Destabilization of Heterologous Proteins Mediated by the GSK3β Phosphorylation Domain of the β-Catenin Protein
Background and Aims: Wnt/β-catenin signaling plays important roles in development and cellular processes. The hallmark of canonical Wnt signaling activation is the stabilization of β-catenin protein in cytoplasm and/or nucleus. The stability of β-catenin is the key to its biological functions and is...
| Main Authors: | , , , , , , , , , , , , , , , , , , , |
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| Format: | Article |
| Language: | English |
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Cell Physiol Biochem Press GmbH & Co KG
2013-11-01
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| Series: | Cellular Physiology and Biochemistry |
| Subjects: | |
| Online Access: | http://www.karger.com/Article/FullText/354518 |
| _version_ | 1819195235622715392 |
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| author | Yuhan Kong Hongyu Zhang Xian Chen Wenwen Zhang Chen Zhao Ning Wang Ningning Wu Yunfeng He Guoxin Nan Hongmei Zhang Sheng Wen Fang Deng Zhan Liao Di Wu Junhui Zhang Xinyue Qin Rex C. Haydon Hue H. Luu Tong-Chuan He Lan Zhou |
| author_facet | Yuhan Kong Hongyu Zhang Xian Chen Wenwen Zhang Chen Zhao Ning Wang Ningning Wu Yunfeng He Guoxin Nan Hongmei Zhang Sheng Wen Fang Deng Zhan Liao Di Wu Junhui Zhang Xinyue Qin Rex C. Haydon Hue H. Luu Tong-Chuan He Lan Zhou |
| author_sort | Yuhan Kong |
| collection | DOAJ |
| description | Background and Aims: Wnt/β-catenin signaling plays important roles in development and cellular processes. The hallmark of canonical Wnt signaling activation is the stabilization of β-catenin protein in cytoplasm and/or nucleus. The stability of β-catenin is the key to its biological functions and is controlled by the phosphorylation of its amino-terminal degradation domain. Aberrant activation of β-catenin signaling has been implicated in the development of human cancers. It has been recently suggested that GSK3βmay play an essential role in regulating global protein turnover. Here, we investigate if the GSK3β phosphorylation site-containing degradation domain of β-catenin is sufficient to destabilize heterologous proteins. Methods and Results: We engineer chimeric proteins by fusing β-catenin degradation domain at the N- and/or C-termini of the enhanced green fluorescent protein (eGFP). In both transient and stable expression experiments, the chimeric GFP proteins exhibit a significantly decreased stability, which can be effectively antagonized by lithium and Wnt1. An activating mutation in the destruction domain significantly stabilizes the fusion protein. Furthermore, GSK3 inhibitor SB-216763 effectively increases the GFP signal of the fusion protein. Conversely, the inhibition of Wnt signaling with tankyrase inhibitor XAV939 results in a decrease in GFP signal of the fusion proteins, while these small molecules have no significant effects on the mutant destruction domain-GFP fusion protein. Conclusion: Our findings strongly suggest that the β-catenin degradation domain may be sufficient to destabilize heterologous proteins in Wnt signaling-dependent manner. It is conceivable that the chimeric GFP proteins may be used as a functional reporter to measure the dynamic status of β-catenin signaling, and to identify potential anticancer drugs that target β-catenin signaling. |
| first_indexed | 2024-12-23T02:09:32Z |
| format | Article |
| id | doaj.art-114fbfa7bb1845b198c44d983dec6897 |
| institution | Directory Open Access Journal |
| issn | 1015-8987 1421-9778 |
| language | English |
| last_indexed | 2024-12-23T02:09:32Z |
| publishDate | 2013-11-01 |
| publisher | Cell Physiol Biochem Press GmbH & Co KG |
| record_format | Article |
| series | Cellular Physiology and Biochemistry |
