A forward RNAi screen identifies MAPK14 as a regulator of human HSPCs. (A) CB CD34⁺ cells were transduced with a pooled lentiviral library and cultured for 20 days. Cell samples harvested after 3 and 20 days were analyzed by next generation sequencing of proviral shRNAs. (B) Relative changes in shRNA distribution calculated as the normalized number of shRNA sequence reads at day 20 divided by the number of reads day 3. The mean from 3 independent screens is plotted in ascending order. (C) The relative enrichment in CD34⁺ cells at day 20 for 3 independent shRNAs (sh1 to sh3) targeting MAPK14. The mean value of all shRNAs in the screen is shown as a dotted line. (D) Western blot of p38 expression in CD34⁺ cells transduced with MAPK14-targeting shRNAs (sh1 to sh3) or control shRNA targeting β-galactosidase (shCtrl). (E) CD34 expression after 17 days of culture of CB CD34⁺ cells transduced with MAPK14-targeting shRNA or control shRNA (n = 3). (F) CD34 expression after 14 days of culture of CB CD34⁺ cells treated with the P38α inhibitor Ly2228820 or vehicle (DMSO; n = 3). Error bars represent SEM.