Alternative splicing regulation by extracellular matrix mechanics: a self-tuning tool to control cell microenvironmental adaptation and tumor progression

细胞外基质力学的选择性剪接调节：控制细胞微环境适应和肿瘤进展的自调节工具

• 批准号: 9224733
• 负责人: Francois Bordeleau
• 金额: $ 11.25万
• 依托单位: VANDERBILT UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-06-01 至 2019-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9224733
• 关键词: Affect; Alpha Cell; Alternative Splicing; Award; Binding; Biomedical Engineering; Cancer Biology; Cell physiology; Cell-Matrix Junction; Cells; Coupling; Cues; Data; Development; Disease; Endothelial Cells; Engineering; Environment; Epigenetic Process; Event; Exhibits; Extracellular Matrix; Faculty; Feedback; Fingerprint; Focal Adhesions; Genes; Genome; Growth; Growth Factor; Growth Factor Receptors; In Vitro; Institution; Integrins; Intermediate Filaments; Invertebrates; Keratin; Knowledge; Laboratories; Lead; Malignant Neoplasms; Mechanics; Mediating; Mentors; Modeling; Molecular; Molecular and Cellular Biology; Mus; Neoplasm Metastasis; Oncogenic; Pathway interactions; Phase; Phenotype; Physics; Play; Positioning Attribute; Process; Protein Family; Protein Isoforms; Protein Splicing; Proteins; RNA Splicing; Receptor Activation; Regulation; Reporting; Research; Role; Signal Pathway; Signal Transduction; Site; Small Interfering RNA; Therapeutic; Time; Tissues; Training; Translations; Tumorigenicity; Universities; Variant; Vertebrates; Work; base; cancer cell; career development; cell behavior; cell motility; cell type; design; differential expression; genetic regulatory protein; genomic tools; in vivo; innovation; insight; mRNA Export; mechanical properties; mechanotransduction; migration; neoplastic cell; new therapeutic target; novel strategies; novel therapeutics; oncology; physical property; physical science; prevent; programs; protein activation; protein kinase C beta; receptor; research and development; response; symposium; targeted treatment; tool; transglutaminase 2; tumor; tumor growth; tumor progression; tumorigenic

Project Summary/Abstract Over the last decade, much effort has been made to define the relationship between stroma physical properties and cancer malignancy. Notably, stiffer tissue is known to constitute a highly favorable environment for tumor emergence and growth. Elevated extracellular matrix (ECM) stiffness affects several processes associated with tumor progression, including the cell response to growth factors and cell migration. Conversely, tumors also exhibit a marked change in the alternative splicing profiles of several key proteins involved in the cell response to growth factors and cell migration. We recently reported that the mechanical properties of the ECM also influence splicing events, revealing a previously unknown regulatory mechanism that could potentially influence tumor progression. This mechanism was proven to be dependent on cell contractility. Interestingly, elevated cell contractility is also a hallmark of aggressive tumor cells (ref) and we and others have shown that cell contractility is also required for proper growth factor receptor activation (ref). Furthermore, my previous doctoral work and my preliminary data demonstrate that specific proteins, such as keratin intermediate filaments and tissue transglutaminase 2, which are differentially expressed in tumor cells, can act as mechanoregulators and as such, can modulate the contractility of cells. By understanding the specific mechanisms that control the altered state of tumor cell contractility and its influence on alternative splicing regulation, this work will reveal an entirely novel strategy in designing cancer therapeutics. I propose to merge prior training in physics and cellular and molecular biology with new training in biomedical engineering and murine tumor models to further investigate and uncover the role of tumor mechanics in modulating key alternative splicing mechanisms to drive tumor progression. In Aim 1 (K99 phase), I will evaluate the effects of altered tumor cell mechanoregulation on the regulation of alternative splicing and the progression of a metastatic phenotype. I will focus on keratin and tissue transglutaminase 2 which are known to be differentially expressed in tumor cells compared to their normal counterparts. In Aim 2 (K99/R00 phase), I will elucidate how alternative splicing of focal adhesion proteins can influence a cell’s ability to adapt to mechanical cues from the extracellular matrix by using a combination of targeted siRNA molecular tools and an engineered matrix of tunable stiffness. These will be further investigated using in vivo mouse tumor models and genomic tools. In Aim 3 (R00 phase), I will investigate the functional crosstalk between growth factor signaling, matrix stiffness, and alternative splicing regulation in the context of tumor progression, combining the in vitro and in vivo training I will have acquired during the mentoring phase of this award, to point toward a targeted therapeutic approach specific to tumor cells within their stiff microenvironment. This Transition to Independence proposal describes research and career development activities, including mentoring, networking opportunities, conference attendance and course training, which will establish me as a competitive candidate for an independent faculty position and will aid in my development of an innovative, successful research program in the field of mechanobiology at the intersection of Physical Sciences and Oncology. These activities will be mentored by Drs. Cynthia Reinhart-King (primary mentor) and Richard Cerione (co-mentor) at Cornell University, a world-class research institution and leader in field of Physical Sciences and Oncology.

