title: Combining integrated genomics and functional genomics to dissect the biology of a cancer-associated, aberrant transcription factor, the ASPSCR1-TFE3 fusion oncoprotein.
abstract: Oncogenic rearrangements of the TFE3 transcription factor gene are found in two distinct human cancers. These include ASPSCR1-TFE3 in all cases of alveolar soft part sarcoma (ASPS) and ASPSCR1-TFE3, PRCC-TFE3, SFPQ-TFE3 and others in a subset of paediatric and adult RCCs. Here we examined the functional properties of the ASPSCR1-TFE3 fusion oncoprotein, defined its target promoters on a genome-wide basis and performed a high-throughput RNA interference screen to identify which of its transcriptional targets contribute to cancer cell proliferation. We first confirmed that ASPSCR1-TFE3 has a predominantly nuclear localization and functions as a stronger transactivator than native TFE3. Genome-wide location analysis performed on the FU-UR-1 cell line, which expresses endogenous ASPSCR1-TFE3, identified 2193 genes bound by ASPSCR1-TFE3. Integration of these data with expression profiles of ASPS tumour samples and inducible cell lines expressing ASPSCR1-TFE3 defined a subset of 332 genes as putative up-regulated direct targets of ASPSCR1-TFE3, including MET (a previously known target gene) and 64 genes as down-regulated targets of ASPSCR1-TFE3. As validation of this approach to identify genuine ASPSCR1-TFE3 target genes, two up-regulated genes bound by ASPSCR1-TFE3, CYP17A1 and UPP1, were shown by multiple lines of evidence to be direct, endogenous targets of transactivation by ASPSCR1-TFE3. As the results indicated that ASPSCR1-TFE3 functions predominantly as a strong transcriptional activator, we hypothesized that a subset of its up-regulated direct targets mediate its oncogenic properties. We therefore chose 130 of these up-regulated direct target genes to study in high-throughput RNAi screens, using FU-UR-1 cells. In addition to MET, we provide evidence that 11 other ASPSCR1-TFE3 target genes contribute to the growth of ASPSCR1-TFE3-positive cells. Our data suggest new therapeutic possibilities for cancers driven by TFE3 fusions. More generally, this work establishes a combined integrated genomics/functional genomics strategy to dissect the biology of oncogenic, chimeric transcription factors.
authors.affiliation: Department of Pediatrics, Memorial Sloan-Kettering Cancer Center, New York, USA.
authors.forename: Rachel
authors.initials: R
authors.lastname: Kobos
authors.affiliation: Department of Pathology, Memorial Sloan-Kettering Cancer Center, New York, USA.
authors.forename: Makoto
authors.initials: M
authors.lastname: Nagai
authors.affiliation: Department of Pathology, Memorial Sloan-Kettering Cancer Center, New York, USA.
authors.forename: Masumi
authors.initials: M
authors.lastname: Tsuda
authors.affiliation: Department of Pathology, Memorial Sloan-Kettering Cancer Center, New York, USA.
authors.forename: Man Yee
authors.initials: MY
authors.lastname: Merl
authors.affiliation: Department of Pathology, Memorial Sloan-Kettering Cancer Center, New York, USA.
authors.forename: Tsuyoshi
authors.initials: T
authors.lastname: Saito
authors.affiliation: Department of Pathology, Memorial Sloan-Kettering Cancer Center, New York, USA.
authors.forename: Marick
authors.initials: M
authors.lastname: Laé
authors.affiliation: Department of Epidemiology and Biostatistics, Memorial Sloan-Kettering Cancer Center, New York, USA.
authors.forename: Qianxing
authors.initials: Q
authors.lastname: Mo
authors.affiliation: Department of Epidemiology and Biostatistics, Memorial Sloan-Kettering Cancer Center, New York, USA.
authors.forename: Adam
authors.initials: A
authors.lastname: Olshen
authors.affiliation: Computational Biology Program, Sloan-Kettering Institute, New York, USA.
authors.forename: Steven
authors.initials: S
authors.lastname: Lianoglou
authors.affiliation: Computational Biology Program, Sloan-Kettering Institute, New York, USA.
authors.forename: Christina
authors.initials: C
authors.lastname: Leslie
authors.affiliation: Department of Epidemiology and Biostatistics, Memorial Sloan-Kettering Cancer Center, New York, USA.
authors.forename: Irina
authors.initials: I
authors.lastname: Ostrovnaya
authors.affiliation: High-throughput Screening Core Facility, Memorial Sloan-Kettering Cancer Center, New York, USA.
authors.forename: Christophe
authors.initials: C
authors.lastname: Antczak
authors.affiliation: High-throughput Screening Core Facility, Memorial Sloan-Kettering Cancer Center, New York, USA.
authors.forename: Hakim
authors.initials: H
authors.lastname: Djaballah
authors.affiliation: Department of Pathology and Human Oncology and Pathogenesis Program, Memorial Sloan-Kettering Cancer Center, New York, USA.
authors.forename: Marc
authors.initials: M
authors.lastname: Ladanyi
mesh_terms: D000818:Animals; D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors / Q000235:genetics* / Q000378:metabolism; D019556:COS Cells; D002467:Cell Nucleus / Q000378:metabolism; D049109:Cell Proliferation; D002470:Cell Survival; D002522:Chlorocebus aethiops; D047369:Chromatin Immunoprecipitation; D016000:Cluster Analysis; D020869:Gene Expression Profiling; D015972:Gene Expression Regulation, Neoplastic; D050939:Gene Fusion; D017930:Genes, Reporter; D023281:Genomics / Q000379:methods; D005838:Genotype; D057809:HEK293 Cells; D006367:HeLa Cells; D057166:High-Throughput Screening Assays; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D061986:MCF-7 Cells; D020411:Oligonucleotide Array Sequence Analysis; D015514:Oncogene Proteins, Fusion / Q000235:genetics* / Q000378:metabolism; D010641:Phenotype; D011401:Promoter Regions, Genetic; D034622:RNA Interference; D015203:Reproducibility of Results; D018234:Sarcoma, Alveolar Soft Part / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D015533:Transcriptional Activation; D014162:Transfection
chemical_list: C422743:ASPSCR1 protein, human; D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; D047908:Intracellular Signaling Peptides and Proteins; D015514:Oncogene Proteins, Fusion; C069203:TFE3 protein, human