title: Validation of chemical compound library screening for transcriptional co-activator with PDZ-binding motif inhibitors using GFP-fused transcriptional co-activator with PDZ-binding motif.
abstract: Transcriptional co-activator with PDZ-binding motif (TAZ) plays versatile roles in cell proliferation and differentiation. It is phosphorylated by large tumor suppressor kinases, the core kinases of the tumor-suppressive Hippo pathway. Phosphorylation induces the cytoplasmic accumulation of TAZ and its degradation. In human cancers, the deregulation of the Hippo pathway and gene amplification enhance TAZ activity. TAZ interacts with TEA domain family members (TEAD), and upregulates genes implicated in epithelial-mesenchymal transition. It also confers stemness to cancer cells. Thus, TAZ activation provides cancer cells with malignant properties and worsens the clinical prognosis. Therefore, TAZ attracts attention as a therapeutic target in cancer therapy. We applied 18 606 small chemical compounds to human osteosarcoma U2OS cells expressing GFP-fused TAZ (GFP-TAZ), monitored the subcellular localization of GFP-TAZ, and selected 33 compounds that shifted GFP-TAZ to the cytoplasm. Unexpectedly, only a limited number of compounds suppressed TAZ-mediated enhancement of TEAD-responsive reporter activity. Moreover, the compounds that weakened TEAD reporter activity did not necessarily decrease the unphosphorylated TAZ. In this study, we focused on three compounds that decreased both TEAD reporter activity and unphosphorylated TAZ, and treated several human cancer cells with these compounds. One compound did not show a remarkable effect, whereas the other two compounds compromised the cell viability in certain cancer cells. In conclusion, the GFP-TAZ-based assay can be used as the first screening for compounds that inhibit TAZ and show anticancer properties. To develop anticancer drugs, we need additional assays to select the compounds.
authors.affiliation: Department of Medical Biochemistry, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Tokyo, Japan.
authors.forename: Shunta
authors.initials: S
authors.lastname: Nagashima
authors.affiliation: Department of Medical Biochemistry, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Tokyo, Japan.
authors.forename: Junichi
authors.initials: J
authors.lastname: Maruyama
authors.affiliation: Department of Medical Biochemistry, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Tokyo, Japan.
authors.forename: Shodai
authors.initials: S
authors.lastname: Kawano
authors.affiliation: Department of Medical Biochemistry, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Tokyo, Japan.
authors.forename: Hiroaki
authors.initials: H
authors.lastname: Iwasa
authors.affiliation: Department of Medical Biochemistry, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Tokyo, Japan.
authors.forename: Kentaro
authors.initials: K
authors.lastname: Nakagawa
authors.affiliation: Chemical Biology Screening Center, Tokyo Medical and Dental University, Tokyo, Japan.
authors.forename: Mari
authors.initials: M
authors.lastname: Ishigami-Yuasa
authors.affiliation: Chemical Biology Screening Center, Tokyo Medical and Dental University, Tokyo, Japan.|Institute of Biomaterials and Bioengineering, Tokyo Medical and Dental University, Tokyo, Japan.
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Kagechika
authors.affiliation: Department of Developmental and Regenerative Biology, Medical Research Institute, Tokyo Medical and Dental University, Tokyo, Japan.
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Nishina
authors.affiliation: Department of Medical Biochemistry, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Tokyo, Japan.|Center for Brain Integration Research, Tokyo Medical and Dental University, Tokyo, Japan.
authors.forename: Yutaka
authors.initials: Y
authors.lastname: Hata
mesh_terms: D020816:Amino Acid Motifs; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D002470:Cell Survival / Q000187:drug effects; D003593:Cytoplasm / Q000187:drug effects / Q000378:metabolism; D004280:Dobutamine / Q000494:pharmacology; D004353:Drug Evaluation, Preclinical / Q000379:methods / Q000592:standards*; D004983:Ethanolamines / Q000032:analysis / Q000494:pharmacology; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism*; D057809:HEK293 Cells; D006575:Heterocyclic Compounds, 3-Ring / Q000032:analysis / Q000494:pharmacology; D000090683:Hippo Signaling Pathway; D006801:Humans; D054731:PDZ Domains / Q000187:drug effects*; D010744:Phosphoric Monoester Hydrolases / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D011485:Protein Binding / Q000187:drug effects; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D011725:Pyridines / Q000032:analysis / Q000494:pharmacology; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D054852:Small Molecule Libraries / Q000032:analysis* / Q000494:pharmacology*; D013890:Thiourea / Q000031:analogs & derivatives / Q000032:analysis / Q000494:pharmacology; D013997:Time Factors; D014157:Transcription Factors / Q000037:antagonists & inhibitors* / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects*; D062367:ortho-Aminobenzoates / Q000032:analysis / Q000494:pharmacology
chemical_list: D004983:Ethanolamines; D006575:Heterocyclic Compounds, 3-Ring; C000613762:IBS000540; C000613761:IBS001594; C000613760:IBS015625; D011725:Pyridines; D011993:Recombinant Fusion Proteins; D054852:Small Molecule Libraries; D014157:Transcription Factors; D062367:ortho-Aminobenzoates; D049452:Green Fluorescent Proteins; D004280:Dobutamine; D017346:Protein Serine-Threonine Kinases; D010744:Phosphoric Monoester Hydrolases; D013890:Thiourea
keywords: Drug screening; GFP; Hippo pathway; TAZ; phosphorylation