Figure 8.
The WW-NLS regulates YAP1-dependent cancer cell stemness. (A) Representative immunostaining images show the subcellular localization of YAP1 in MDA-MB-231 cells following 1-wk culture with 1 μM of synthesized PPPY peptide, and the scatter chart shows YAP1 nucleocytoplasmic ratios (representative of three independent experiments, n represents the total number of the measured cells, two-tailed ANOVA test). (B) The expression of CYR61 in MDA-MB-231 and AsPC1 cells following 1-wk culture with 1 μM of PPPY peptide (representative of three independent experiments, N = 3, all data shown as the average values ± SD, two-tailed Student’s t test). The representative western blotting shows YAP1 expression in both cells. YAP1nuc, YAP1 nuclear localization; YAP1cyto, YAP1 cytoplasmic localization. (C and D) The expressions of OCT4 (C) and NANOG (D) in MDA-MB-231 and AsPC1 cells following 1-wk culture with 1 μM of PPPY peptide (representative of three independent experiments, N = 3, all data shown as the average values ± SD, two-tailed Student’s t test). (E) Ectopic expression of CPY6 peptide suppresses YAP1-activated OCT4 gene promoter activity in HEK293T cells (representative of three independent experiments, N = 3, all data shown as the average values ± SD, two-tailed Student’s t test). EV, empty vector. (F) Depletion of YAP1 suppresses the expressions of OCT4 and NANOG in MDA-MB-231 cells (representative of three independent experiments, N = 3, all data shown as the average values ± SD, two-tailed Student’s t test). Scr, scramble. (G) Representative images show spheroids of MDA-MB-231 cells following 1-wk culture with 1 μM of synthesized PPPY peptide, and the column chart shows diameters of spheroids (representative of three independent experiments, n = 8 spheroids, all data shown as the average values ± SD, two-tailed ANOVA test). (H) Representative images show spheroids of MDA-MB-231 cells following YAP1 knockdown, and the column chart shows diameters of spheroids (representative of three independent experiments, n = 8 spheroids, all data shown as the average values ± SD, two-tailed ANOVA test). (I) The schematic diagram illustrates that the WW domain regulates YAP1 nuclear import as an NLS, bypassing the hitchhiking mechanism. Source data are available for this figure: SourceData F8.

The WW-NLS regulates YAP1-dependent cancer cell stemness. (A) Representative immunostaining images show the subcellular localization of YAP1 in MDA-MB-231 cells following 1-wk culture with 1 μM of synthesized PPPY peptide, and the scatter chart shows YAP1 nucleocytoplasmic ratios (representative of three independent experiments, n represents the total number of the measured cells, two-tailed ANOVA test). (B) The expression of CYR61 in MDA-MB-231 and AsPC1 cells following 1-wk culture with 1 μM of PPPY peptide (representative of three independent experiments, N = 3, all data shown as the average values ± SD, two-tailed Student’s t test). The representative western blotting shows YAP1 expression in both cells. YAP1nuc, YAP1 nuclear localization; YAP1cyto, YAP1 cytoplasmic localization. (C and D) The expressions of OCT4 (C) and NANOG (D) in MDA-MB-231 and AsPC1 cells following 1-wk culture with 1 μM of PPPY peptide (representative of three independent experiments, N = 3, all data shown as the average values ± SD, two-tailed Student’s t test). (E) Ectopic expression of CPY6 peptide suppresses YAP1-activated OCT4 gene promoter activity in HEK293T cells (representative of three independent experiments, N = 3, all data shown as the average values ± SD, two-tailed Student’s t test). EV, empty vector. (F) Depletion of YAP1 suppresses the expressions of OCT4 and NANOG in MDA-MB-231 cells (representative of three independent experiments, N = 3, all data shown as the average values ± SD, two-tailed Student’s t test). Scr, scramble. (G) Representative images show spheroids of MDA-MB-231 cells following 1-wk culture with 1 μM of synthesized PPPY peptide, and the column chart shows diameters of spheroids (representative of three independent experiments, n = 8 spheroids, all data shown as the average values ± SD, two-tailed ANOVA test). (H) Representative images show spheroids of MDA-MB-231 cells following YAP1 knockdown, and the column chart shows diameters of spheroids (representative of three independent experiments, n = 8 spheroids, all data shown as the average values ± SD, two-tailed ANOVA test). (I) The schematic diagram illustrates that the WW domain regulates YAP1 nuclear import as an NLS, bypassing the hitchhiking mechanism. Source data are available for this figure: SourceData F8.

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