ETS class I (ETS1/ERG/FLI1)
ETS proteins are a family of 28 human transcription factors that each bind DNA through a single ~85-residue winged helix-turn-helix domain and read a nine-base site whose informative part is the four-base core GGA(A/T). The class I members - ETS1 and ETS2, ERG, FLI1, FEV, the PEA3 proteins ETV1/4/5, the ternary complex factors ELK1/3/4, the repressors ERF and ETV3, and GABPA - select essentially the same sequence, ACCGGAAGT, so a match names the class rather than a protein. These sites sit just upstream of a large fraction of human promoters and, in enhancers, next to RUNX, GATA, PAX and SRF sites. Class I also contains every ETS gene recurrently rearranged in cancer: ERG in prostate cancer, FLI1 and ERG in Ewing sarcoma, ETV1 and ETV4 in both.
The factors
The human genome contains 28 ETS genes. All encode one copy of the ETS domain: about 85 residues folded as a winged helix-turn-helix, three alpha-helices packed on a four-stranded antiparallel beta-sheet, that binds DNA as a monomer 8602247,21548782. Profiling the DNA-binding preference of every human and mouse ETS protein split the family into four classes, and class I is by far the largest: the ETS subfamily (ETS1, ETS2), the ERG subfamily (ERG, FLI1, FEV), the PEA3 proteins (ETV1, ETV4, ETV5), the ternary complex factors (ELK1, ELK3, ELK4), the repressors ERF and ETV3, and GABPA 20517297. GABPA is the structural exception: it functions only in complex with the ankyrin-repeat subunit GABPB, which it recruits using the ETS domain plus a carboxy-terminal extension 9461436. Several class I proteins carry autoinhibitory segments flanking the ETS domain that are relieved by partner binding 10075931,21548782. Expression ranges from near-ubiquitous (GABPA, ELK1, ERF, ETV3) through lymphoid-enriched (ETS1), endothelial and hematopoietic (ERG, FLI1), to narrowly restricted (FEV). SPDEF is grouped here in this atlas, but biochemically it is the sole class IV factor and prefers a GGAT core 20517297,21548782.
The motif
The site is nine base pairs, ACCGGAAGT, and nearly all of its information is in the GGA(A/T) core 21548782. Two invariant arginines in recognition helix H3 hydrogen-bond the two core guanines; a conserved tyrosine (Tyr395 in ETS1) supports the interface; a lysine in the wing contacts DNA 5' of the core and a lysine in the H2-H3 turn contacts the opposite strand 3' of it 8602247,21548782. The core is read base by base, the flanks indirectly, through sequence-dependent backbone geometry 21548782. The flanks separate subfamilies, and separate them weakly: class I prefers A three bases 5' of the core and disfavors C there, the ELF/EHF group prefers C at that position (CCCGGAAGT), SPI1/SPIB/SPIC use a purine-rich 5' flank (AGAGGAAGTG), and SPDEF alone prefers GGAT 20517297. Quantitatively, different ETS proteins vary only about ten-fold in affinity for the same consensus, whereas one protein's affinity across variant sites spans five- to eighty-fold 21548782; sequence therefore grades site quality much better than it identifies a paralogue, and no class I member is separable from another by motif alone. The CCGG contains a CpG, and methylating it blocks binding 20019798,21548782. Tandem arrangements are a distinct case: GGAA microsatellites of four or more consecutive repeats are the response elements of the EWS-FLI1 fusion 18626011,21548782, and some models grouped here (JASPAR MA0149.1, archetype AC0561) are GGAA repeats rather than single sites.
Where it acts
ETS occupancy follows two logics. At promoters it is largely redundant: a strong CCGGAAGT sits 20-40 bp upstream of the transcription start site in CpG-island housekeeping promoters - roughly a quarter of human promoters carry an ETS site near the start site 17652178,21548782 - and different ETS proteins bind interchangeably, with most GABPA-occupied promoters in Jurkat cells also occupied by ETS1 17652178,20019798. These sites lie in hypomethylated CpG islands, so the CpG in the core stays unmethylated 21548782. At distal, DNase-sensitive enhancers, occupancy is factor-specific, the sequence departs from consensus 17652178,20019798, and specificity comes from composite elements: ETS1 with RUNX1 at AGGA(A/T)GTGG, which binds more than 30-fold cooperatively and marks over a thousand ETS1-specific regions in T cells 10075931,20019798,21548782; ETS with GATA in hemogenic endothelium, where FLI1, GATA2 and SCL cross-regulate through neighboring sites 17962413; ETS1 with PAX5 at the mb-1 promoter, where Gln22 of the PAX5 paired domain hydrogen-bonds Tyr395 of ETS1 11779502,21548782; the ternary complex factors with SRF at serum response elements 11715049,21548782; and ERG with the androgen receptor in prostate cancer, where the two share about 44% of sites 21548782. The ETS-IRF composite element (EICE) belongs to the SPI1 subfamily and is described there.
