TIF1γ Counteracts Ferroptosis to Drive Erythroid Progenitor Differentiation

Understanding in-vivo mechanisms of hematopoiesis is critical for developing directed blood differentiation approaches to treat blood disorders such as leukemias. Zebrafish moonshine (mon) mutant embryos defective for transcriptional intermediary factor 1 gamma (tif1γ), a conserved transcription elongation and chromatin factor, lack red blood cells due to a block in hematopoietic stem cell differentiation along the erythroid lineage. We recently showed that TIF1γ plays a critical role in mitochondrial metabolism, including maintaining adequate coenzyme Q levels. Through a chemical suppressor screen for the mon mutant, we identified inhibitors of the essential mitochondrial pyrimidine synthesis enzyme dihydroorotate dehydrogenase to promote erythroid differentiation in mon mutants due to its functional link to the electron transport chain. In agreement, our in-vivo metabolomics analyses identified nucleotide metabolism as the most significantly altered processes in mon mutants, with elevated levels of uridine monophosphate and low levels of N-carbamoyl-L-aspartate. Interestingly, imbalances in nucleotide metabolism similar to those in mon mutants have also been reported in the presence of active ferroptosis, a pathway of programmed cell death due to iron-dependent lipid peroxidation. Through transcriptome profiling upon tif1γ loss of function in zebrafish embryos at the onset of hematopoiesis, we uncovered an expression signature indicative of activated ferroptosis. gpx4 and fsp1, inhibitors of ferroptosis acting in parallel pathways, were both downregulated, and chac1 and ptgs2, an enhancer and marker of ferroptosis, respectively, were upregulated. In addition, parallel genome-wide expression and chromatin immunoprecipitation analyses identified anti-ferroptotic genes as direct TIF1γ targets in human cells. In support of these results, we found mon mutants to exhibit increased levels of lipid peroxidation. Functionally, tif1γ loss of function synergizes with loss of gpx4 in blocking erythroid differentiation. These results demonstrate a tight coordination of nucleotide metabolism, mitochondrial respiration, and the regulation of lipid peroxidation as a key function of tif1γ-dependent transcription that drives cell fate decisions in the early erythroid lineage. Our work highlights the importance of the plasticity achieved by transcription regulatory processes such as transcription elongation for metabolic processes during lineage differentiation and could have therapeutic potential for blood diseases.