title: SDHDPromoter Mutations Ablate GABP Transcription Factor Binding in Melanoma.
abstract: SDHDencodes subunit D of the succinate dehydrogenase complex, an integral membrane protein. Across cancer types, recurrentSDHDpromoter mutations were reported to occur exclusively in melanomas, at a frequency of 4% to 5%. These mutations are predicted to disrupt consensus ETS transcription factor-binding sites and are correlated with both reducedSDHDgene expression and poor prognosis. However, the consequence of these mutations onSDHDexpression in melanoma is still unclear. Here, we found that expression ofSDHDin melanoma correlated with the expression of multiple ETS transcription factors, particularly inSDHDpromoter wild-type samples. Consistent with the predicted loss of ETS transcription factor binding, we observed that recurrent hotspot mutations resulted in decreased luciferase activity in reporter assays. Furthermore, we demonstrated specific GABPA and GABPB1 binding to probes containing the wild-type promoter sequences, with binding disrupted by theSDHDhotspot promoter mutations in both quantitative mass spectrometry and band-shift experiments. Finally, using siRNA-mediated knockdown across multiple melanoma cell lines, we determined that loss of GABPA resulted in reduced SDHD expression at both RNA and protein levels. These data are consistent with a key role for GABPA/B1 as the critical ETS transcription factors deregulatingSDHDexpression in the context of highly recurrent promoter mutations in melanoma and warrant a detailed search for other recurrent promoter mutations that create or disrupt GABPA consensus sequences.Cancer Res; 77(7); 1649-61. ©2017 AACR.
authors.affiliation: Laboratory of Translational Genomics, Division of Cancer Epidemiology and Genetics, NCI, NIH, Bethesda, Maryland.
authors.forename: Tongwu
authors.initials: T
authors.lastname: Zhang
authors.affiliation: Laboratory of Translational Genomics, Division of Cancer Epidemiology and Genetics, NCI, NIH, Bethesda, Maryland.
authors.forename: Mai
authors.initials: M
authors.lastname: Xu
authors.affiliation: Radboud Institute of Molecular Life Sciences, Radboud University Nijmegen, Nijmegen, the Netherlands.
authors.forename: Matthew M
authors.initials: MM
authors.lastname: Makowski
authors.affiliation: Laboratory of Translational Genomics, Division of Cancer Epidemiology and Genetics, NCI, NIH, Bethesda, Maryland.
authors.forename: Christine
authors.initials: C
authors.lastname: Lee
authors.affiliation: Laboratory of Translational Genomics, Division of Cancer Epidemiology and Genetics, NCI, NIH, Bethesda, Maryland.
authors.forename: Michael
authors.initials: M
authors.lastname: Kovacs
authors.affiliation: Laboratory of Translational Genomics, Division of Cancer Epidemiology and Genetics, NCI, NIH, Bethesda, Maryland.
authors.forename: Jun
authors.initials: J
authors.lastname: Fang
authors.affiliation: Radboud Institute of Molecular Life Sciences, Radboud University Nijmegen, Nijmegen, the Netherlands.
authors.forename: Esther
authors.initials: E
authors.lastname: Willems
authors.affiliation: The Translational Genomics Research Institute (TGen), Phoenix, Arizona.
authors.forename: Jeffrey M
authors.initials: JM
authors.lastname: Trent
authors.affiliation: QIMR Berghofer Medical Research Institute, Brisbane, Queensland, Australia.
authors.forename: Nicholas K
authors.initials: NK
authors.lastname: Hayward
authors.affiliation: Radboud Institute of Molecular Life Sciences, Radboud University Nijmegen, Nijmegen, the Netherlands. Kevin.Brown3@nih.gov M.Vermeulen@ncmls.ru.nl.
authors.forename: Michiel
authors.initials: M
authors.lastname: Vermeulen
authors.affiliation: Laboratory of Translational Genomics, Division of Cancer Epidemiology and Genetics, NCI, NIH, Bethesda, Maryland. Kevin.Brown3@nih.gov M.Vermeulen@ncmls.ru.nl.
authors.forename: Kevin M
authors.initials: KM
authors.lastname: Brown
mesh_terms: D045744:Cell Line, Tumor; D051268:GA-Binding Protein Transcription Factor / Q000378:metabolism*; D006801:Humans; D008545:Melanoma / Q000235:genetics* / Q000378:metabolism; D009154:Mutation; D011401:Promoter Regions, Genetic; D050783:Proto-Oncogene Proteins c-ets / Q000378:metabolism; D013385:Succinate Dehydrogenase / Q000235:genetics*; D053719:Tandem Mass Spectrometry
chemical_list: D051268:GA-Binding Protein Transcription Factor; C495661:GABPA protein, human; C495662:GABPB1 protein, human; D050783:Proto-Oncogene Proteins c-ets; C491545:SDHD protein, human; D013385:Succinate Dehydrogenase