Fig. 1

ASORL1 expression is significantly higher in breast cancer cell lines with high ERBB3 expression (CCLE; N = 59). RMA: robust multi-array average. Data are mean ± SD; statistical analysis: Mann–Whitney U. BSORL1 expression is higher in tumors with high ERBB3 and ERBB2 expression (cBioPortal; N = 476). Violin plot boxes represent median and 25th and 75th percentiles (interquartile range), and whiskers extend to maximum and minimum values. See methods for details on how the groups were defined. C Hrg β-1 increases SorLA levels. BT-474 cells were stimulated with 20 ng.mL⁻¹ Hrg β-1 for the indicated times. Representative immunoblotting of SorLA, AKT(p)S473, total AKT, ERK1/2(p)T202/Y204, total ERK1/2, with α-tubulin as a loading control. D Quantification of SorLA levels normalized to loading control and relative to non-stimulated (0 h) cells. E Hrg β-1 increases SORL1 expression. Quantification of SORL1 mRNA levels, relative to HPRT1, determined with RT-qPCR in BT-474 cells stimulated with 20 ng.mL⁻¹ Hrg β-1 for the indicated time points relative to non-stimulated (0 h) cells. F Representative immunoblotting of SorLA, AKT(p)S473, total AKT, ERK1/2(p)T202/Y204, total ERK1/2, with α-tubulin as a loading control from control (mCherry)- or Hrg β-1-overexpressing BT-474 cells. G Quantification of SorLA levels normalized to loading control and relative to control cells. H SMDF increases SorLA expression. BT-474 cells were cultured on a monolayer of mCherry-positive control or SMDF-overexpressing fibroblasts (TIFF). BT-474 cells were FACS sorted (see “Methods”) and cell lysates were analyzed for SorLA, AKT(p)S473, total AKT, ERK1/2(p)T202/Y204, total ERK1/2, with α-tubulin as a loading control. I Quantification of SorLA levels normalized to loading control and relative to control TIFF cocultured cells. J Representative immunoblotting of SorLA, HER2, and HER3, with α-tubulin as a loading control from parental BT-474 and brain-tropic metastasis variant BT-474-Br cells. K Quantification of indicated protein levels normalized to loading control and relative to BT-474 cells. L Quantification of SORL1 mRNA levels, normalized to HPRT1, determined with RT-qPCR in parental BT-474 and BT-474-Br cells relative to BT-474 cells. Unless otherwise indicated, data are mean ± SD from three independent biological experiments; statistical analysis: Student’s t test (unpaired, two-tailed, unequal variance).