Extended Data Figure 4. MSI2 KD impairs secondary CFU replating potential and HSC engraftment capacity.
a, Left: Schematic of MSI2- and control RFP-targeted shRNA lentiviruses. Right: Confirmation of MSI2 protein knockdown (both isoforms that can be detected by western blot) in transduced NB4 cells. b, CFU production by shMSI2 and shControl transduced Lin− CB (n=8 cultures from 4 experiments). c, Secondary CFU output from shMSI2 and images of representative secondary CFU (scale bar = 200 μm, performed on n=4 cultures from 2 experiments). d, Fold change in transduced cell number after 7 days in culture (n=4 experiments). e, Growth curves of cultures initiated with transduced Lin− CB cells (n=4 experiments). f, Experimental design to readout changes in HSC capacity with MSI2 KD. g, Left: Representative flow analysis of transduced CD34+ CD38− derived human chimerism in NSG mouse BM. Right: Shown is the ratio of the percentage of GFP+ cells in the CD45+ population post-transplant to the initial pre-transplant GFP+ cell percentage. Dotted line indicates the proportion of GFP+ cells is unchanged relative to input. (One sample t-test, no change = 1, n = 6 shControl and n = 8 shMSI2 mice pooled from 2 experiments). h, Representative flow plots and summary of multilineage engraftment with shControl and shMSI2 (gated on GFP+ cells). Western blot source data is shown in Supplementary Figure 1. Data presented as mean ± SEM. *p<0.05; ***p<0.001.