Mechanisms guiding the fibrillar assembly of SNED1 in the extracellular matrix

指导细胞外基质中 SNED1 纤维组装的机制

• 批准号: 10733534
• 负责人: Alexandra Naba
• 金额: $ 29.88万
• 依托单位: UNIVERSITY OF ILLINOIS AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-08-04 至 2027-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10733534
• 关键词: Address; Adhesions; Adhesives; Antibodies; Architecture; Binding; Binding Sites; Biochemical; Biological Assay; Breast Cancer Patient; Cardiovascular Diseases; Cell Adhesion; Cell Line; Cell Proliferation; Cell physiology; Cell surface; Cells; Collagen; Complex; Confocal Microscopy; Connective Tissue Diseases; Coupled; Craniofacial Abnormalities; Data; Defect; Development; Developmental Process; Disease; Disease Progression; EGF gene; EGF-Like Domain; Electron Microscopy; Embryonic Development; Essential Genes; Extracellular Matrix; Extracellular Matrix Proteins; Fiber; Fibroblasts; Fibronectins; Fibrosis; Future; Genetic; Goals; Homeostasis; Immunofluorescence Immunologic; In Vitro; Integrin Binding; Integrins; Interferometry; Knock-out; Knockout Mice; Knowledge; Label; Laboratories; Lead; Link; Malignant Neoplasms; Mammary Neoplasms; Maps; Mass Spectrum Analysis; Mediating; Metabolic; Microscopy; Musculoskeletal Diseases; Mutation; Neonatal; Neoplasm Metastasis; Neural Crest Cell; Nidogen; Organ; Patients; Phenotype; Prognosis; Property; Protein Biochemistry; Proteins; Proteomics; Role; Sequence Analysis; Signal Transduction; Structure; Sushi Domain; System; Tertiary Protein Structure; Testing; Therapeutic; Time; Tissues; Variant; Work; adhesion receptor; aggressive breast cancer; cell behavior; cell motility; clinically relevant; craniofacial development; deoxycholate; dimer; fibrillin; insight; interest; migration; molecular modeling; mouse model; neoplastic cell; novel; pharmacologic; prevent; promoter; protein complex; protein purification; receptor; scaffold; therapeutic development; tool; tumor progression

Project Summary The extracellular matrix (ECM) is a complex protein meshwork that constitutes the architectural scaffold of all tissues. In addition to its structural role, the ECM conveys biochemical signals to cells interpreted by receptors, like integrins, and controlling diverse cellular functions including adhesion and migration. The ECM is thus a key regulator of developmental processes and tissue homeostasis. Consequently, alterations in the composition and assembly of the ECM meshwork have been linked to a plethora of diseases including fibrosis, and cancer. Important progress toward understanding how the ECM meshwork is built and how the ECM govern cellular phenotypes have been made by studying major ECM proteins such as fibronectin and collagens. However, using sequence analysis, we have predicted that nearly 300 proteins can contribute to the ECM meshwork. Knowledge regarding the roles of these other ECM proteins remains preliminary. This represents a significant gap in our understanding of the ECM and in our ability to correct disease-causing ECM defects. We have recently become interested in one of these understudied ECM proteins, SNED1, after having found that it was associated with more highly aggressive breast cancers. Of clinical relevance, we found that higher SNED1 expression correlated with a worse prognosis for breast cancer patients. To gain insights into SNED1’s functions, we generated the first knockout (KO) mouse model of Sned1 and showed that Sned1 is an essential gene, since its KO resulted in early neonatal lethality due, in part, to craniofacial malformations. Importantly, we recently identified the first patients with SNED1 variants and they present with craniofacial malformations. Despite these observations, the mechanisms by which SNED1 contributes to embryonic development and cancer metastasis are unknown. Using novel tools we developed (antibodies, mouse models, cell lines, purified proteins), we have shown that SNED1 forms fibers within the ECM scaffold and contributes to its overall organization. In this proposal, we will test the hypothesis that SNED1 assembly in the ECM and its role in regulating ECM architecture depend on SNED1’s interactions with other ECM proteins and integrins. Leveraging our unique toolkit and combining our unique expertise in ECM protein biochemistry and ECM proteomics with state-of-the-art microscopy, we will conduct a time-resolved structure/function analysis to map which domains of SNED1 mediate its incorporation in the ECM (Aim 1); determine the role of ECM proteins/SNED1 interactions in SNED1 ECM assembly (Aim 2); and identify the SNED1 receptors at the cell surface governing SNED1-dependent ECM organization and responsible for the adhesive property of SNED1 (Aim 3)? Our goal is to fill critical gaps in our understanding of the fundamental mechanisms leading to ECM assembly, with respect to SNED1. This is a necessary step toward deciphering how perturbations of these mechanisms can lead to developmental defects and cancer progression. This work will also pave the way to the development of future therapeutic strategies to correct disease-causing alterations in ECM structure.

项目摘要 细胞外基质（ECM）是一种复杂的蛋白质网络，构成了所有细胞的结构支架。 组织中除了其结构作用外，ECM还将生物化学信号传递给由受体解释的细胞， 如整联蛋白，并控制多种细胞功能，包括粘附和迁移。因此，ECM是一个关键 发育过程和组织内环境稳定的调节器。因此，组成和 ECM网络的组装与包括纤维化和癌症在内的多种疾病有关。 了解ECM网络如何构建以及ECM如何管理细胞的重要进展 通过研究主要的ECM蛋白质如纤连蛋白和胶原蛋白已经确定了表型。但使用 通过序列分析，我们已经预测了近300种蛋白质可以有助于ECM网络。知识 关于这些其他ECM蛋白的作用仍然是初步的。这代表了我们的一个重大差距， 对ECM的理解以及我们纠正致病ECM缺陷的能力。我们最近 在发现SNED 1与细胞外基质蛋白质的合成有关后， 更高侵袭性的乳腺癌在临床相关性方面，我们发现SNED 1的高表达与 乳腺癌患者的预后更差。为了深入了解SNED 1的功能，我们生成了 Sned 1的第一个敲除（KO）小鼠模型，并表明Sned 1是一个必需的基因，因为它的KO导致 在早期新生儿死亡中，部分原因是颅面畸形。重要的是，我们最近发现了第一个 SNED 1变异的患者，他们表现为颅面畸形。尽管有这些意见， SNED 1促进胚胎发育和癌症转移的机制尚不清楚。 使用我们开发的新工具（抗体，小鼠模型，细胞系，纯化蛋白质），我们已经证明， SNED 1在ECM支架内形成纤维并有助于其整体组织。在本提案中，我们将 检验SNED 1在ECM中的组装及其在调节ECM结构中的作用取决于以下假设： SNED 1与其他ECM蛋白和整合素的相互作用。利用我们独特的工具包， 我们将利用最先进的显微镜技术，在ECM蛋白质生物化学和ECM蛋白质组学方面拥有独特的专业知识， 进行时间分辨结构/功能分析，以绘制SNED 1的哪些结构域介导其掺入 确定ECM蛋白/SNED 1相互作用在SNED 1 ECM组装中的作用（Aim 2）; 并鉴定在细胞表面控制SNED 1依赖性ECM组织的SNED 1受体， 负责SNED 1的粘合性能（目标3）？ 我们的目标是填补我们对ECM组装基本机制的理解中的关键空白， 关于SNED 1这是一个必要的步骤，以破译如何扰动这些机制， 可能导致发育缺陷和癌症进展。这项工作也将铺平道路的发展 未来的治疗策略，以纠正导致疾病的细胞外基质结构改变。