKs, modulate TKI response, and thus has a broad impact on GBM in vivo. These mechanistic and focused aims are needed to better define the role of lineage-specific splicing processes in tumorigenesis, which remains understudied. Our results will determine that ANXA7 I1 is sufficient to simultaneously inhibit signaling by multiple RTKs and thus holds potential to reveal a new therapeutic direction. This work will define the role of aberrant lineage-specific alternative splicing in GBM pathogenesis, offer a broader understanding of this process in malignant transformation, and provide a potential foundation for future studies to therapeutically target developmentally regulated splicing factors.

项目摘要 谱系特异性选择性剪接，其中剪接以组织调节的方式发生，涉及进化 保守的替代外显子，在脑发育中具有决定性作用。组织特异性 恶性转化和肿瘤发展中的替代外显子尚未确定。血统的影响- 胶质母细胞瘤（GBM）发病机制的特异性剪接仍不清楚，但在概念上很有吸引力 考虑到这一过程在决定潜在GBM起源的祖细胞命运中的普遍性。 我们发现脑富集剪接因子PTBP 1介导ANXA 7的谱系特异性选择性剪接， 肿瘤抑制基因PTBP 1在神经和神经胶质前体细胞（NPC/GPC）中表达，并结合 ANXA 7前mRNA跳过外显子6;这产生剪接的ANXA 7同种型2（I2）。在血统规范期间， PTBP 1被下调，这允许表达未剪接的ANXA 7 I1（I1），包括外显子6。我们 确定了I2剪接变体在脑中的模式化表达仅限于 代表起源的潜在GBM细胞，但I1在这些细胞中几乎不存在。我们的初步数据 说明谱系特异性剪接可以增强遗传机制， 途径。具体来说，我们发现ANXA 7 I1，而不是I2，靶向致癌受体酪氨酸激酶（RTK） 例如EGFR、MET和PDGFRA用于内体降解。在成人大脑中，PTBP 1几乎不存在; GBM、PTBP 1异常持续，RTK水平升高。然而，我们证明，在PTBP 1 敲低和/或ANXA 7 I1再表达，RTK信号传导减少，GBM致瘤性降低。 因此，我们假设GBM受益于持续的PTBP 1表达，因为这抑制了GBM的表达。 ANXA 7 I1防止RTK内体降解，并维持GBM中的RTK信号传导。我们的具体目标 是：1.定义ANXA 7 I1调节内体和细胞运输的机制 EGFR对在此，我们将确定ANXA 7 I1与细胞相互作用并调节细胞凋亡的详细机制。 作为原型RTK的EGFR的内体和细胞外囊泡转运。2.证明ANXA 7 I1是一个 用于多种RTK的信号调节剂和通过持续内体的RTK靶向治疗 降解在此，我们将证明ANXA 7 I1能够通过多种途径减弱信号传导， 促肿瘤发生RTK调节TKI应答，因此对体内GBM具有广泛影响。这些 需要机械的和集中的目标来更好地定义谱系特异性剪接过程在以下方面的作用： 肿瘤发生，这仍然是研究不足。我们的结果将确定ANXA 7 I1足以 同时抑制多个RTK的信号传导，因此具有揭示新的治疗方向的潜力。 这项工作将确定异常谱系特异性选择性剪接在GBM发病机制中的作用， 提供了一个更广泛的理解这一过程中的恶性转化，并提供了一个潜在的 为将来的研究奠定了基础，以治疗为目标的发育调控剪接因子。