项目总结/摘要 在过去的十年中，已经做了大量的努力来定义基质物理和生物学之间的关系。 性质和癌症恶性程度。值得注意的是，已知较硬的组织构成高度有利的环境 for tumor肿瘤emergency出现and growth生长.细胞外基质（ECM）硬度升高影响几个过程 与肿瘤进展相关，包括细胞对生长因子的反应和细胞迁移。 相反，肿瘤中的几个关键蛋白质的可变剪接模式也发生了显著变化 参与细胞对生长因子的反应和细胞迁移。我们最近报道说， ECM的性质也影响剪接事件，揭示了一种以前未知的调节机制 可能会影响肿瘤的发展这一机制被证明是依赖于细胞 收缩性有趣的是，细胞收缩性升高也是侵袭性肿瘤细胞的标志（参考文献）， 其他人已经表明细胞收缩性也是适当的生长因子受体活化所需要的（参考文献）。 此外，我以前的博士工作和我的初步数据表明，特定的蛋白质，如 角蛋白中间丝和组织转氨酶2，它们在肿瘤细胞中差异表达， 可以作为机械调节剂，并因此可以调节细胞的收缩性。通过了解 控制肿瘤细胞收缩性改变状态的特定机制及其对替代性 这项工作将揭示设计癌症治疗方法的全新策略。 我建议将先前在物理学、细胞和分子生物学方面的培训与生物医学方面的新培训合并起来。 工程和小鼠肿瘤模型，以进一步研究和揭示肿瘤力学在 调节关键的可变剪接机制以驱动肿瘤进展。在目标1（K99阶段），我将 评估改变的肿瘤细胞机械调节对可变剪接调节的影响， 转移表型的进展。我将集中在角蛋白和组织转氨酶2，这是已知的 在肿瘤细胞中与正常细胞相比有差异表达。在目标2（K99/R 00阶段）中， 将阐明粘着斑蛋白的选择性剪接如何影响细胞适应 通过使用靶向siRNA分子工具的组合， 一个可调刚度的工程矩阵。这些将使用体内小鼠肿瘤模型进行进一步研究 和基因组工具。在目标3（R 00阶段），我将研究生长因子之间的功能串扰， 信号传导、基质硬度和可变剪接调节在肿瘤进展的背景下，结合 在体外和体内的培训，我将获得在指导阶段的这个奖项，指向一个 靶向治疗方法特异性针对肿瘤细胞在其坚硬的微环境中。 这份向独立过渡的提案描述了研究和职业发展活动，包括 指导，网络机会，会议出席和课程培训，这将使我成为一个 一个独立的教师职位的竞争力的候选人，并将在我的创新， 在机械生物学领域的成功研究计划，在物理科学和 肿瘤科这些活动将由Cynthia Reinhart-King博士（主要导师）和Richard Cerione（共同导师）康奈尔大学，世界级的研究机构和物理领域的领导者 科学和肿瘤学。