Cell density controls signal propagation waves in a multicellular synthetic gene circuit

Abstract: During organismal development, biochemical reaction networks sense and respond to mechanical forces to coordinate embryonic patterning with embryo morphogenesis. Factors such as cortical tension, cell density, and matrix mechanical properties influence differentiation and cell fate decisions by modulating gene regulatory signaling networks. A major goal in synthetic development is to construct gene regulatory circuits that program the patterning and morphogenesis of synthetic multicellular structures. However, in the synthetic context, little is known regarding how the physical properties of the growth environment impact the behavior of synthetic gene circuits. Here, we exploit physical-chemical coupling observed in a synthetic patterning circuit in order to control the size and spatial distribution of patterned synthetic cell sheets. We show that cell density attenuates the propagation of signal between neighboring cells in a multicellular sheet containing a contact-dependent patterning circuit based on the synNotch signaling system. Density-dependent attenuation leads to a signal propagation wave that exhibits distinct qualitative phases of persistent propagation, transient propagation, and no propagation. Through computational modeling, we demonstrate that cell growth parameters determine the phase of propagation observed within a growing cell sheet. Using growth-modulating drugs and spatial density gradients, we control the size of synNotch-activated cell populations and generate tissue-scale activation gradients and kinematic waves. Our study reveals that density-dependent synNotch activity can be exploited to control a synthetic multicellular patterning circuit. More broadly, we show that synthetic gene circuits can be critically impacted by their physical context, providing an alternate means for programming circuit behavior.