Creating Smart Biomaterials using Engineered Bacteria that Cooperatively Reprogram Mammalian Cells

非技术性摘要重新编程哺乳动物细胞功能的能力对于探索细胞行为的科学基础以及改善人类健康至关重要。这一努力将导致智能生物材料和实验系统的开发，这些材料和实验系统由携带工程细菌的微粒组成。这些混合的、有生命的、无生命的生物材料将被用作重新编程哺乳动物细胞信号的工具。一种可扩展的合成生物材料方法的一个优点是，使用工程菌来调节哺乳动物的钙信号可能会消除直接改变哺乳动物细胞以重新编程它们的行为的需要。因此，可以减少对病毒等载体或创建转基因动物模型的需求。为了这一努力，将开发与细菌结合的微粒，这种微粒可以将合成的遗传成分输送到哺乳动物细胞，并重新编程钙信号。钙信号是一种无处不在的系统，它影响着从神经到肠道细胞的各种细胞，并控制着缓慢和快速的细胞过程。下一步，这些粒子将被制成？智能？生物材料是它们的生命成分吗？细菌？将被设计成能够合作并共同决定何时将基因成分传递给哺乳动物细胞。最后，将使用数学模型和计算模拟来探索哺乳动物细胞中的钙信号动力学，以确定何时改变将导致细胞信号动力学中最显著的变化。然后，智能生物材料将被用来重新编程这一信号。此外，这项工作将与生物工程夏令营的多个教学模块的开发相结合，最终课程将广泛传播给更广泛的科学和教育社区。技术摘要这一努力将导致创建智能生物材料，这些材料连接到细菌协同入侵所需的工程细胞内网络，然后对哺乳动物细胞中的钙信号进行重新编程。首先，将建立一个合成组件，使工程细菌能够共价结合到微粒上，以创建针对特定哺乳动物细胞表面标记的智能生物传感材料。其次，不同的细菌株将被编程为具有工程基因电路，使它们能够相互合作识别对方，然后激活哺乳动物细胞入侵能力。这种合作进一步确保了只有特定的哺乳动物细胞才是目标。最后，人工结合和协同侵入回路都将被用来重新编程哺乳动物细胞中的钙信号。入侵的细菌将使用RNA干扰，通过下调钙途径酶的表达来改变钙信号的动态。这些目标酶的选择将通过单细胞钙动力学的建模来提供信息。这一努力将结合工程化细胞入侵、哺乳动物合成生物学、细胞表面展示和工程化群体感应方面的最新进展，以创造携带细菌的智能生物材料，对哺乳动物细胞进行重新编程。拟议的努力包括促进智能生物材料领域的发现和理解的研究，同时还通过开发一项针对高中生的新活动来促进教学、培训和学习。

NON-TECHNICAL ABSTRACTAn ability to reprogram the function of mammalian cells is critical for exploring the scientific underpinnings of cell behavior as well as improving human health. This effort will result in the development of smart biomaterials and experimental systems made up of microparticles carrying engineered bacteria. These hybrid, living-nonliving biomaterials will be used as a tool to reprogram mammalian cell signaling. One advantage of an extendable synthetic biomaterials approach that uses engineered bacteria to regulate mammalian calcium signaling could be the abrogation of the need to directly alter mammalian cells to reprogram their behavior. Thus, the need for vectors such as viruses or the creation of transgenic animal models could be reduced. For this effort, bacteria-bound microparticles will be developed that can deliver synthetic genetic components to mammalian cells and reprogram calcium signaling. Calcium signaling is a ubiquitous system that effects cells ranging from nerves to the cells that line the gut, and controls both slow and fast cellular processes. Next, these particles will be made into ?smart? biomaterials as their living component ? the bacteria ? will be engineered with an ability to collaborate and collectively determine when to transmit genetic components to mammalian cells. Finally, mathematical modeling and computational simulation will be used to explore calcium signaling dynamics in mammalian cells in order to determine when alterations will cause the most significant changes in cell signaling dynamics. The smart biomaterials will then be used to reprogram this signaling. In addition, this effort will be integrated with the development of multiple teaching modules for a bioengineering summer camp, and the final curriculum will be widely disseminated to the broader scientific and educational community.TECHNICAL ABSTRACTThis effort will result in the creation of smart biomaterials that link to engineered intracellular networks necessary for bacteria to cooperatively invade and then reprogram calcium signaling in mammalian cells. First, a synthetic component will be built to enable engineered bacteria to covalently bond to microparticles to create smart biosensing materials that target specific mammalian cell surface markers. Second, different bacterial lines will be programmed with engineered gene circuits that enable them to cooperatively recognize each other and then activate mammalian cell invasion capabilities. This cooperation further ensures that only specific mammalian cells are targeted. Finally, both the synthetic bonding and the cooperative invasion circuitry will be used to reprogram calcium signaling in mammalian cells. Invading bacteria will use RNA interference to alter the dynamics of calcium signaling by knocking down expression of calcium pathway enzymes. Selection of these target enzymes will be informed by modeling of single cell calcium dynamics. This effort will combine recent advances in engineered cellular invasion, mammalian synthetic biology, cell surface display, and engineered quorum sensing to create bacteria-laden, smart biomaterials that reprogram mammalian cells. The proposed effort includes research to advance discovery and understanding in the field of smart biomaterials while also promoting teaching, training, and learning through the development of a new activities targeted toward high school students.