Sensing and controlling cell signaling events with bioluminescent kinase sensors and gene circuits - Cell signaling events like growth factors and cell cycle play crucial roles in regulating the behavior of cells. Growth factors are signaling molecules that promote cell growth, division, and survival. They bind to cell-surface receptors that are linked to kinases, which initiate intracellular signaling pathways to exert a cellular response. On the other hand, the cell cycle represents a completely different class of cell signaling events where a complex network of proteins interacts with each other to ensure that the cell divides only when it is ready and that the daughter cells are identical to the parent cell. More recently, cell cycle regulation or dysregulation has been shown to be involved in disease processes that are not related to cell division. The cell cycle is regulated by both positive and negative cues, such as growth factors and DNA damage, respectively. In both cases, these cell signaling processes are highly relevant in both health and disease and the roles of these signaling events are still being discovered. We aim to develop a series of bioluminescent kinase sensors designed to monitor various cell signaling processes. We are starting with two examples to apply this concept, (1) Detecting and manipulating cell cycle state and progression and (2) detecting and manipulating growth factor signaling. These sensors will provide real-time, non-invasive insights into cellular processes, enhancing our understanding of cell biology and disease progression. We also aim to apply these light emitting sensors to control optogenetic actuators to amplify or inhibit these cell signaling events through light activated transcription that will occur in response to both the cell signaling events of interest and presence of a luciferin that can be added to cell culture or injected in model organisms occurring simultaneously. The proposed research represents a significant advancement in the field of cell biology, creating novel tools for manipulation of cellular function to study cell signaling processes in health and disease. The successful development of these sensors will not only enhance our understanding of cell signaling but also open new avenues for therapeutic intervention either through mechanistic understanding of cellular processes or by adapting this system to correct aberrant cell signaling events. The use of bioluminescent based sensors paired with optogenetic actuators offers several advantages over current approaches to probing cell signaling events in that manipulations are dependent on a specific kinase’s activity and the presence of luciferin, brightness can be tuned based on luciferin concentration and the on/off kinetics of the sensor can be tuned. Long-term, development of these systems will enable a variety of new basic science questions to be answered by allowing kinase dependent cell signaling events in various cell types to be manipulated in vitro and in vivo. The goal over the next five years is to successfully develop two distinct cell signaling sensing and control systems that can be used for basic science applications as proof of principle systems that we will apply in our lab and collaborators labs with the goal of expanding to other cell signaling categories.
