title: CAUSEL: an epigenome- and genome-editing pipeline for establishing function of noncoding GWAS variants.
abstract: The vast majority of disease-associated single-nucleotide polymorphisms (SNPs) mapped by genome-wide association studies (GWASs) are located in the non-protein-coding genome, but establishing the functional and mechanistic roles of these sequence variants has proven challenging. Here we describe a general pipeline in which candidate functional SNPs are first evaluated by fine mapping, epigenomic profiling, and epigenome editing, and then interrogated for causal function by using genome editing to create isogenic cell lines followed by phenotypic characterization. To validate this approach, we analyzed the 6q22.1 prostate cancer risk locus and identified rs339331 as the top-scoring SNP. Epigenome editing confirmed that the rs339331 region possessed regulatory potential. By using transcription activator-like effector nuclease (TALEN)-mediated genome editing, we created a panel of isogenic 22Rv1 prostate cancer cell lines representing all three genotypes (TT, TC, CC) at rs339331. Introduction of the 'T' risk allele increased transcription of the regulatory factor 6 (RFX6) gene, increased homeobox B13 (HOXB13) binding at the rs339331 region, and increased deposition of the enhancer-associated H3K4me2 histone mark at the rs339331 region compared to lines homozygous for the 'C' protective allele. The cell lines also differed in cellular morphology and adhesion, and pathway analysis of differentially expressed genes suggested an influence of androgens. In summary, we have developed and validated a widely accessible approach that can be used to establish functional causality for noncoding sequence variants identified by GWASs.
authors.affiliation: Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, Massachusetts, USA.|Center for Functional Cancer Epigenetics, Dana-Farber Cancer Institute, Boston, Massachusetts, USA.
authors.forename: Sándor
authors.initials: S
authors.lastname: Spisák
authors.affiliation: Department of Preventive Medicine, Keck School of Medicine, University of Southern California, Los Angeles, California, USA.
authors.forename: Kate
authors.initials: K
authors.lastname: Lawrenson
authors.affiliation: Molecular Pathology Unit, Massachusetts General Hospital, Charlestown, Massachusetts, USA.|Center for Cancer Research, Massachusetts General Hospital, Charlestown, Massachusetts, USA.|Center for Computational and Integrative Biology, Massachusetts General Hospital, Charlestown, Massachusetts, USA.|Department of Pathology, Harvard Medical School, Boston, Massachusetts, USA.
authors.forename: Yanfang
authors.initials: Y
authors.lastname: Fu
authors.affiliation: Department of Physics of Complex Systems, Eötvös Loránd University, Budapest, Hungary.
authors.forename: István
authors.initials: I
authors.lastname: Csabai
authors.affiliation: Molecular Pathology Unit, Massachusetts General Hospital, Charlestown, Massachusetts, USA.|Center for Cancer Research, Massachusetts General Hospital, Charlestown, Massachusetts, USA.|Center for Computational and Integrative Biology, Massachusetts General Hospital, Charlestown, Massachusetts, USA.|Program in Biological and Biomedical Sciences, Harvard Medical School, Boston, Massachusetts, USA.
authors.forename: Rebecca T
authors.initials: RT
authors.lastname: Cottman
authors.affiliation: Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, Massachusetts, USA.|Center for Functional Cancer Epigenetics, Dana-Farber Cancer Institute, Boston, Massachusetts, USA.
authors.forename: Ji-Heui
authors.initials: JH
authors.lastname: Seo
authors.affiliation: Department of Preventive Medicine, Keck School of Medicine, University of Southern California, Los Angeles, California, USA.|Norris Comprehensive Cancer Center, University of Southern California, Los Angeles, California, USA.
authors.forename: Christopher
authors.initials: C
authors.lastname: Haiman
authors.affiliation: Department of Preventive Medicine, Keck School of Medicine, University of Southern California, Los Angeles, California, USA.
authors.forename: Ying
authors.initials: Y
authors.lastname: Han
authors.affiliation: Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, Massachusetts, USA.|Center for Functional Cancer Epigenetics, Dana-Farber Cancer Institute, Boston, Massachusetts, USA.
