Long noncoding RNA GATA2AS influences human erythropoiesis by transcription factor and chromatin landscape regulation

• Human-specific lncRNA GATA2AS is a regulator of erythroid differentiation

• GATA2AS is a novel GATA2 and HBG activator through distinct mechanisms

LncRNAs are extensively expressed in eukaryotic cells and have been revealed to be important for regulating cell differentiation. Many lncRNAs have been found to regulate erythroid differentiation in the mouse. However, given the low sequence conservation of lncRNAs between mouse and human, our understanding of lncRNAs in human erythroid differentiation remains incomplete. LncRNAs are often transcribed opposite to protein coding genes and regulate their expression. Here we characterized a human erythrocyte-expressed lncRNA, GATA2AS, which is transcribed opposite to erythroid transcription regulator GATA2. GATA2AS is a 2080 bp-long, primarily nuclear-localized non-coding RNA that is expressed in erythroid progenitor cells and decreases during differentiation. Knockout of GATA2AS in human HUDEP2 erythroid progenitor cells using CRISPR-Cas9 genome editing to remove the transcription start site accelerated erythroid differentiation and dysregulated erythroblast gene expression. We identified GATA2AS as a novel GATA2 and HBG activator. Chromatin Isolation by RNA Purification (ChIRP) showed that GATA2AS binds to thousands of genomic sites and colocalizes at a subset of sites with erythroid transcription factors including LRF and KLF1. RNA-pulldown and RIP confirmed interaction between GATA2AS and LRF and KLF1. ChIP-sequencing showed that knockout of GATA2AS reduces binding of these transcription factors genome wide. ATAC-seq and H3K27ac ChIP-seq showed that GATA2AS is essential to maintain the chromatin regulatory landscape during erythroid differentiation. Knockdown of GATA2AS in human primary CD34+ cells mimicked results in HUDEP2 cells. Overall, our results implicate human-specific lncRNA GATA2AS as a regulator of erythroid differentiation by influencing erythroid transcription factor binding and the chromatin regulatory landscape.