title: Emergent Properties of EWS/FLI Regulation via GGAA Microsatellites in Ewing's Sarcoma.
abstract: ETS proteins are a family of transcription factors that play important roles in the development of cancer. The Ewing's sarcoma EWS/ETS fusion oncoproteins control a number of cancer-relevant phenotypes in that disease. We recently demonstrated that EWS/FLI, the most common EWS/ETS fusion in Ewing's sarcoma, regulates a portion of its target genes, including the critical target NR0B1, via GGAA-containing microsatellites in their promoters. Given the unusual nature of microsatellites as EWS/FLI response elements, we sought to elucidate the mechanism of EWS/FLI activity at these sites. We found that the ability to bind GGAA microsatellites is shared by multiple ETS family members from distinct phylogenetic subfamilies. Importantly, however, only EWS/ETS-containing fusions are capable of mediating transcriptional activation via these elements, highlighting a neomorphic function of the Ewing's sarcoma fusion proteins. Additional analysis revealed that the GGAA microsatellite binds EWS/FLI with an affinity that is 2 to 3 orders of magnitude lower than previously identified high-affinity consensus/redundant binding sites. The stoichiometry of this interaction is 2 protein molecules for each DNA molecule, suggesting that EWS/FLI binds these elements as a homodimer. The isolated FLI ETS domain bound microsatellite sequences in a nearly identical fashion to full-length EWS/FLI, thus indicating that residues required for homodimeric binding are localized to the ETS domain. These data suggest a new paradigm for an ETS family member binding to DNA at cancer-relevant genetic loci and highlight emergent properties of EWS/FLI that are required for the development of Ewing's sarcoma.
authors.affiliation: Department of Oncological Sciences, University of Utah School of Medicine, and Center for Children's Cancer Research, Huntsman Cancer Institute, Salt Lake City, Utah.
authors.forename: Kunal
authors.initials: K
authors.lastname: Gangwal
authors.forename: Devin
authors.initials: D
authors.lastname: Close
authors.forename: Camille A
authors.initials: CA
authors.lastname: Enriquez
authors.forename: Christopher P
authors.initials: CP
authors.lastname: Hill
authors.forename: Stephen L
authors.initials: SL
authors.lastname: Lessnick
references.citation: Proc Natl Acad Sci U S A. 1993 Jun 15;90(12):5752-6
references.pmid: 8516324
references.citation: Proc Natl Acad Sci U S A. 2008 Jul 22;105(29):10149-54
references.pmid: 18626011
references.citation: Mol Cell Biol. 1994 May;14(5):3230-41
references.pmid: 8164678
references.citation: Nature. 1992 Sep 10;359(6391):162-5
references.pmid: 1522903
references.citation: Cancer Res. 2006 Oct 15;66(20):9937-47
references.pmid: 17047056
references.citation: J Biol Chem. 2000 Sep 15;275(37):28363-70
references.pmid: 10867009
references.citation: Nature. 1996 Nov 28;384(6607):322
references.pmid: 8934514
references.citation: Oncogene. 1993 Aug;8(8):2167-73
references.pmid: 8336942
references.citation: Oncogene. 2009 Nov 19;28(46):4126-32
references.pmid: 19718047
references.citation: Methods Cell Biol. 2008;84:243-62
references.pmid: 17964934
references.citation: Genes Dev. 1989 May;3(5):606-11
references.pmid: 2744457
references.citation: Cancer Cell. 2002 May;1(4):393-401
references.pmid: 12086853
references.citation: Cell Cycle. 2006 Dec;5(23):2753-9
references.pmid: 17172842
references.citation: J Bacteriol. 2005 Jul;187(14):5008-12
references.pmid: 15995218
references.citation: Oncogene. 1997 Mar 13;14(10):1159-64
references.pmid: 9121764
references.citation: Cancer Genet Cytogenet. 1988 Jun;32(2):229-38
references.pmid: 3163261
references.citation: Nat Genet. 1994 Feb;6(2):146-51
references.pmid: 8162068
references.citation: PLoS One. 2009;4(3):e4932
references.pmid: 19305498
references.citation: Science. 2005 Jul 1;309(5731):142-5
references.pmid: 15994560
references.citation: Oncogene. 1995 Mar 16;10(6):1229-34
references.pmid: 7700648
references.citation: Oncogene. 2003 Oct 9;22(44):6819-29
references.pmid: 14534527
references.citation: Nat Rev Mol Cell Biol. 2001 Nov;2(11):827-37
references.pmid: 11715049
references.citation: J Biol Chem. 1996 Sep 20;271(38):23329-37
references.pmid: 8798534
references.citation: Biochem Biophys Res Commun. 1996 Feb 15;219(2):608-12
references.pmid: 8605035
references.citation: Mol Cancer Res. 2006 Nov;4(11):851-9
references.pmid: 17114343
references.citation: Cancer Res. 2009 Dec 1;69(23):9047-55
references.pmid: 19920188
references.citation: Genes Dev. 1992 Jun;6(6):975-90
references.pmid: 1592264
references.citation: Genes Chromosomes Cancer. 1996 Feb;15(2):115-21
references.pmid: 8834175
references.citation: Mol Cell Biol. 1993 Dec;13(12):7393-8
references.pmid: 8246959
references.citation: Mol Cell Biol. 2000 Aug;20(15):5643-52
references.pmid: 10891501
references.citation: EMBO J. 2008 Jul 23;27(14):2006-17
references.pmid: 18566588
references.citation: J Mol Biol. 2008 Oct 17;382(4):1014-30
references.pmid: 18692067
references.citation: Brief Funct Genomic Proteomic. 2006 Mar;5(1):8-14
references.pmid: 16769671
references.citation: Oncogene. 2008 Oct 9;27(46):6034-43
references.pmid: 18591936
references.citation: Cancer Cell. 2006 May;9(5):405-16
references.pmid: 16697960
references.citation: Oncogene. 1999 Feb 11;18(6):1351-9
references.pmid: 10022817
references.citation: Genes Dev. 2007 Aug 1;21(15):1882-94
references.pmid: 17652178
references.citation: Cell Cycle. 2008 Oct;7(20):3127-32
references.pmid: 18927503
references.citation: Cell Res. 2007 Apr;17(4):286-8
references.pmid: 17426699
references.citation: Cell Cycle. 2006 Sep;5(18):2049-53
references.pmid: 16969112
references.citation: Eur J Cancer. 2005 Nov;41(16):2462-78
references.pmid: 16213704
references.citation: J Biol Chem. 1994 Jul 8;269(27):18216-22
references.pmid: 7517940
references.citation: PLoS Genet. 2009 Dec;5(12):e1000778
references.pmid: 20019798
references.citation: Mol Cell. 2001 Dec;8(6):1267-76
references.pmid: 11779502
references.citation: Oncogene. 1995 Feb 2;10(3):423-31
references.pmid: 7845667