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Supplementary Tables S1 to S8. S1 shows de novo AML patient characteristics. S2 shows JAM-C expression in monocytic AML. S3 shows multivariate Cox regression data. S4 shows genes selected for single cell gene expression study. S5 shows statistical analysis of genes used in Fig 3C. S6 shows limit dilution assay data in vitro. S7 shows limit dilution assay data in vivo. S8 shows frequencies of human and mouse cells from Fig 5E.

Supplementary Figures S1 to S7. S1 shows JAM-C, CD90 and CD45RA expression in leukemic fraction. S2 shows JAM-C expression in leukemic cells and HSPCs. S3 shows JAM-C expression in cell lines and genes differentially expressed between KG1/JAM-CNeg and KG1/JAM-CPos variant cell lines. S4 shows characterization of engrafting and non-engrafting AML patient samples. S5 shows cell cycle and proliferation properties of KG1/JAM-CNeg and KG1/JAM-CPos variant cells. S6 shows signaling pathways in KG1/JAM-CNeg and KG1/JAM-CPos. S7 shows comparison of SFK phosphorylation in CD45dimCD34+ and CD45dimCD34- leukemic cells.

Acute myeloid leukemia (AML) originates from hematopoietic stem and progenitor cells that acquire somatic mutations, leading to disease and clonogenic evolution. AML is characterized by accumulation of immature myeloid cells in the bone marrow and phenotypic cellular heterogeneity reflective of normal hematopoietic differentiation. Here, we show that JAM-C expression defines a subset of leukemic cells endowed with leukemia-initiating cell activity (LIC). Stratification of de novo AML patients at diagnosis based on JAM-C–expressing cells frequencies in the blood served as an independent prognostic marker for disease outcome. Using publicly available leukemic stem cell (LSC) gene expression profiles and gene expression data generated from JAM-C–expressing leukemic cells, we defined a single cell core gene expression signature correlated to JAM-C expression that reveals LSC heterogeneity. Finally, we demonstrated that JAM-C controls Src family kinase (SFK) activation in LSC and that LIC with exacerbated SFK activation was uniquely found within the JAM-C–expressing LSC compartment. Cancer Res; 77(23); 6627–40. ©2017 AACR.

Supplementary methods describing flow-cytometry, cumulative relapse incidence calculation and mass spectrometry.

Galectins are glycan binding proteins translating the sugar-encoded information of cellular glycoconjugates into many physiological activities including immunity, cell migration, and signaling. During early B lymphocytes (BL) development at the pre-B cell stage, BL express the pre-B cell receptor (pre-BCR) and are supported by mesenchymal stromal cells secreting Galectin-1 (Gal-1). Gal-1 interacts with glycosylated receptors from stromal and pre-B cell surfaces but also with the pre-BCR through a direct carbohydrate-independent contact. How this interaction might interplay with the glycan-decoding function of Gal-1 is unknown. Here, we investigated Gal-1 binding to cell surface ligands using NMR spectroscopy on native membranes. We showed that pre-BCR regulates Gal-1 binding to specifically target α2,3-sialylated receptors on pre-B cells. Upon pre-BCR interaction, dynamic changes resulted in additional contacts with α2,3-sialylated glycans converting Gal-1 from an exo- to an endo-type lectin. Remarkably, this selectivity switch is able to promote pre-B cell survival. Altogether, we shed light on a new mechanism allowing fine-tuning of Galectin specificity at the cell surfaces.