A mosaic gastruloid model highlights the developmental stage-specific restriction of cell competition in mammalian pre-gastrulation

Abstract

In early mammalian embryogenesis, the selective elimination of suboptimal cells is critical for developmental integrity. Cell competition (CC) is a cell non-autonomous quality control in which "winner" cells eliminate viable but suboptimal "loser" cells based on their relative difference in fitness. Due to its central role in fitness perception, loss of p53 results in the emergence of "supercompetitor" cells, which stand at the apex of cell competition and induce apoptosis in neighboring wild-type (WT) cells. Here, we investigate CC dynamics using mosaic 3D mouse gastruloids, an embryonic stem cell (ESC)-based in vitro model of gastrulation, composed of defined numbers of WT and p53-KO cells. In mosaic gastruloids, even low numbers of p53-KO cells robustly outcompete WT cells, and introduction of as few as two p53-KO cells is sufficient to measurably impair neighboring WT cell growth. CC in gastruloids is independent of cell proliferation rates, nutrient availability, or reactive oxygen species (ROS), and not influenced by Nodal and ERK signaling. However, we observe that Wnt and BMP signaling protect from CC, which is exclusively driven by intrinsic apoptosis, as indicated by Bcl2-mediated complete rescue of WT cells. During gastruloid development, CC is temporally restricted to a window of 48-96 hours after aggregation, mirroring embryonic days E5.5-E7.5 in the mouse. Heterochronic mosaic gastruloid experiments demonstrate that relative differences in pluripotency levels are neither necessary nor sufficient to cause supercompetition, but that CC is contingent on both competitors residing within the developmental window permissive to CC. Neither pluripotent mosaic 3D aggregates, nor 3D EpiGastruloids, which model more advanced developmental processes, display any competition, supporting the hypothesis that developmental CC is specific to the onset of gastrulation. Our findings offer insights into the mechanisms of cell fitness evaluation in mammalian embryogenesis and establish gastruloids as a powerful 3D model for investigating developmental stage-specific cell competition.