title: Systematic Screening of Promoter Regions Pinpoints Functional Cis-Regulatory Mutations in a Cutaneous Melanoma Genome.
abstract: With the recent discovery of recurrent mutations in the TERT promoter in melanoma, identification of other somatic causal promoter mutations is of considerable interest. Yet, the impact of sequence variation on the regulatory potential of gene promoters has not been systematically evaluated. This study assesses the impact of promoter mutations on promoter activity in the whole-genome sequenced malignant melanoma cell line COLO-829. Combining somatic mutation calls from COLO-829 with genome-wide chromatin accessibility and histone modification data revealed mutations within promoter elements. Interestingly, a high number of potential promoter mutations (n = 23) were found, a result mirrored in subsequent analysis of TCGA whole-melanoma genomes. The impact of wild-type and mutant promoter sequences were evaluated by subcloning into luciferase reporter vectors and testing their transcriptional activity in COLO-829 cells. Of the 23 promoter regions tested, four mutations significantly altered reporter activity relative to wild-type sequences. These data were then subjected to multiple computational algorithms that score the cis-regulatory altering potential of mutations. These analyses identified one mutation, located within the promoter region of NDUFB9, which encodes the mitochondrial NADH dehydrogenase (ubiquinone) 1 beta subcomplex 9, to be recurrent in 4.4% (19 of 432) of TCGA whole-melanoma exomes. The mutation is predicted to disrupt a highly conserved SP1/KLF transcription factor binding motif and its frequent co-occurrence with mutations in the coding sequence of NF1 supports a pathologic role for this mutation in melanoma. Taken together, these data show the relatively high prevalence of promoter mutations in the COLO-829 melanoma genome, and indicate that a proportion of these significantly alter the regulatory potential of gene promoters.
  
  
  CONCLUSIONS
  Genomic-based screening within gene promoter regions suggests that functional cis-regulatory mutations may be common in melanoma genomes, highlighting the need to examine their role in tumorigenesis.
authors.affiliation: Prince of Wales Clinical School, University of New South Wales Australia, Sydney, Australia. Lowy Cancer Research Centre, University of New South Wales Australia, Sydney, Australia.
authors.forename: Rebecca C
authors.initials: RC
authors.lastname: Poulos
authors.affiliation: Prince of Wales Clinical School, University of New South Wales Australia, Sydney, Australia. Lowy Cancer Research Centre, University of New South Wales Australia, Sydney, Australia.
authors.forename: Julie A I
authors.initials: JA
authors.lastname: Thoms
authors.affiliation: Prince of Wales Clinical School, University of New South Wales Australia, Sydney, Australia. Lowy Cancer Research Centre, University of New South Wales Australia, Sydney, Australia.
authors.forename: Anushi
authors.initials: A
authors.lastname: Shah
authors.affiliation: Prince of Wales Clinical School, University of New South Wales Australia, Sydney, Australia. Lowy Cancer Research Centre, University of New South Wales Australia, Sydney, Australia.
authors.forename: Dominik
authors.initials: D
authors.lastname: Beck
authors.affiliation: Prince of Wales Clinical School, University of New South Wales Australia, Sydney, Australia. Lowy Cancer Research Centre, University of New South Wales Australia, Sydney, Australia. Department of Haematology, Prince of Wales Hospital, Sydney, Australia.
authors.forename: John E
authors.initials: JE
authors.lastname: Pimanda
authors.affiliation: Prince of Wales Clinical School, University of New South Wales Australia, Sydney, Australia. Lowy Cancer Research Centre, University of New South Wales Australia, Sydney, Australia. jason.wong@unsw.edu.au.
authors.forename: Jason W H
authors.initials: JW
authors.lastname: Wong
mesh_terms: D045744:Cell Line, Tumor; D002843:Chromatin / Q000378:metabolism; D017930:Genes, Reporter; D005821:Genetic Techniques; D015894:Genome, Human; D006801:Humans; D008545:Melanoma / Q000235:genetics*; D008957:Models, Genetic; D009154:Mutation; D009245:NADH Dehydrogenase / Q000235:genetics; D011401:Promoter Regions, Genetic; D050436:Regulatory Elements, Transcriptional; D012878:Skin Neoplasms / Q000235:genetics*; D016329:Sp1 Transcription Factor / Q000378:metabolism; D014158:Transcription, Genetic; D000096142:Cutaneous Malignant Melanoma
chemical_list: D002843:Chromatin; D016329:Sp1 Transcription Factor; D009245:NADH Dehydrogenase; C118906:NDUFB9 protein, human