EKLF/Klf1 regulates erythroid transcription by its pioneering activity and selective control of RNA Pol II pause-release

EKLF/Klf1 is a Zinc-finger transcription activator essential for erythroid lineage commitment and terminal differentiation. Using ChIP-Seq, we investigate EKLF DNA binding and transcription activation mechanisms during mouse embryonic erythropoiesis. Our study focuses on global EKLF binding dynamics during embryonic erythropoiesis in primary WT and Nan/+ mouse fetal liver, and its correlation with chromatin accessibility, CBP occupancy, histone acetylation, and finally its effect on RNA Polymerase II pausing and elongation. Our goal is to elucidate the mechanisms of transcription activation by EKLF/Klf1 during embryonic erythropoiesis in vivo and in the context of RNA pol II pause-release control. Additionally, we aim to understand the unusually severe effects of conservative E to D change in Nan-EKLF and the molecular mechanisms leading to dominant anemia through global gene dysregulation. Overall design: Chromatin Immunoprecipitation and NGS (ChIP-seq) for EKLF/Klf1, CBP, H3K27ac, RNA Polymerase II C-terminal domain phospho-Serine5, RNA Polymerase II C-terminal domain phospho-Serine2, ATAC-Seq in mouse E13.5 fetal liver from WT and Nan/+ embryos. All ChIPs except RNA Pol II are biological triplicates from littermate fetal livers. RNA Pol II was performed in duplicate for each experiment. ATAC-Seq was performed in biological duplicates