Pooled CRISPR screens identify genes and non-coding genomic regions that regulate red blood cell density

Some of these genes have previously been implicated in RBC biology (e.g. ATP2B4, CCND3, EPOR) although many are novel (e.g. CHTF8, CTU2, DNASE2). We confirmed that deletions in the osmotic stress response kinase gene OXSR1 increase cell density, and a phosphoproteome analysis in OXSR1-depleted cells indicated that this phenotype is accompanied with a dephosphorylation of the upstream kinase WNK1 and the downstream target KCC3 (SLC12A6).

We also combined CRISPR perturbations and RNA-sequencing to show how a non-coding genomic sequence near rs13255015 regulates the expression of the transcription factor ZFAT in cis and SLC4A1 in trans. SLC4A1 encodes Band3, a known regulator of RBC hydration and volume. Our results suggest experimental strategies to characterize GWAS findings and provide new molecular insights into the regulation of complex RBC traits.