| spelling | doaj.art-114fbfa7bb1845b198c44d983dec68972022-12-21T18:03:48ZengCell Physiol Biochem Press GmbH & Co KGCellular Physiology and Biochemistry1015-89871421-97782013-11-013251187119910.1159/000354518354518Destabilization of Heterologous Proteins Mediated by the GSK3β Phosphorylation Domain of the β-Catenin ProteinYuhan KongHongyu ZhangXian ChenWenwen ZhangChen ZhaoNing WangNingning WuYunfeng HeGuoxin NanHongmei ZhangSheng WenFang DengZhan LiaoDi WuJunhui ZhangXinyue QinRex C. HaydonHue H. LuuTong-Chuan HeLan ZhouBackground and Aims: Wnt/β-catenin signaling plays important roles in development and cellular processes. The hallmark of canonical Wnt signaling activation is the stabilization of β-catenin protein in cytoplasm and/or nucleus. The stability of β-catenin is the key to its biological functions and is controlled by the phosphorylation of its amino-terminal degradation domain. Aberrant activation of β-catenin signaling has been implicated in the development of human cancers. It has been recently suggested that GSK3βmay play an essential role in regulating global protein turnover. Here, we investigate if the GSK3β phosphorylation site-containing degradation domain of β-catenin is sufficient to destabilize heterologous proteins. Methods and Results: We engineer chimeric proteins by fusing β-catenin degradation domain at the N- and/or C-termini of the enhanced green fluorescent protein (eGFP). In both transient and stable expression experiments, the chimeric GFP proteins exhibit a significantly decreased stability, which can be effectively antagonized by lithium and Wnt1. An activating mutation in the destruction domain significantly stabilizes the fusion protein. Furthermore, GSK3 inhibitor SB-216763 effectively increases the GFP signal of the fusion protein. Conversely, the inhibition of Wnt signaling with tankyrase inhibitor XAV939 results in a decrease in GFP signal of the fusion proteins, while these small molecules have no significant effects on the mutant destruction domain-GFP fusion protein. Conclusion: Our findings strongly suggest that the β-catenin degradation domain may be sufficient to destabilize heterologous proteins in Wnt signaling-dependent manner. It is conceivable that the chimeric GFP proteins may be used as a functional reporter to measure the dynamic status of β-catenin signaling, and to identify potential anticancer drugs that target β-catenin signaling.http://www.karger.com/Article/FullText/354518β-cateninWnt signalProtein degradationGreen fluorescent proteinDestabilized GFPChimeric proteinsHuman cancer |
| spellingShingle | Yuhan Kong Hongyu Zhang Xian Chen Wenwen Zhang Chen Zhao Ning Wang Ningning Wu Yunfeng He Guoxin Nan Hongmei Zhang Sheng Wen Fang Deng Zhan Liao Di Wu Junhui Zhang Xinyue Qin Rex C. Haydon Hue H. Luu Tong-Chuan He Lan Zhou Destabilization of Heterologous Proteins Mediated by the GSK3β Phosphorylation Domain of the β-Catenin Protein Cellular Physiology and Biochemistry β-catenin Wnt signal Protein degradation Green fluorescent protein Destabilized GFP Chimeric proteins Human cancer |
| title | Destabilization of Heterologous Proteins Mediated by the GSK3β Phosphorylation Domain of the β-Catenin Protein |
| title_full | Destabilization of Heterologous Proteins Mediated by the GSK3β Phosphorylation Domain of the β-Catenin Protein |
| title_fullStr | Destabilization of Heterologous Proteins Mediated by the GSK3β Phosphorylation Domain of the β-Catenin Protein |
| title_full_unstemmed | Destabilization of Heterologous Proteins Mediated by the GSK3β Phosphorylation Domain of the β-Catenin Protein |
| title_short | Destabilization of Heterologous Proteins Mediated by the GSK3β Phosphorylation Domain of the β-Catenin Protein |
| title_sort | destabilization of heterologous proteins mediated by the gsk3β phosphorylation domain of the β catenin protein |
| topic | β-catenin Wnt signal Protein degradation Green fluorescent protein Destabilized GFP Chimeric proteins Human cancer |
| url | http://www.karger.com/Article/FullText/354518 |
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