Billions of cells in the colon are replaced daily due to natural turnover and repair. During this process, tissue stem cells are activated from renewal/dormancy. This activation, known as stem cell commitment, involves exiting their local niches, leading to increased proliferation and progression to differentiation. How do dying or damaged cells signal to the stem cell compartment, hundreds of microns away, to initiate replacement? The balance between stem cell renewal and commitment depends on mechanical and growth factor signals. These signals interact, driving stem cells to divide symmetrically or asymmetrically, a key factor in maintaining the stem cell pool and producing specialized cells for tissue repair. The long-term goal of this research is to control and understand these processes to advance regenerative medicine and develop tissue repair therapies.

Despite the known significance of signaling dynamics in stem cell commitment, the balance between mechanical and growth factor signaling in the colon remains under-explored. Comprehensive Temporal Analysis and Control (CTAC) is an innovative system we developed, combining biology, physics, and optics, to investigate signaling dynamics in each cell of organoids. CTAC uses patient-derived colonoids, high-content live-cell imaging, innovative signaling reporters, and optogenetic tools to analyze the interaction of mechanical and growth factor signals. Our preliminary studies using CTAC in a high-content screen using over 700 kinase inhibitors in live organoids identified mechanically regulated kinases as top regulators of stem cell commitment, highlighting the importance of mechanical signals in addition to growth factor pathways. Following up on these hits revealed that maintaining contractility is essential for both maintaining stem cells and preserving the wound healing function of differentiated cells.