Abstract: Localization of biochemical signaling internal to cells is critical for cellular function, affecting a variety of processes of importance in human health, including cell division, differentiation, motility, and inter- and intracellular signaling. As such, biologists are keenly interested in studying and probing the role of biomolecule localization, and have developed molecular biological tools in order to selectively perturb local environments internal to cells. Although these tools, including gene knockouts, gene fusions, and photoactivated ligands, have allowed some investigation into localization effects, a simple and robust approach that is broadly applicable to precisely localize a range of biomolecules intracellularly is critically lacking. Magnetic nanoparticles can be conjugated to a range of biomolecules of interest and are readily uptaken by most cells, but intracellular manipulation and localization has been limited by the strong field gradients required to create appreciable forces on such small particles. This limitation has been overcome by using magnetic-gradient enhancing substrates (""""""""nano-active slides"""""""") that elicit modal behavior. Using this approach allows for the robust transformation of macroscale movements of an external magnet to nanoscale movements of intracellular environment-modifying nanoparticle ensembles. Dynamically localized nanoparticles can then act to transduce a variety of chemical, mechanical, and thermal signals with unprecedented control. Achieving the full potential of nanoparticle-based intracellular engineering, including joystick-based dynamic control of the intracellular nanoenvironment and precision perturbation of intracellular locations in massively parallel arrays will require additional investigation and development. Thrust areas will include nanoparticle conjugation and delivery, transparent substrate development, cell patterning and alignment to magnetic elements, and development of joystick-based control systems for nanoparticle movement. Although the most significant impact will be in creating a general platform for biological experimentation, concurrent with developing tools for precision engineering of intracellular environments we will also use these tools for initial pioneering studies of the effect of localization in cellular motility and Ca2+ propagation internal to cells. These experiments will potentially culminate in the ability to remotely control cell movement and extract quantitative understanding of key factors required for cell migration through engineering its outcome. Public Health Relevance: By developing simple tools for precise control of nanoparticles locally within cells it will be possible to understand and control aspects of cell behavior that could not be probed previously. Similar to how we investigate large-scale physiology by perturbing local organ systems, these type of tools will allow a more mathematical understanding of cell behavior through perturbation of sub-cellular locations and organelles in time and space. Ultimately, this type of tool will be broadly used by biologists to uncover unique aspects of cellular function that directly impact human health.
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