Figure S6. Related to Fig 7. Stromal SDF-1 and TGF-β mediate the formation of invasive and metastatic breast tumor clusters with E^hi and E/M states via Src activation.

(A) Real-time PCR of DCIS^cnt2cy and DCIS^CAF2cy using the indicated primers. The DCIS^cnt2cy and DCIS^CAF2cy were extracted from tumor xenografts generated by DCIS cells admixed with control fibroblasts and CAFs expressing GFP or TβRII ecto, respectively. (B) Real-time PCR of DCIS^cnt1cy and DCIS^CAF1cy expressing the indicated shRNAs measuring E-cad expression. The DCIS^cnt1cy and DCIS^CAF1cy were extracted from 30-d-old tumor xenografts generated by DCIS cells expressing the indicated shRNAs admixed with control fibroblasts and CAFs, respectively. (C) Immunofluorescence of DCIS cells treated with recombinant SDF-1 (100 ng/ml) and/or TGF-β1 (10 ng/ml) for 48 h using anti–E-cad or ZEB1 antibody. E-cad⁺ cancer cells (simple arrows) and nuclear ZEB1⁺ cancer cells (triangular arrows) are shown. Scale bars, 10 μm. (D) Real-time PCR of MCF-7-ras cells treated with recombinant SDF-1 and/or TGF-β1 for 24 h to measure the indicated gene expressions. Data information: Asterisk indicates a significant difference relative to DCIS^CAF2cy extracted from tumor with CAFs expressing GFP (A) and GFP-shRNA–expressing DCIS^CAF1cy (B), and between the depicted groups (D). t test (A, B, D). n.s.: not significant (D). Error bars, SE.