3097 – RNA EDITOR-EXONUCLEASE AXIS SCULPTS THE TRANSCRIPTOME DURING TERMINAL ERYTHROID DIFFERENTIATION

Abstract

Erythropoiesis is a committed process by which erythroid progenitors become mature red blood cells (RBCs). Reticulocytes are terminal-staged, immature RBCs which contain selected residual RNA after enucleation. Normally, reticulocytes are efficiently processed and typically represent a small percentage of cells in human peripheral blood. In contrast, when differentiated from iPS or CD34+ hematopoietic stem/progenitor cells in vitro, erythroid cells tend to arrest at the reticulocyte stage. This is the major bottleneck for the complete recapitulation of erythropoiesis in vitro, and a hindrance to modeling hemoglobinopathies in cell cultures. Recent studies have highlighted that uridylation by Terminal Uridylyl Transferases (TUTases 7/4, official names are ZCCHC6/11) occurs on a broad spectrum of RNA classes in mammalian cells. Oligo-uridylated RNA is then recognized by exoribonucleases and targeted for decay. We posited that this is the machinery behind RNA degradation that accompanies terminal erythropoiesis. Utilizing both constitutional and erythroid-specific conditional murine knockout models, we found that blood from TUTase Zcchc6 RNA editor knockout embryos indeed exhibit reticulocytosis with a terminal maturation defect, as documented by FACS, histology, and hematological profiling. Murine strains deficient in the downstream exonuclease, Dis3l2, phenocopied the RNA decay defect of the Zcchc6 KO. Furthermore, knockout models did not show signs of hemolysis suggesting a cell intrinsic and niche-independent role for the TUTase7-Dis3l2 axis in promoting red blood cell maturation. Modulating the expression of this axis holds promise for enhancing in vitro erythropoiesis and modeling of hemoglobinopathies. Future mechanism studies will also reveal a post-transcriptional regulatory machinery governing maturation from reticulocytes to erythrocytes.