Multicellular organisms rely on the coordinated function of cells and tissues to support homeostasis. This cooperation begins very early in mammalian development, when embryos consist of single-digit cell numbers and functionally specialized groups of cells are formed by the time the embryo is a few hundred cells. Noise in gene expression generates early cellular heterogeneity that is utilized by signaling mechanisms to drive stable commitment to cell fates. Though the pathways required for development are characterized, we do not understand the temporal dynamics of signaling activity that govern cellular decision making or how individual decisions are collectively regulated to robustly generate embryos of consistent lineage composition. We investigated a critical lineage segregation event in preimplantation embryos using a single cell reporter for ERK kinase and describe variable activation that is coordinated with cell cycle progression. We employ a novel reporter of the lineage defining transcription factor NANOG to analyze gene expression and posttranscriptional regulation dynamics underlying this process and propose that ERK variability at mitotic exit reinforces cell fates in the embryo and embryonic stem cells. Direct measurement of kinase signaling dynamics and cell type specification in the same embryos required methodological advancement and we shared our novel techniques with the field.
Our study includes the first reported measurements of ERK activity in live mammalian embryos and our analysis of cell cycle specific regulation of kinase activity and gene expression dynamics prompt reconsideration of earlier models of FGF-dependent signaling in the embryo. The method we developed enables further molecular characterization of ERK, cell cycle proteins, transcriptional regulators, and their interactions. Future investigation of cellular decision making in pluripotent cells will elucidate how single cells cooperate to build complex tissues and expand our ability to control cells in technology and medicine.