Stem cell antigen (Sca)-1/Ly6A, a glycerophosphatidylinositollinked surface protein, was found to be associated with murine stem cell-and progenitor cell-enriched populations, and also has been linked to the capacity of tumor-initiating cells. Despite these interesting associations, this protein's functional role in these processes remains largely unknown. To identify the mechanism underlying the protein's possible role in mammary tumorigenesis, Sca-1 expression was examined in Sca-1 +/EGFP mice during carcinogenesis. Mammary tumor cells derived from these mice readily engrafted in syngeneic mice, and tumor growth was markedly inhibited on down-regulation of Sca-1 expression. The latter effect was associated with significantly elevated expression of the TGF-β ligand growth differentiation factor-10 (GDF10), which was found to selectively activate TGF-β receptor (TβRI/II)-dependent Smad3 phosphorylation. Overexpression of GDF10 attenuated tumor formation; conversely, silencing of GDF10 expression reversed these effects. Sca-1 attenuated GDF10-dependent TGF-β signaling by disrupting the heterodimerization of TβRI and TβRII receptors. These findings suggest a new functional role for Sca-1 in maintaining tumorigenicity, in part by acting as a potent suppressor of TGF-β signaling.S tem cell antigen (Sca)-1 is a member of the Ly6A superfamily of glycerophosphatidylinositol (GPI)-anchored membrane proteins (1), which is associated with murine stem and progenitor cell populations in several tissues. In the mammary gland, Sca-1-positive cells are able to reconstitute the cleared fat pad (2, 3) and give rise to alveolar and ductal structures (4). In addition to Sca-1's normal role in stem cell self-renewal, Sca-1 expression is elevated in malignant tissues, such as retinoblastomas (5), prostate tumors (6), mammary tumors (7, 8) and chronic myeloid leukemia (9), which generally reflects a more aggressive phenotype (10). Despite these associations, the role of Sca-1 in these processes remains largely unknown. To address this question, we used Sca-1 +/EGFP mice, in which EGFP is under the control of the Sca-1 locus (11), to study mammary tumorigenesis. We found that Sca-1 suppresses TGF-β signaling by inhibiting expression of the TGF-β family ligand GDF10 and binding to the type I TGF-β receptor (TβRI), thereby inhibiting ligand-induced TGF-β receptor complex formation and Smad3 phosphorylation. This study is the first to demonstrate specific functions for Sca-1 and GDF10 in tumorigenesis.
ResultsSca-1 Is Increased Early in Mammary Tumorigenesis. To explore the role of Sca-1 in tumorigenesis, we induced mammary tumors in heterozygous Sca-1 +/EGFP mice (11) using medroxyprogesterone (MPA) and 7,12-dimethylbenz[a]anthracene (DMBA) (8,12,13). Cultures of primary mammary epithelial cells exhibited enhanced EGFP fluorescence coincident with increased Sca-1 expression immediately after treatment with MPA and DMBA (( Fig. S1 A and B ), suggesting that the Sca-1 locus is activated at the onset of carcinogenesis. Both Sca-1 +/EGFP a...