authors.forename: Romina
authors.initials: R
authors.lastname: Lenci
authors.affiliation: Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, Massachusetts, USA.|Center for Functional Cancer Epigenetics, Dana-Farber Cancer Institute, Boston, Massachusetts, USA.|Medical College, Xiamen University, Xiamen, China.
authors.forename: Qiyuan
authors.initials: Q
authors.lastname: Li
authors.affiliation: Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, Massachusetts, USA.|Computational Health Informatics Program (CHIP), Boston Children's Hospital, Boston, Massachusetts, USA.
authors.forename: Viktória
authors.initials: V
authors.lastname: Tisza
authors.affiliation: Computational Health Informatics Program (CHIP), Boston Children's Hospital, Boston, Massachusetts, USA.|Department of Systems Biology, Technical University of Denmark, Lyngby, Denmark.|Second Department of Pathology, Semmelweis University, Budapest, Hungary.
authors.forename: Zoltán
authors.initials: Z
authors.lastname: Szállási
authors.affiliation: Molecular Biology Core Facilities at Dana-Farber Cancer Institute, Boston, Massachusetts, USA.
authors.forename: Zachery T
authors.initials: ZT
authors.lastname: Herbert
authors.affiliation: Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, Massachusetts, USA.
authors.forename: Matthew
authors.initials: M
authors.lastname: Chabot
authors.affiliation: Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, Massachusetts, USA.
authors.forename: Mark
authors.initials: M
authors.lastname: Pomerantz
authors.affiliation: Department of Animal Hygiene, Szent István University, Budapest, Hungary.
authors.forename: Norbert
authors.identifier: http://orcid.org/0000-0003-1783-2041
authors.initials: N
authors.lastname: Solymosi
authors.affiliation: Department of Preventive Medicine, Keck School of Medicine, University of Southern California, Los Angeles, California, USA.|The Eli and Edythe L. Broad Institute, Cambridge, Massachusetts, USA.
authors.forename: Simon A
authors.initials: SA
authors.lastname: Gayther
authors.affiliation: Molecular Pathology Unit, Massachusetts General Hospital, Charlestown, Massachusetts, USA.|Center for Cancer Research, Massachusetts General Hospital, Charlestown, Massachusetts, USA.|Center for Computational and Integrative Biology, Massachusetts General Hospital, Charlestown, Massachusetts, USA.|Department of Pathology, Harvard Medical School, Boston, Massachusetts, USA.|Program in Biological and Biomedical Sciences, Harvard Medical School, Boston, Massachusetts, USA.
authors.forename: J Keith
authors.initials: JK
authors.lastname: Joung
authors.affiliation: Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, Massachusetts, USA.|Center for Functional Cancer Epigenetics, Dana-Farber Cancer Institute, Boston, Massachusetts, USA.|The Eli and Edythe L. Broad Institute, Cambridge, Massachusetts, USA.
authors.forename: Matthew L
authors.initials: ML
authors.lastname: Freedman
mesh_terms: D000483:Alleles; D045744:Cell Line, Tumor; D002874:Chromosome Mapping; D002896:Chromosomes, Human, Pair 6 / Q000235:genetics*; D004268:DNA-Binding Proteins / Q000235:genetics; D057890:Epigenomics; D015972:Gene Expression Regulation, Neoplastic / Q000235:genetics*; D020022:Genetic Predisposition to Disease; D055106:Genome-Wide Association Study; D042421:Histone Code; D006657:Histones / Q000378:metabolism; D018398:Homeodomain Proteins / Q000235:genetics; D006801:Humans; D008297:Male; D020641:Polymorphism, Single Nucleotide; D011471:Prostatic Neoplasms / Q000235:genetics*; D012333:RNA, Messenger / Q000378:metabolism*; D000071701:Regulatory Factor X Transcription Factors; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014157:Transcription Factors / Q000235:genetics*
chemical_list: D004268:DNA-Binding Proteins; C484414:HOXB13 protein, human; D006657:Histones; D018398:Homeodomain Proteins; D012333:RNA, Messenger; D000071701:Regulatory Factor X Transcription Factors; C547964:Rfx6 protein, human; D014157:Transcription Factors