Upon productive immunoglobulin gene rearrangement, expression of a functional pre-B cell receptor (pre-BCR) initiates positive selection of pre-B cells, clonal expansion and self-renewal 1-2 . Studying mechanisms driving this first wave of B-lymphopoiesis, we identified the G-protein coupled receptor Lgr5 as an essential initiator of positive selection. Lgr5 was extensively studied as determinant of stem cell populations in multiple tissues 3-6 , but not in B-cells. While undetectable throughout the hematopoietic system, positively selected pre-B cells were marked with a sharp peak of Lgr5 expression. Conditional deletion of Lgr5 preceding the pre-BCR checkpoint induced negative selection and complete abortion of B-cell development. Proteomic studies of Lgr5-ablation revealed massive (>250-fold) accumulation of -catenin and suppression of MYC. Lgr5-deficient pre-B cells fully recovered by concurrent -catenin-deletion, demonstrating a central role of Lgr5-mediated restraint of -catenin at the pre-BCR checkpoint. In other cell types, -catenin/TCF4 complexes drive transcriptional activation of MYC 7-9 . Instead of TCF4, proximity-based interactome studies in pre-B cells identified the Blymphoid transcription factors IKZF1 and IKZF3 10-11 as -catenin-binding partners, which had the opposite effect and caused transcriptional repression of MYC. On positively selected pre-B cells, Lgr5 prevented accumulation of -catenin and formation of complexes with IKZF1 and IKZF3, which relieved transcriptional repression of MYC. Activating β-catenin-mutations are common throughout all main types of cancer 7-8 , but were conspicuously absent in pre-B leukemia (B-ALL). Like pre-B cells, B-ALL cells were uniquely sensitive to genetic and pharmacological β-catenin hyperactivation, which recapitulated the effects of Lgr5-deletion and compromised colony formation and leukemia-initiation. A new LGR5 antibody-drug conjugate targeted leukemia-initiating cells in patient-derived B-ALL and achieved long-term disease-control. Likewise, small molecule hyperactivation of -catenin selectively killed B-ALL but not other cell types. Hence, Lgr5-mediated restraint of -catenin activation is essential for B-lymphopoiesis and revealed an unexpected vulnerability that can be leveraged for the treatment of drug-resistant B-ALL. Lgr5 is essential for the initiation of B-lymphopoiesisOnce early B-lymphocyte precursors have productively rearranged immunoglobulin (Ig) V, D and J gene segments, expression of a functional Ig -heavy chain as part of the pre-B cell receptor (pre-BCR) results in a strong positive selection signal to initiate clonal expansion and the first wave of B-lymphopoiesis 1, 2 . To study mechanisms of positive selection and pre-B cell self-renewal, we analyzed gene expression changes that are induced at the onset of Ig -heavy chain signaling. Among the most prominent changes at the pre-BCR checkpoint, we identified upregulation of the Leucine-rich repeat-containing G-protein coupled receptor 5 (Lgr5; Fig. 1a and b, Extended data ...