Genetically encoded tools to control any mammalian cell function with any desired stimulus

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The development of genetically encoded tools like channelrhodopsin-2 has allowed the stimuli of light to control membrane depolarization of neurons, sparking a steady stream of breakthroughs in our understanding of brain function. While light as a stimuli has strengths (e.g. precise spatial and temporal activation), it also has weaknesses (e.g. low tissue penetration and invasive animal surgery). Furthermore, other than membrane depolarization as a cell function, biologists have sought control over cell death to determine the biological role of the particular cell types and control over gene expression to determine the function of particular genes under regulation. ******My ultimate research objective is to engineer genetically encoded tools that will allow the control of any mammalian cell function with any desired stimuli. Our group believes this is possible through Ca2+ rewiring combining proteins that generate Ca2+ signals upon binding desired stimuli with proteins that control cell function when activated by Ca2+ signals.******First in the proposal, we will develop genetically-encoded tools to allow Ca2+ influx and membrane depolarization to be controlled by magnetic fields. Magnetic fields are permeable through tissues that will allow neurons to be stimulated deep inside the brain with no surgery in contrast with light-based or electrode-based approaches. Second, we will develop Ca2+-activated caspases. Caspases are central cysteine proteases in the apoptotic cell death that function to dismantle the cell machinery by cleaving after an aspartate residue in their substrate. By co-expressing the chimeric caspase with a chimeric receptor that generates a Ca2+ signal in response to binding its target ligand, we aim to induce cell death in response to any stimuli via Ca2+ rewiring. Lastly, we will develop inducible gene expression systems. Through the ubiquitous NFAT pathway, Ca2+ oscillations activate gene expression. By co-expressing a chimeric receptor with natural or synthetic genes that enhance Ca2+ oscillations, we aim to induce gene expression in response to any stimuli via Ca2+ rewiring.******The ability to control any mammalian cell function (e.g. Ca2+ influx, cell death and gene expression) with any desired stimuli (e.g. magnetic fields) would allow us to ask biological questions that are currently not possible. For example, magnetic field stimuli would allow activation of the areas deep inside the brain such as the hippocampus to probe questions about the formation of memories. Cell ablation rewired to any stimuli would allow cell death to be customized to the microenvironment (e.g. patterns of morphogens) to probe questions about their importance in development of tissues. Gene expression rewired to any stimuli would expand the current paucity of reliable inducible promoters for biomedical research. If rewired to magnetic fields, we could perform targeted deep tissue ablation and gene expression in the brain and spinal cord.