Extended Data Fig. 7: Depletion of ITCH triggers p63-mediated basal differentiation and metastatic outgrowth of mammary tumours.
From: Autophagy-targeted NBR1–p62/SQSTM1 complexes promote breast cancer metastasis by sequestering ITCH
a, Quantification of total number of metastases per section of lung for CTRL (n = 12 mice), WT-NBR1 (n = 11) and del-ZnF-NBR1 (n = 10). shCTRL and WT-NBR1 cohorts were also used as controls in Extended Data Fig. 3d. b, Quantification of total number of metastases per section of lung for shCTRL (n = 13 mice), shATG7 (n = 6), shITCH, #1 (n = 25), shITCH, #2 (n = 24), shKEAP1, #1 (n = 12) and shKEAP1, #2 (n = 12). Mice in shCTRL and shATG7 cohorts are also used in Extended Data Fig. 1d. Related to Fig. 8g–l. c,d, Control (shCTRL) and ITCH-depleted MDA-MB-231 cells immunostained for p63 (green) and CK14 (red; c) or for p62 (green) and CK5 (red; d). Nuclei counterstained with DAPI (blue). e, Immunoblot analysis of MDA-MB-231 cells for the indicated proteins (left), 48 h following treatment with CTRL, NEDD4 or ITCH shRNAs. Samples were run on separate gels with equal loading of total proteins; loading control (actin) corresponds to ITCH blot. f, Model for breast cancer cells with normal autophagy (top) versus cells with autophagy inhibition (bottom). Normal autophagic degradation of ACRs reduces steady state levels of p63 due to ITCH-mediated UPS turnover. Inhibiting autophagy results in accumulation of NBR1–p62/SQSTM1 complexes, which sequester ITCH away from p63. As a result, stabilized p63 promotes the increased expression of basal cytokeratins and increased metastasis. Each dot (a,b) represents one animal. Statistical significance (a,b) was determined by one-way ANOVA, followed by Dunnett’s multiple comparisons test. Data are represented by mean ± S.E.M. Data and unprocessed blots are provided.
