Dissecting subcellular and cellular organization by spatial molecular neighborhood networks

通过空间分子邻域网络剖析亚细胞和细胞组织

• 批准号: 10713565
• 负责人: Ahmet F. Coskun
• 金额: $ 37.21万
• 依托单位: GEORGIA INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-22 至 2028-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10713565
• 关键词: 3-Dimensional; Affect; Automobile Driving; Benchmarking; Bioinformatics; Biological Assay; Biology; Biomanufacturing; Cell Survival; Cell model; Cell physiology; Cells; Collaborations; Crowding; Data Analyses; Data Set; Disease; Environment; Epithelial Cells; Event; Fluorescent in Situ Hybridization; Functional disorder; Gene Expression; Genes; Genetic; Genomics; Goals; Growth; Health; Human; Image; Imaging technology; Individual; Intervention; Ligation; Light; Mathematics; Measurement; Mesenchymal; Modeling; Molecular; Molecular Evolution; Neighborhoods; Normal Cell; Normal tissue morphology; Outcome; Pathology; Pharmaceutical Preparations; Phenotype; Population; Proliferating; Protein Analysis; Proteins; Public Health; RNA; Regulation; Research; Resolution; Role; Signal Transduction; Signaling Protein; Specificity; Specimen; Stimulus; System; Technology; Tissues; Transcript; Visualization; Work; biological systems; experience; high throughput screening; molecular imaging; molecular modeling; molecular scale; response; small molecule; superresolution microscopy; tissue/cell culture; treatment response

Subcellular organization of intracellular molecules regulates basic cell functions such as growth, division, proliferation, and differentiation. By regulating subcellular neighborhoods of RNAs and proteins, cells survive but also specialize in a multicellular system. While subcellular localization mechanisms are widely studied, the role of subcellular neighborhoods in determining the cellular response to stimuli and multiple drugs has been limited to a few low-throughput and low-sensitivity molecular studies. Little is known about how these highly coordinated subcellular neighborhoods give rise to tissue-specificity, tissue-sate, and how they regulate cell function in distinct subcellular volumes of tissue states and phenotypes. Image-based visualization of molecular neighborhoods has explored only a few target molecules, yielding evidence of the importance of subcellular localization of transcripts and protein factors. One bottleneck in subcellular regulation is the lack of models explaining neighborhood interactions of molecules that are driving cellular response in tissues from distinct abnormalities. Previous work established multiplexed gene expression measurements to model subcellular RNA neighborhoods and multiparameter protein analysis to quantify spatial signaling protein neighborhoods that are altered in response to perturbations. However, dominant signaling protein and RNA neighborhoods have not been established at a physically meaningful resolution in widely employed mesenchymal and epithelial cells as they respond to perturbations, experience differentiation events, or adapt to tissue environments of individuals treated with cell-based or small molecule interventions for correcting abnormalities due to genetic or molecular dysfunction. Thus, the long-term goal of this project is to leverage advanced high-throughput screening and spatially resolved genomic and protein measurements in single cells for modeling molecular neighborhoods in populations and tissue contexts. To achieve these tasks, using recent advances in sequential fluorescence in situ hybridization and multiplexed protein imaging, this proposal plans to 1) mechanistically dissect the molecular RNA and protein neighborhoods using proximity ligation assays, 2) enhance the spatial information capacity of molecular neighborhoods using 3D super-resolution microscopy and temporal evolution of molecular neighborhoods by pseudo-temporal differentiation models, 3) evaluate the outcome of subcellular neighborhood organization and tissue context of mesenchymal cells near vessels and epithelial cells near linings from human specimens isolated from health restoring interventions. Molecular benchmarking and normal tissue annotations will be performed in collaboration with signaling biology, bioinformatics, biomanufacturing, and pathology experts. Cross-scale molecular control from subcellular neighborhoods to tissue organization will reveal how a systemic response to treatments can be re-stratified by network variance of molecular neighborhoods. This MIRA proposal sheds light on the spatially resolved subcellular organization in health and disease, providing a predictive metric for deciphering tissue-state control and alterations in many disorders.

细胞内分子的亚细胞组织调节基本细胞功能，例如生长，分裂，增殖和分化。通过调节RNA和蛋白质的亚细胞邻域，细胞可以存活，但也专门研究多细胞系统。尽管广泛研究了亚细胞定位机制，但亚细胞邻居在确定刺激和多种药物的细胞反应中的作用已限于一些低通量和低敏感性分子研究。对于这些高度协调的亚细胞邻域如何产生组织特异性，组织酸盐以及它们如何调节细胞功能在组织态和表型的不同亚细胞体积中的细胞功能。基于图像的分子邻域的可视化仅探索了几个靶分子，从而证明了转录本和蛋白质因子亚细胞定位的重要性。亚细胞调节中的一个瓶颈是缺乏解释分子的邻域相互作用的模型，这些模型正在驱动组织中细胞反应的不同异常。先前的工作建立了多路复用基因表达测量值，以模拟亚细胞RNA邻域和多参数蛋白分析，以量化对扰动而改变的空间信号蛋白邻域。然而，在广泛使用的间质和上皮细胞中，尚未建立出主要的信号蛋白和RNA社区，因为它们对扰动，经历了分化事件或适应细胞基于基于细胞或小分子干预的个体的组织环境，以纠正因遗传或分子的遗传性或分子而受到的小分子干预措施的组织环境。因此，该项目的长期目标是利用单个细胞中的先进的高通量筛选和空间分辨的基因组和蛋白质测量，以模拟种群和组织环境中的分子邻居。为了实现这些任务，使用序列荧光原位杂交和多路复用蛋白成像的最新进展，该建议计划使用接近性连接测定法进行机械机械地解剖分子RNA和蛋白质邻域，2）使用3D超级分辨率显微镜和临时模型，以pse的模型来增强分子邻域的空间信息能力，评估从健康恢复干预措施中分离出的人类标本附近血管和上皮细胞附近的间充质细胞和上皮细胞附近的间充质细胞和上皮细胞附近的间充质细胞的结局。分子基准和正常组织注释将与信号生物学，生物信息学，生物制造和病理专家合作进行。从亚细胞邻域到组织组织的跨尺度分子控制将揭示如何通过分子邻域网络方差重新分解对治疗的全身反应。该MIRA提案阐明了健康和疾病中的空间解决亚细胞组织，为解密的组织状态控制和许多疾病的改变提供了预测指标。