Early enforcement of cell identity by a functional component of the terminally differentiated state

How progenitor cells can attain a distinct differentiated cell identity is a challenging problem given that critical transcription factors are often not unique to a differentiation process and the fluctuating signaling environment in which cells exist. Here we test the hypothesis that a unique differentiated cell identity can result from a core component of the differentiated state doubling up as a signaling protein that also drives differentiation. Using live single-cell imaging in the adipocyte differentiation system, we show that progenitor fat cells (preadipocytes) can only commit to terminally differentiate after upregulating FABP4, a lipid buffer that is highly enriched in mature adipocytes. Upon induction of adipogenesis, we show that after a long delay, cells first abruptly start to engage a positive feedback between CEBPA and PPARG before then engaging, after a second delay, a positive feedback between FABP4 and PPARG. These sequential positive feedbacks both need to engage in order to drive PPARG levels past the threshold for irreversible differentiation. In the last step before commitment, PPARG transcriptionally increases FABP4 expression while fatty-acid loaded FABP4 binds to and increases PPARG activity. Together, our study suggests a control principle for robust cell identity whereby a core component of the differentiated state also promotes differentiation from its own progenitor state. HIGHLIGHTSO_LIFatty-acid loaded FABP4 binds to and increases PPARG expression, thereby turning on PPARG positive feedback loops that further increase PPARG expression. C_LIO_LIFABP4 critically controls the second phase of adipogenesis between activation of the feedback loops and reaching the threshold to differentiate. C_LIO_LIOnly a small fraction ([~]10%) of the FABP4 levels typically attained in mature fat cells is needed to commit cells to the differentiated state, thus providing an explanation for why maintenance of the mature adipocyte state is so robust. C_LI