The orchestration of cell behavior into multicellular pattern formation is a central issue for developmental, regenerative, cancer, evolutionary, and synthetic biology. Molecular and cell biology has made remarkable strides because tools were developed for the experimental control of biochemical signals. However, Bioelectric signals - a fascinating and important physical layer of pattern control - remain poorly understood. Functional experiments using targeted, quantitative, molecular-level changes in ion fluxes in a variety of vertebrate and invertebrate model systems implicated specific bioelectric events in development (left-right asymmetry, craniofacial patterning, eye induction), regeneration (tail and limb regeneration in vertebrates, and anterior-posterior polarity in planaria), and is now being applied to detection and suppression of cancer in situ. By developing 1) tools that allow molecular-level changes in physiological properties in vivo, 2) building quantitative computer models synthesizing physiological and molecular data, 3) fleshing out pathways that show in detail how biophysical signals are produced, propagated, and transduced into downstream canonical biochemical/transcriptional responses, and 4) disseminating protocols and reagents to many labs in related fields, bioelectricity has been brought into a new age. Crucially, the field is held back by a lack of tools: transformative impact requires the ability for many labs to be able to exert tight spatio-temporal control over ion flux and transmembrane voltage in vivo.
Optogenetics (expression of light-sensitive ion channels) is an exciting advance, but has never been applied outside of excitable cells (nerve and muscle) because available devices do not allow flexible control of sufficiently-bright light delivered over large areas - a necessity when working with developing or regenerating systems (organs, whole animals, or bioengineered constructs). This project will develop an automated, highly versatile, optogenetics research station that extends past existing technology to enable experimental control of voltage gradients in model systems. Built around a computer controlled microscope that is suitable for working with anything from small animals to individual cells, this platform will allow any lab that has access to molecular biology and microscopy to perform screens and targeted experiments on the role of physiology and bioelectricity in any context. This will significantly impact several basic fields by transforming the state of the art in how functional in vivo physiology experiments are planned, executed, and analyzed. Aim 1 capitalizes on commercial partnerships and local collaborators in engineering and optics to build a platform for light-based control of resting potential in any desired cell groups. Aim 2 validates the system by proof-of-principle applications in the control of stem cell derivatives in vivo and regulation of organ patterning in the Xenopus laevis system. Crucially, protein/mRNA profile does not fully determine cell behavior; this IDBR platform will transform the field of functional electrophysiology, and help to crack the bioelectrical code, by, for the first time, allowing many labs to easily collect functional physiomic data. Direct control of voltage in any cell/tissue of interest will enable quantitative understanding of how biophysical (post-translational) parameters interact with gene regulatory networks in determining pattern formation and tissue/organ function. Bringing a whole new aspect of regulation to several communities will allow the field to truly understand the role and information content of biophysical gradients.
The proposed activity will involve the training of a minority PhD student and talented young post-doctoral fellow in highly interdisciplinary techniques at the forefront of developmental biology, at the collaborative edge between basic science and engineering industry. Natural bioelectric fields form a kind of subtle "scaffold" that determines the growth and form of biological structures, and guides the activity of gene networks. The ability for any lab to control the natural bioelectric fields inside living tissues will transform our ability to understand and control the shape of tissues and organs. This will have important implications for understanding evolution of complex body parts, as well as ultimately driving novel the synthetic biology and bioengineering applications. The direct beneficiaries of this technology will be not only the undergraduates and other students who will use such devices in state-of-the-art learning modules, but society as a whole, which will ultimately benefit from applications of this technology to biomedicine (bioengineering of organs in vitro, regenerative sleeves, and development of robust computational devices made of living tissues).
