RNA-sequencing comparing activation of wild-type transcription factor KLF1 with mutant KLF1 (KLF1-325K) in iPSC-derived macrophages.
Congenital dyserythropoietic anaemia (CDA) type IV has been associated with an amino acid substitution, Glu325Lys (E325K), in the transcription factor KLF1. Patients with CDA type IV present with a range of symptoms, including the persistence of nucleated red blood cells (RBCs) in the peripheral blood which reflects the known role for KLF1 within the erythroid cell lineage. The final stages of RBCs maturation and enucleation take place within the erythroblastic island (EBI) niche in close association with EBI macrophages. It is not known whether the detrimental effects of the E325K mutation in KLF1 are restricted to the erythroid lineage or whether deficiencies in macrophages associated with their niche also contribute to the disease pathology. To address this question, we generated iPSC lines genetically modified to express a KLF1-E325K-ERT2 protein that could be activated with 4OH-tamoxifen. We performed bulk RNA-sequencing on macrophages generated from these iPSCs, macrophages generated from one KLF1-E325K-ERT2 iPSC line (iCDA4.1) was compared to macrophages generated from one inducible KLF1-WT-ERT2 (K2) iPSC line which was derived from the same parental iPSCs (SFCi55) as the KLF1-E325K-ERT2 line. Overall design: Bulk RNA-sequencing was performed on macrophages generated from the parental (SFCi55), inducible KLF1-WT activation iPSC line (K2), and the inducible KLF1-E325K activation iPSC line. Macrophages were either untreated or treated with 100 nM of 4OH-tamoxifen (5 replicates of each for a total of 30 samples).