Photo-inducible Cell Ablation and Fluorescent Chemosensors for Chemical Biology

用于化学生物学的光诱导细胞消融和荧光化学传感器

• 批准号: RGPIN-2017-06488
• 负责人: Beharry, Andrew
• 金额: $ 1.97万
• 依托单位: University of Toronto
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2022
• 资助国家: 加拿大
• 起止时间: 2022-01-01 至 2023-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=746965
• 关键词: Photo; inducible; Cell; Ablation; Fluorescent

We are interested in developing chemical tools to probe biological complexity in vivo. Specifically, we are proposing two areas of research:1) Photo-induced Cell Ablation. Deciphering the role of entire cells during biological events has been a difficult challenge since cells function in complex networks. A powerful approach to probing their complexity is to turn “off” cells then monitor developmental/behavioral changes in the organism. To achieve this effectively, a promising strategy is to use small molecule photosensitizers that can produce reactive oxygen species in the presence of light, which will kill or turn “off” cells. However, once administered, photosensitizers accumulate in all tissues making it difficult to probe cellular function at a specific location. To address this issue, we are proposing to develop a series of photosensitizers that will require not only light but also an enzyme for its activation. With this chemical genetic approach, cells can be destroyed efficiently (low light dosage) at a specific site with minimal non-specific cell destruction of the surrounding region. Overall this strategy will enable us to accurately assign functions to cells and tissues.2) Fluorescent Chemosensors. Monitoring enzymes in their native cellular environments would be a powerful approach to truly understand their function. Tools that are capable of directly reporting on their behavior could provide an accurate map of activity in both normal and diseased states. To this regard, we are proposing to construct a series of fluorescent chemosensors for the enzyme class, methyltransferases. Our sensors will be general and selective to permit real-time sensing of all methyltransferase activity in live cells, and will be used to answer specific questions regarding their native roles that would otherwise be difficult to obtain by conventional biochemical methods. In turn, these novel tools will permit a better understanding of the network of methyltransferases in epigenetic pathways.

我们对开发化学工具来探索体内生物复杂性感兴趣。具体地说，我们提出了两个研究领域：1)光诱导细胞消融。破译整个细胞在生物事件中的作用一直是一个困难的挑战，因为细胞在复杂的网络中发挥作用。探索其复杂性的一个有效方法是关闭细胞，然后监测有机体的发育/行为变化。为了有效地实现这一点，一个有希望的策略是使用小分子光敏剂，这种光敏剂可以在光存在的情况下产生活性氧物种，从而杀死或关闭细胞。然而，一旦给予光敏剂，光敏剂就会在所有组织中积累，因此很难探测特定位置的细胞功能。为了解决这个问题，我们建议开发一系列光敏剂，不仅需要光，而且需要一种酶来激活它。使用这种化学遗传方法，细胞可以在特定的位置被有效地破坏(低光剂量)，而对周围区域的非特定细胞的破坏最小。总体而言，这一策略将使我们能够准确地为细胞和组织分配功能。2)荧光化学传感器。监测它们天然细胞环境中的酶将是真正了解它们功能的有力途径。能够直接报告它们的行为的工具可以提供正常和患病状态下的准确活动地图。在这方面，我们建议构建一系列针对甲基转移酶这一酶类的荧光化学传感器。我们的传感器将是通用的和选择性的，以允许实时检测活细胞中的所有甲基转移酶活性，并将用于回答有关其天然角色的特定问题，否则将很难通过传统的生化方法获得这些问题。反过来，这些新的工具将允许更好地理解表观遗传途径中的甲基转移酶网络。