miR-125b inhibits B-cell release from the bone marrow through direct targeting of S1PR1. (A) Schematic representation of the S1PR1 3′ UTR showing the conserved miR-125b seed region. (B) S1PR1 transcript expression levels in B cells purified from the bone marrow of Eμ/miR-125b-Tg and control mice. (C) Relative luciferase expression in HEK293T cells transfected with luciferase reporter construct bearing the S1PR1 3′ UTR immediately downstream (S1PR1UTR) and either a miR-125b overexpression vector (MG-125b) or a control vector (MG). (D) S1pr1 protein expression levels in B cells purified from the bone marrow of Eμ/miR-125b-Tg and control mice. Data represent 2 independent experiments. (E) Enumeration of B cells (B220⁺) and immature B cells (B220⁺CD93⁺) in the peripheral blood of reconstituted mice at 7 weeks after reconstitution (n = 10 mice per group). (F) Schematic representation of small gRNA target (red) and protospacer adjacent motif (green) sequence designed to edit genomic sequence in the mouse S1PR1 3′UTR locus. (G) Detection of genome editing in the S1PR1 3′ UTR locus using a surveyor assay. (H) Enumeration of B cells (B220⁺) in peripheral blood of reconstituted mice at 7 weeks after reconstitution with CRISPR/Cas9-edited HSPCs (n = 10-12 mice per group). Data represent 2 independent experiments and represent mean ± SEM. *P < .05; ***P < .001; NS, not significant.