Yongwen Luo1,2, Jun Zhou3, Jianing Tang2, Fengfang Zhou1, Zhiwen He1, Tongzu Liu4, Tao Liu5. 1. Department of Urology, Zhongnan Hospital of Wuhan University, Wuhan, China. 2. Department of Biological Repositories, Zhongnan Hospital of Wuhan University, Wuhan, China. 3. The Interventional Diagnosis and Treatment Center, Zhongnan Hospital of Wuhan University, Wuhan, China. 4. Department of Urology, Zhongnan Hospital of Wuhan University, Wuhan, China. liutongzu@163.com. 5. Department of Urology, Zhongnan Hospital of Wuhan University, Wuhan, China. liutaozaiwuda@whu.edu.cn.
Abstract
BACKGROUND: Bladder cancer is one of the most commonly diagnosed urological malignant tumor. The Hippo tumor suppressor pathway is highly conserved in mammals and plays an important role in carcinogenesis. YAP is one of major key effectors of the Hippo pathway. However, the mechanism supporting abnormal YAP expression in bladder cancer remains to be characterized. METHODS: Western blot was used to measure the expression of MINDY1 and YAP, while the YAP target genes were measured by real-time PCR. CCK8 assay was used to detect the cell viability. The xeno-graft tumor model was used for in vivo study. Protein stability assay was used to detect YAP protein degradation. Immuno-precipitation assay was used to detect the interaction domain between MINDY1 and YAP. The ubiquitin-based Immuno-precipitation assays were used to detect the specific ubiquitination manner happened on YAP. RESULTS: In the present study, we identified MINDY1, a DUB enzyme in the motif interacting with ubiquitin-containing novel DUB family, as a bona fide deubiquitylase of YAP in bladder cancer. MINDY1 was shown to interact with, deubiquitylate, and stabilize YAP in a deubiquitylation activity-dependent manner. MINDY1 depletion significantly decreased bladder cancer cell proliferation. The effects induced by MINDY1 depletion could be rescued by further YAP overexpression. Depletion of MINDY1 decreased the YAP protein level and the expression of YAP/TEAD target genes in bladder cancer, including CTGF, ANKRD1 and CYR61. CONCLUSION: In general, our findings establish a previously undocumented catalytic role for MINDY1 as a deubiquitinating enzyme of YAP and provides a possible target for the therapy of bladder cancer.
BACKGROUND:Bladder cancer is one of the most commonly diagnosed urological malignant tumor. The Hippotumor suppressor pathway is highly conserved in mammals and plays an important role in carcinogenesis. YAP is one of major key effectors of the Hippo pathway. However, the mechanism supporting abnormal YAP expression in bladder cancer remains to be characterized. METHODS: Western blot was used to measure the expression of MINDY1 and YAP, while the YAP target genes were measured by real-time PCR. CCK8 assay was used to detect the cell viability. The xeno-graft tumor model was used for in vivo study. Protein stability assay was used to detect YAP protein degradation. Immuno-precipitation assay was used to detect the interaction domain between MINDY1 and YAP. The ubiquitin-based Immuno-precipitation assays were used to detect the specific ubiquitination manner happened on YAP. RESULTS: In the present study, we identified MINDY1, a DUB enzyme in the motif interacting with ubiquitin-containing novel DUB family, as a bona fide deubiquitylase of YAP in bladder cancer. MINDY1 was shown to interact with, deubiquitylate, and stabilize YAP in a deubiquitylation activity-dependent manner. MINDY1 depletion significantly decreased bladder cancer cell proliferation. The effects induced by MINDY1 depletion could be rescued by further YAP overexpression. Depletion of MINDY1 decreased the YAP protein level and the expression of YAP/TEAD target genes in bladder cancer, including CTGF, ANKRD1 and CYR61. CONCLUSION: In general, our findings establish a previously undocumented catalytic role for MINDY1 as a deubiquitinating enzyme of YAP and provides a possible target for the therapy of bladder cancer.
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