Interrogating Post-Transcriptional Mechanisms of Fetal Hemoglobin Regulation

Elevated levels of fetal hemoglobin (HbF) significantly ameliorate clinical outcomes for patients with beta-hemoglobinopathies, such as sickle cell disease (SCD). The only FDA-approved drug for treating SCD through inducing HbF is hydroxyurea, however the mechanism of action is unknown with variable effectiveness among patients. Thus, there remains a strong interest to identify more robust means of upregulating HbF, such as specific inhibition of HbF repressors. BCL11A and LRF are well-characterized transcription factors that independently repress the fetal type b-globin like genes HBG1 and HBG2 but their therapeutic potential is limited by challenging druggability and critical developmental function. However, upstream regulation of these factors, such as post-transcriptional mechanisms, are not well studied and may house novel therapeutic targets. To this end, we employed a CRISPR/Cas9 based screening approach to interrogate a library of RNA binding proteins (RBP) in the context of HbF regulation. Using HUDEP2 cells, a human adult-type erythroid progenitor cell line, we screened 341 human RBPs and identified four candidate RBPs, none of which have previously been implicated in HbF regulation. Of these candidates, RNA Binding Motif 12 (RBM12) showed the greatest level of HbF induction following in vitro depletion.