Abstract
Colon cancer cells frequently lose expression of the tumor suppressor adenomatous polyposis coli (APC). As result, beta-catenin accumulates and activates transcription of Tcf-responsive genes. Here we describe a novel mammalian two-hybrid system that selectively kills APC-mutated cells. This system consists of GAL4/beta-catenin, VP16/Tcf4, and a gene that is transcribed when GAL4 and VP16 associate. In APC-mutated human colon cancer cells, such as SW480, GAL4/beta-catenin accumulates, and in the presence of VP16/Tcf4, induces high levels of expression of the reporter gene. Expression of wild-type APC reduced GAL4/beta-catenin and intact beta-catenin levels and inhibited reporter gene expression. In colon cancer cells such as SW48 that have wild-type APC, GAL4/beta-catenin was degraded, and expression levels of the output gene were low. Replacement of the reporter gene with a suicide gene resulted in selective killing of SW480 cells. This system may be applicable for broader use of gene therapy by targeting diseases that involve protein degradation.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Adenomatous Polyposis Coli Protein
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Colonic Neoplasms / genetics*
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Colonic Neoplasms / metabolism
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Colonic Neoplasms / therapy
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Cytoskeletal Proteins / biosynthesis
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Cytoskeletal Proteins / genetics*
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Cytoskeletal Proteins / metabolism
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DNA-Binding Proteins
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Fungal Proteins / genetics
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Fungal Proteins / metabolism
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Genes, APC / genetics*
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Genetic Therapy / methods*
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Herpes Simplex Virus Protein Vmw65 / genetics
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Herpes Simplex Virus Protein Vmw65 / metabolism
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Humans
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Kidney / cytology
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Kidney / metabolism
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Kidney / physiology
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Mutation
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Osteosarcoma / genetics
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Osteosarcoma / metabolism
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Plasmids
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Recombinant Fusion Proteins / biosynthesis
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Recombinant Fusion Proteins / genetics*
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Recombinant Fusion Proteins / metabolism
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Saccharomyces cerevisiae Proteins*
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TCF Transcription Factors
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Trans-Activators*
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Transcription Factor 7-Like 2 Protein
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Transcription Factors / biosynthesis
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Transcription Factors / genetics*
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Transcription Factors / metabolism
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Transfection
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Tumor Cells, Cultured
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Tumor Suppressor Protein p53 / biosynthesis
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beta Catenin
Substances
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Adenomatous Polyposis Coli Protein
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CTNNB1 protein, human
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Cytoskeletal Proteins
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DNA-Binding Proteins
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Fungal Proteins
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GAL4 protein, S cerevisiae
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Herpes Simplex Virus Protein Vmw65
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Recombinant Fusion Proteins
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Saccharomyces cerevisiae Proteins
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TCF Transcription Factors
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TCF7L2 protein, human
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Trans-Activators
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Transcription Factor 7-Like 2 Protein
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Transcription Factors
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Tumor Suppressor Protein p53
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beta Catenin