Fig. 2: Formation of acini structures is perturbed in autophagy-deficient primary mammary epithelial cells.

a (left) Representative bright-field images and BrdU immunostaining in primary mammary epithelial cells (pMECs) isolated from wild-type and Atg16L1 hypomorph (Atg16hyp) mice. Scale bars are 200 µm. (right) Percentage of BrdU-positive pMECs isolated from wild-type (n = 2) and Atg16hyp (n = 3) mice. Bars represent the mean ± s.d. (*P < 0.5; two-way ANOVA). b Size of pMECs isolated from wild-type (n = 3) and Atg16hyp (n = 4) mice. Bars represent the mean ± s.d. (**P < 0.01; two-tailed t-test). c Representative confocal images of MCF10A cells plated on either soft or stiff ECM. MCF10A cells were previously exposed to control, ATG16L1 or ATG7/10 siRNAs. Scale bars are 200 µm. The bars represent the percentage of organoid/ spherical structures in soft (n = 3 independent experiments) or stiff (n = 5 independent experiments) ECM: mean ± s.d. (****P < 0.0001; two-tailed t-test). d Representative images of wild-type (n = 3 independent experiments) and Atg16hyp (n = 5 independent experiments) pMECs plated on stiff ECM. Scale bars are 400 µm. The bars represent the percentage of branched structures: mean ± s.d. (****P < 0.0001; two-tailed t-test). e Diameter of soft acini in MCF10A cells exposed to control, ATG16L1 or ATG7/10 siRNAs. Bars represent the mean ± s.d. (n = 3 independent experiments; **P < 0.01; two-tailed t-test). f Diameter of soft acini in wild-type or Atg16hyp pMECs. Bars represent the mean ± s.e.m. (n = 33 (wild-type), n = 27 (Atg16hyp) soft acini; ***P < 0.001; two-tailed t-test). The experiment was repeated with similar results. g Representative images of wild-type and Atg16hyp pMECs plated on soft ECM. Scale bars are 400 µm. The experiment was repeated twice with similar results. Exact P values for asterisks: a 0.0163; b 0.0050; e (from left to right) 0.0017, 0.0018; and f 0.0001.