GATA Factor Switching during Erythroid Differentiation Is Facilitated By FBW7 Mediated Clearance of GATA2

Erythroid differentiation is controlled by the dynamic exchange of GATA family transcription factors. During early erythroid maturation, high GATA2 levels activate progenitor genes such as c-kit-, c-myc, and GATA2 itself. In contrast, GATA1 levels are low in early progenitor cells, but rise during terminal maturation. During this process GATA1 turns off GATA2 controlled early progenitor genes and activates terminal maturation genes, such as globin genes, heme biosynthesis enzymes, and iron transporters. This involves the exchange of GATA1 for GATA2 at key chromatin sites, the so-called "GATA factor switch". GATA factor switching is facilitated by the much shorter half-life of GATA2 (~30-60 min) compared to GATA1 (>4-6 hrs). We and others recently demonstrated that the E3 ubiquitin ligase adaptor protein FBW7 contributes to GATA2's relative instability. This prompted us to dissect the role of FBW7 during GATA switching and erythroid differentiation. We deleted the Fbw7 gene using CRISPR/Cas9 gene editing in the inducible G1-ER murine erythroid cell line. This resulted in the delayed clearance of GATA2 during differentiation. RNA-seq analysis at an early time points (7 hr) demonstrated impaired repression of GATA2 regulated genes and reduced activation of GATA1 target genes. Globally, altered gene expression was enriched for GATA factor switch genes. This ultimately resulted in delayed erythroid maturation. We also found that Fbw7 mRNA transcript levels increase during erythroid maturation in wild type cells. We identified a site ~40kb upstream of the Fbw7 gene transcriptional start site, which is itself a GATA factor switch site. We propose that FBW7 facilitates GATA factor switching by promoting the clearance of GATA2 from GATA factor switch sites. Moreover, we suggest that GATA factor switching at the Fbw7 locus itself reinforces the commitment of erythroid cells to terminal maturation, by enhancing the clearance of GATA2 and other Fbw7 progenitor target gene proteins such as c-Myc and c-Myb. As Fbw7 recognition of GATA2 requires phosphorylation of GATA2's degron motif, this suggests that signaling pathways, acting through Fbw7, may modulate erythroid maturation kinetics.