ARID1B, a molecular suppressor of erythropoiesis, is essential for the prevention of Monge’s disease

In this work, we used genomic and molecular functional assays to understand the role of ARID1B in regulating red blood cells under hypoxia using iPS cells derived from non-CMS adapted individuals and comparing the non-CMS wild type cells to non-CMS cells with ARID1B-KD using in-vitro iPSC-derived model system. hiPSC-derived cell erythroid cells were generated from the adapted non-CMS individuals (referred to as non-CMS) and cell line was created in which ARID1B was knocked down (using shRNA lentivirus) in these non-CMS cells (Referred to as ARID1B-KD). For ATAC-Seq, transposition reactions were performed on 50,000 non-CMS and non-CMS with ARID1B KD (replicates, n = 3) hiPSC-derived erythroid cells grown under hypoxic and normoxic conditions for two weeks as previously described Buenrostro et al., 2013 (52). Briefly Cells were washed with PBS followed by lysis with cold lysis buffer. Cells were then suspended in 50 μl 1X reaction buffer. Transposase reactions were carried out at 37°C for 30 minutes and DNA was purified using ChIP DNA Clean & Concentrator kits. DNA was amplified using the dual barcoded primers (Illumina) using NEBNext High-Fidelity 2XPCR Master Mix for 14 cycles. Resulting libraires were sequenced on an Illumina Novaseq platform generating Paired-End, 50bp (PE50) reads with an average of over 100 million reads per sample.