Embryonic stem cell ERK, AKT, plus STAT3 response dynamics combinatorics are heterogeneous but NANOG state independent

• Published in: Stem cell reports Vol. 18; no. 6; pp. 1295 - 1307
• Main Authors: Reimann, Andreas, Kull, Tobias, Wang, Weijia, Dettinger, Philip, Loeffler, Dirk, Schroeder, Timm
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 13.06.2023; Elsevier
• Subjects: AKT; dynamics; ERK; ESC; heterogeneity; signaling; STAT3; time-lapse; dynamics; signaling; ESC; time-lapse; STAT3; AKT; heterogeneity; ERK
• ISSN: 2213-6711; 2213-6711
• DOI: 10.1016/j.stemcr.2023.04.008

Signaling is central in cell fate regulation, and relevant information is encoded in its activity over time (i.e., dynamics). However, simultaneous dynamics quantification of several pathways in single mammalian stem cells has not yet been accomplished. Here we generate mouse embryonic stem cell (ESC) lines simultaneously expressing fluorescent reporters for ERK, AKT, and STAT3 signaling activity, which all control pluripotency. We quantify their single-cell dynamics combinations in response to different self-renewal stimuli and find striking heterogeneity for all pathways, some dependent on cell cycle but not pluripotency states, even in ESC populations currently assumed to be highly homogeneous. Pathways are mostly independently regulated, but some context-dependent correlations exist. These quantifications reveal surprising single-cell heterogeneity in the important cell fate control layer of signaling dynamics combinations and raise fundamental questions about the role of signaling in (stem) cell fate control. •Simultaneous dynamics quantification of three signaling pathways in single ESCs•Unexpected multilevel heterogeneity of signaling dynamics and their combinations•Link of ERK signaling dynamics to ESC morphology, cell cycle, and pluripotency state Reimann et al. simultaneously quantify the activity dynamics of ERK, AKT, and STAT3 in single ESCs and find striking multilevel heterogeneity, even in ESCs currently assumed to be homogeneous. Dynamics are partly dependent on cell cycle but independent of pluripotency state. The single-cell dynamics quantification approach for multiple pathways is now also applicable to other pathways and cell systems.