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. 2017 Nov 14;7:15528. doi: 10.1038/s41598-017-14764-4

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© The Author(s) 2017

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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YAP -promoted LRP1 was depended on transcription in the A375 cells. (a,b) Western blots of LRP1 in melanoma A375 cells infected with GFP-sh or LRP1-sh1or LRP1-sh2 (a); relative LRP1 protein levels were shown as the ratio between LRP1 and GAPDH, and protein levels of the A375 cells infected with GFP-sh was arbitrarily set to 100% (b). (c,d) Western blots of YAP in melanoma A375 cells a transfected with GFP-sh or YAP-FLAG (c); relative LRP1 protein levels were shown as the ratio between YAP and GAPDH, and protein levels of melanoma A375 cells infected with GFP-sh were arbitrarily set to 100% (d). (e,f) Western blots of LRP1 in melanoma A375 cells transfected with GFP-sh or LRP1-FLAG (e); relative LRP1 protein levels were shown as the ratio between LRP1 and GAPDH, and protein levels of the A375 cells infected with GFP-sh were arbitrarily set to 100% (f). (g,h) Western blots of LRP1 in melanoma A375 cells infected with GFP-sh or YAP-sh1or YAP-sh2 (g); relative LRP1 protein levels were shown as the ratio between LRP1 and GAPDH, and protein levels of the A375 cells infected with GFP-sh were arbitrarily set to 100% (h). (i) The LRP1 protein in melanoma A375 cells transfected with empty or YAP-FLAG plasmids was detected using Western blot after adding CHX (50 μg/ml) for indicated time (upper panel). And the LRP1 protein was normalized to that of GAPDH. The 0 h time point was arbitrarily set to 100% (lower panel). (j) Time-dependent Caspase 3/7 activities after CHX (50 μg/ml) of LRP1 in the indicated A375 cells. The blue line was represented the control group; The red line was represented the YAP-FLAG group. (k) Time-dependent Caspase 3/7 activities after CHX (50 μg/ml) of YAP in the indicated A375 cells. The blue line was represented the control group; The red line was represented the YAP-FLAG group. (l) Relative mRNA of LRP1 in melanoma A375 cells infected with GFP-sh or YAP-sh1 or YAP-sh2.(m,n) Immunofluorescence assays using anti-YAP and anti-LRP1 antibodies (lower panel). Scale bar, 10 μM. (o,p) Knockdown of YAP inhibited the activity of LRP1 promoter and over-expressed YAP stimulated the activity of LRP1 promoter, LRP1 promoter containing −2000 bp to +150 bp luciferase activities were measured in melanoma A375 cells with control which were transfecting PGL4/Basic. Data were shown as mean ± SD from three independent experiments (including WB). *P < 0.05; **P < 0.01; ***P < 0.001 versus control measured by the student t test.