Intracellular Biophotonic Nanoswitches

最近对特定细胞中蛋白质之间所有物理相互作用的绘图证实了蛋白质之间的相互作用是高度调节的，是所有细胞过程的基础。研究人员和技术人员面临着一个重大挑战——如何对这样复杂的系统提出具体的问题，特别是当给定细胞中蛋白质相互作用随时间变化并导致不同的最终状态时。例如，当人类细胞对压力作出反应时，主调节蛋白之间的特定相互作用开始驱动恢复过程或启动对细胞死亡的受控承诺。这个项目旨在产生一种解决这个问题的通用技术——将合成开关引入活细胞，通过远程控制“微调”蛋白质的相互作用。这种令人兴奋的方法，基于非常有前途的初步工作，将允许研究人员通过引入小干扰分子，通过精心选择的波长的光来开启和关闭这些小干扰分子，从而对已定义的蛋白质-蛋白质相互作用进行编程。外部光脉冲诱导肽主链的构象重排，从而改变细胞内生物光子纳米开关（IBN）的生物学特性，从而触发小分子结构的变化。ibn是一种基于光敏纳米粒子的分子结构，它与短肽序列相连，可以识别蛋白质表面的特征，这些特征已经被靶向转换。传统的和新颖的IBN传递到活细胞的方法将允许转换成细胞群的模式。通过光操作这些ibn将使研究人员能够在如此复杂的细胞群中对开关的激活进行建模，或者对单个细胞中的开关过程进行“编程”——这是在操纵离散细胞内通路的主调节器技术上迈出的一步。我们的提案的冒险和风险与IBN设计的问题和它们在活细胞中自我报告的潜力有关。ibn将允许研究人员通过生物物理手段切换或编程活细胞中主调节器的状态，并探索整个系统的后果，以揭示不同途径的内部联系。此外，我们的建议解决了如何跟踪选择开关的下游后果，即使在不同的谱系中，揭示细胞如何对不同的信号（幅度或频率）做出反应，即使这些信号在不同的细胞世代中迅速出现或确实缓慢发展。由于我们的愿景是为生命科学界提供基于强大化学系统的新颖，强大且易于实施的技术，因此该提案包括与IBN技术的潜在用户和下游需求的接触，重点是在复杂的“系统”层面上理解细胞生物学的新兴需求。令人兴奋的前景是获得可编程光子控制正常生理（指导干细胞分化，操纵伤口愈合和延缓细胞衰老），肿瘤（细胞周期检查点功能障碍和光控制治疗的癌症生物学），构建细胞群落（光定向组织工程）和分子靶标识别（寻找新的药物和产品）。

Recent mapping of all physical interactions between proteins in a given cell has confirmed the notion that interactions between proteins are highly regulated and underpin all cellular processes. Researchers and technologists have been presented with a major challenge - how to ask specific questions of such complex systems especially when protein interactions change with time in a given cell and result in different end states. For example when a human cell responds to stress, specific interactions between master regulatory proteins start to drive a recovery process or initiate a controlled commitment to cell death. This project aims to generate a generic technology for solving this problem - introducing synthetic switches into live cells that can 'fine-tune' protein interactions by remote control. This exciting approach, based on highly promising preliminary work, would allow the investigator to programme changes in defined protein-protein interactions by the introduction of small interfering molecules engineered to be switched on and off by light of carefully selected wavelengths. Changes in the structure of a small molecule are triggered by external light pulses inducing conformational rearrangements in the peptide backbone and hence alterations of the biological properties of the Intracellular Biophotonic Nanoswitch (IBN). IBNs are light-sensitive nanoparticle-based molecular structures linked to the short peptide sequences that recognize features on the surface of a protein that has been targeted for switching. Conventional and novel methods for IBN delivery into live cells will allow patterning of the swiches into populations of cells. Operating these IBNs by light will allow the researcher to pattern the activation of switches in such complex cell populations or to 'programme' the switching process in single cells - a step-forward in the technology of manipulating master regulators of discrete intracellular pathways. Our proposal's adventure and risk relates to the problems of IBN design and their potential for self-reporting in live cells.IBNs will allow a researcher to switch or programme the state of a master regulator in a live cell by biophysical means and explore the consequences on the whole system to reveal the internal linking of different pathways. Furthermore, our proposal addresses how to track the downstream consequences of selective switching, even in different lineages, to reveal how cells respond to different signals (amplitude or frequency) in developing their responses even if these arise quickly or indeed develop slowly through different cell generations.Since our vision is to provide the life sciences community with novel, robust and readily implemented technologies based on robust chemical systems, the proposal encompasses engagement with potential user & downstream demands of IBN technology with a focus on the burgeoning demand to understand cellular biology at the complex 'systems' level. The exciting prospect looms of gaining programmable photonic control over normal physiology (directing stem cell differentiation, manipulating wound healing and delaying cell senescence), neoplasia (cancer biology of cell cycle checkpoint dysfunction and photonically-controlled therapeutics), constructed cell communities (light-directed tissue engineering) and molecular target identification (the search for new medicines and products).