The transcription factor PU.1 supports myeloid cell lineage differentiation by establishing transcriptional networks that involve several feedback loops. Here, Rothenberg and colleagues describe a previously unappreciated mechanism of positive feedback regulation by PU.1 that is controlled by the cell cycle.

Credit: NPG

The authors used knock-in PU.1 reporter mice to screen PU.1 expression levels during the differentiation of lineage-negative KIT+CD27+ fetal liver progenitor cells. Consistent with previous findings, these early multipotent haematopoietic progenitor cells expressed intermediate PU.1 levels, and they increased their expression of PU.1 as they differentiated into macrophages but decreased their expression of PU.1 as they differentiated into B cells. Remarkably, analysis of time-lapse microscopy movies showed that PU.1 synthesis rates did not change during the differentiation of myeloid progenitor cells into macrophages, but the length of the cell cycle increased. As the half-life of PU.1 is longer than the duration of the cell cycle of haematopoietic progenitor cells, this cell cycle lengthening led to PU.1 accumulation. By contrast, the length of the cell cycle in B cells was comparable with the length of the cell cycle in progenitor cells, and PU.1 expression was downregulated through transcriptional regulation during B cell differentiation.

transcription factors that have a long half-life might prolong the cell cycle to sustain a stable cell fate

So, is cell cycle control essential for PU.1 accumulation and myeloid cell differentiation? To test this, the authors increased the length of the cell cycle in progenitor cells by either transducing them with cyclin-dependent kinase (CDK) inhibitors or treating them with small-molecule cell cycle inhibitors. Cell cycle-arrested cells had higher levels of PU.1, as well as a higher output of myeloid cells, which required PU.1 activity as it was suppressed in cells that had been transduced with a PU.1 antagonist. Moreover, the transduction of progenitor cells with exogenous PU.1 did not increase the synthesis rate of PU.1, but instead increased the length of the cell cycle, which led to PU.1 accumulation. Consistent with these observations, the forced expression of exogenous PU.1 did not further activate the transcription of endogenous PU.1, but it decreased the transcription of cell cycle-promoting factors, including cyclin D2, CDC25A (also known as M phase inducer phosphatase 1), MYB and MYC.

Thus, PU.1 levels are not only controlled through direct transcriptional feedback regulation (as seen in differentiating B cells) but also through cell cycle lengthening, which seems to result from, and to maintain, high PU.1 levels in developing macrophages. Mathematical modelling confirmed that lymphoid and myeloid cell differentiation can depend on these two regulatory principles, and the authors suggest that transcription factors that have a long half-life might prolong the cell cycle to sustain a stable cell fate.