Endogenous variability in transcription factor concentrations shapes their genome-wide occupancy

The control of transcription factor (TF) concentrations is crucial for the precise regulation of gene expression and cell fate decisions during development. Yet, TF concentrations can display substantial temporal fluctuations and intercellular variations. How TF levels quantitatively shape genome-wide occupancy patterns remains largely unexplored. Here, we systematically investigate how physiological fluctuations in the concentrations of the pluripotency TFs OCT4, SOX2, and NANOG impact their genomic binding profiles in mouse embryonic stem cells. We uncover distinct concentration-dependent binding behaviors for each TF. NANOG occupancy increased monotonously with its concentration, even though a common set of regions made accessible by OCT4 and SOX2 are highly and uniformly bound at all NANOG concentrations. In contrast, SOX2 occupancy does not increase with its concentrations, while OCT4 displayed an intermediate behavior. Our results challenge the notion that TF binding follows simple mass-action dynamics and reveal a core set of pluripotency regions that are bound by all three TFs at all TF concentrations, revealing how pluripotency can be maintained despite marked fluctuations in core pluripotency TFs.