A common theme in the self-organization of multicellular tissues is the use of cell-cell signaling networks to induce morphological changes. We used the modular synNotch juxtacrine signaling platform to engineer artificial genetic programs in which specific cell-cell contacts induced changes in cadherin cell adhesion. Despite their simplicity, these minimal intercellular programs were sufficient to yield assemblies with hallmarks of natural developmental systems: robust self-organization into multidomain structures, well-choreographed sequential assembly, cell type divergence, symmetry breaking, and the capacity for regeneration upon injury. The ability of these networks to drive complex structure formation illustrates the power of interlinking cell signaling with cell sorting: Signal-induced spatial reorganization alters the local signals received by each cell, resulting in iterative cycles of cell fate branching. These results provide insights into the evolution of multicellularity and demonstrate the potential to engineer customized self-organizing tissues or materials.

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Apoptotic cells expose phosphatidylserine on their surface as an "eat me" signal, and macrophages respond by engulfing them. Although several molecules that specifically bind phosphatidylserine have been identified, the molecular mechanism that triggers engulfment remains elusive. Here, using a mouse pro-B cell line, Ba/F3, that grows in suspension, we reconstituted the engulfment of apoptotic cells. The parental Ba/F3 cells did not engulf apoptotic cells. Ba/F3 transformants expressing T cell immunoglobulin-and mucin-domain-containing molecule 4 (Tim4), a type I membrane protein that specifically binds phosphatidylserine, efficiently bound apoptotic cells in a phosphatidylserine-dependent manner but did not engulf them. However, Ba/F3 transformants expressing both Tim4 and the integrin ␣ v ␤ 3 complex bound to and engulfed apoptotic cells in the presence of milk fat globule epidermal growth factor factor VIII (MFG-E8), a secreted protein that can bind phosphatidylserine and integrin ␣ v ␤ 3 . These results indicate that the engulfment of apoptotic cells proceeds in two steps: Tim4 tethers apoptotic cells, and the integrin ␣ v ␤ 3 complex mediates engulfment in coordination with MFG-E8. A similar two-step engulfment of apoptotic cells was observed with mouse resident peritoneal macrophages. Furthermore, the Tim4/integrin-mediated engulfment by the Ba/F3 cells was enhanced in cells expressing Rac1 and Rab5, suggesting that this system well reproduces the engulfment of apoptotic cells by macrophages. Every day, billions of cells that are toxic, useless, and senescent die by apoptosis and are engulfed by macrophages, presumably to prevent the release of noxious materials from the dead cells (18). The system that efficiently removes apoptotic cells from the body appears to be quite elaborate, and some details of this process are unclear (25). Apoptotic cells present an "eat me" signal to macrophages, triggering their own engulfment. Among the various molecules proposed to be involved in this process, phosphatidylserine (PS) is a strong candidate for the "eat me" signal (11). PS is transferred caspase dependently from the inner leaflet to the outer leaflet of the plasma membrane (13), and masking PS inhibits the engulfment of apoptotic cells by macrophages (2, 6, 11).By identifying monoclonal antibodies that have positive or negative effects on the engulfment of apoptotic cells, we previously identified two molecules, milk fat globule epidermal growth factor (EGF) factor VIII (MFG-E8) and T cell immunoglobulinand mucin-domain-containing molecule 4 (Tim4), that enhance engulfment (6, 15). MFG-E8 is a secreted protein of 75 kDa that binds to PS via its factor VIII-homologous domain. It also binds to the integrin ␣ v ␤ 3 complex in macrophages via an RGD motif in MFG-E8's EGF domain, thus bridging apoptotic cells and macrophages. Tim4 is a type I membrane protein of 70 kDa that binds to PS via the immunoglobulin-like domain in its extracellular region. Its cytoplasmic region is 43 amino acids long and is dispensable for...

In metazoan tissues, cells decide their fates by sensing positional information provided by specialized morphogen proteins. To explore what features are sufficient for positional encoding, we asked whether arbitrary molecules (e.g., green fluorescent protein or mCherry) could be converted into synthetic morphogens. Synthetic morphogens expressed from a localized source formed a gradient when trapped by surface-anchoring proteins, and they could be sensed by synthetic receptors. Despite their simplicity, these morphogen systems yielded patterns reminiscent of those observed in vivo. Gradients could be reshaped by altering anchor density or by providing a source of competing inhibitor. Gradient interpretation could be altered by adding feedback loops or morphogen cascades to receiver cell response circuits. Orthogonal cell-cell communication systems provide insight into morphogen evolution and a platform for engineering tissues.