A genetically-encoded sensor for imaging intracellular fatty acids

描述(申请人提供)：脂类的储存和动员是基本的细胞过程，在后生动物进化过程中是保守的，几乎存在于所有哺乳动物细胞中。细胞内游离脂肪酸(FFAi)通常受到严格控制，因为过量的FFAI是有毒的。事实上，细胞内FFA和FFA代谢物的过度积累会导致细胞功能障碍，被称为“脂毒性”，这被认为是肥胖导致糖尿病和心血管疾病的主要途径。因此，从机制上理解细胞如何同化、动员和引导游离脂肪酸是一个重要的生物学问题，对健康和疾病具有广泛的影响。细胞内FFA的摄取和动员似乎具有重要的时间和空间域。然而，由于目前还不能对细胞内的FFA水平进行成像，所以对活细胞内FFA代谢发生的时间和地点的研究是不可能的。因此，R21应用的目标是开发和实现一种用于生物应用的具有高空间和时间分辨率的FFAI光学传感器。该传感器可用作一般的胞浆传感器，也可专门针对建议的亚细胞内FFA摄取、动员和氧化部位。我们预计，经过验证的传感器将有广泛的应用，并将导致对细胞脂肪运输生物学的新见解。 与公共卫生相关：储存和动员脂质是几乎所有哺乳动物细胞中都存在的基本细胞过程。细胞内游离脂肪酸(FFAi)通常受到严格控制，因为过量的FFAI是有毒的，可能导致肥胖相关的糖尿病和心血管疾病。因此，从机制上理解细胞如何同化、动员和引导游离脂肪酸是一个重要的生物学问题，对健康和疾病具有广泛的影响。有证据表明，细胞内FFA的摄取和动员具有重要的时间和空间域，然而，这些动力学的分析并不困难，因为目前还不能对细胞内FFA水平进行成像。因此，R21应用的目标是开发和实现一种用于生物应用的具有高空间和时间分辨率的细胞内FFA光学传感器。该传感器可以作为一般的胞浆传感器使用，也可以专门针对FFA摄取、动员和氧化的关键亚细胞部位。我们预计，经过验证的传感器将有广泛的应用，并将导致对细胞脂肪运输生物学的新见解。

DESCRIPTION (provided by applicant): The storage and mobilization of lipid are fundamental cellular processes that are conserved throughout metazoan evolution and are present in virtually all mammalian cells. Intracellular FFA (FFAi) are normally kept under tight control since excessive FFAi are toxic. Indeed, excessive accumulation of intracellular FFA and FFA metabolites leads to cellular dysfunction, termed 'lipotoxicity', which is thought to be a major means by which obesity contributes to diabetes and cardiovascular disease. Thus, a mechanistic understanding of how cells assimilate, mobilize and channel FFA is an important biological question with broad implications for health and disease. The uptake and mobilization of FFA within cells appear to have important temporal and spatial domains. Nonetheless, investigation of the "when and where" FFA metabolism occurs within live cells has not been possible because intracellular FFA levels cannot presently be imaged. Therefore, the aim of the R21 application is to develop and implement an optical sensor of FFAi having high spatial and temporal resolution for biological applications. This sensor is designed to be used as a general cytosolic sensor, or can be specifically targeted to proposed subcellular sites of FFA uptake, mobilization and oxidation. We anticipate that a validated sensor will have wide applications and will lead to new insights into biology of cellular lipid trafficking. PUBLIC HEALTH RELEVANCE: The storage and mobilization of lipid are fundamental cellular processes that are present in virtually all mammalian cells. Intracellular FFA (FFAi) are normally kept under tight control and since excessive FFAi are toxic and can contribute to obesity-related diabetes and cardiovascular disease. Thus, a mechanistic understanding of how cells assimilate, mobilize and channel FFA is an important biological question with broad implications for health and disease. Evidence indicates that the uptake and mobilization of FFA within cells has important temporal and spatial domains, however, analysis of these dynamics has not been difficult because intracellular FFA levels cannot presently be imaged. Therefore, the aim of the R21 application is to develop and implement an optical sensor of intracellular FFA with high spatial and temporal resolution for biological applications. This sensor can be used as a general cytosolic sensor, or can be specifically targeted to key subcellular sites of FFA uptake, mobilization and oxidation. We anticipate that a validated sensor will have wide applications and will lead to new insights into biology of cellular lipid trafficking.