Biology and disease
Class I factors run housekeeping transcription, growth-factor responses and lineage programs. Gabpa-null mouse embryos die before implantation 15199140, consistent with GABPA's occupancy of housekeeping promoters. Ets1 loss impairs T-cell and natural killer cell development. ERG and FLI1 are required for endothelial and hematopoietic specification. The clinical weight of the class is oncogenic rearrangement: TMPRSS2-ERG and related fusions place ERG, ETV1, ETV4 or ETV5 under androgen control in roughly half of prostate cancers 16254181,21548782, and EWSR1-FLI1 or EWSR1-ERG defines Ewing sarcoma, where the fusion's prion-like EWSR1 domain converts GGAA microsatellites into de novo enhancers 18626011,28844694.
Reading a hit
Treat a single match as weak evidence. The four-base GGAA core is expected about 12 million times per strand in a 3.1-Gb genome and the full nine-mer ACCGGAAGT on the order of 10,000 times per strand by base-composition arithmetic; CCGG-containing words are rarer than that because the genome is CpG-depleted, but the site is still common enough that isolated hits mean little without context: conservation, accessibility, a nearby partner site, or a promoter position close to the start site. A hit identifies an ETS site, not an ETS protein: ETS1, ERG, FLI1, ELK1 and GABPA are not distinguishable from sequence. ChIP-derived models carry partner signal: HOCOMOCO's ChIP-based ETS1 model (ETS1.H14CORE.1.P.B) has consensus ACAGGAAGTGG, whose trailing GTGG is the RUNX1 half-site of the ETS1-RUNX1 composite 20019798, and several other ChIP-derived ETS models converge on CAGGAAG rather than the in vitro CCGGAAGT 20019798,21548782. Half of the JASPAR entries grouped here (16 of 32) are CAP-SELEX heterodimer models such as ERF::SREBF2 or ETV5::FIGLA, where much of the matrix is the partner's site, not an ETS site. Finally, consensus strings are stored on either strand: GABPA's JASPAR model reads CACTTCCTGT, the reverse complement of ACAGGAAGTG.
How this was worked out
The ETS domain is named as a new DNA-binding motif shared by the ETS proteins and shown to recognize a purine-rich core centred on GGA.
Still open then: How one short domain read so small a core, and whether family members differed in what they bound, was unknown.
The first ETS domain-DNA crystal structure (PU.1) shows a winged helix-turn-helix contacting ten base pairs through a loop-helix-loop: two arginines of the recognition helix read the GGA core, a wing lysine binds 5' of it and a turn lysine 3' of it on the opposite strand.
Still open then: No base contacts outside the core were seen, so the flanking preferences that separate subfamilies were left to indirect readout.
Outlier-expression analysis finds ERG and ETV1 overexpressed in prostate cancer through fusion to the 5' end of the androgen-regulated TMPRSS2 gene; 23 of 29 tumors examined by FISH carry an ERG or ETV1 rearrangement.
Still open then: Which genomic sites the overexpressed ETS proteins act through, and whether they differ from those of the endogenous ETS factors, was not addressed.
Promoter-array ChIP for three ETS proteins in a T-cell line finds that redundant occupancy is the rule: consensus sites near the start sites of housekeeping genes are shared, while factor-specific sites diverge from consensus, lie farther away, and include an ETS1-RUNX1 cooperative site.
Still open then: Promoter arrays could not see enhancers, where specific occupancy turned out to concentrate.
EWS-FLI1 regulates targets such as NR0B1 through GGAA microsatellites; the NR0B1 repeat binds the fusion in vitro and in vivo, is necessary and sufficient for regulation, and activity depends on the number of consecutive GGAA motifs.
Still open then: Whether repeat binding was a property of the FLI1 ETS domain or required the EWS portion of the fusion was not settled.
ETS1 ChIP-seq in Jurkat T cells separates redundant occupancy with GABPA at housekeeping promoters from ETS1-specific occupancy at T-cell enhancers, which carry either a CpG-free CAGGAAGT variant or an ETS-RUNX composite; more than a thousand specific regions show AGGA(A/T)GTGG.
Still open then: FLI1 and ERG also co-occupy sites with RUNX1 in stem cells, so how specific the composite is for ETS1 was left open.
DNA-binding profiles of all human and mouse ETS proteins, from two in vitro methods, fall into four classes; every cancer-linked ETS factor (ERG, ETV1, ETV4, FLI1) is in class I, and ChIP-seq for one member of each class confirms that the small in vitro differences affect site choice in cells.
Still open then: Members within class I remained indistinguishable, so paralogue specificity had to come from elsewhere.
- 2017mediumChIP-seq in mesenchymal stem cells and Ewing sarcoma cells, mutagenesis, proteomics, in vitro phase-transition assays28844694
Wild-type FLI1 occupies canonical ETS sites but not GGAA repeats, whereas EWS-FLI1 binds repeats, recruits BAF complexes and opens de novo enhancers, and needs the tyrosines that drive phase transitions of the EWSR1 prion-like domain to bind them. This revises the earlier view that the EWS portion contributes only transcriptional activation.
Still unresolved
- How class I paralogues reach different genomic sites despite indistinguishable in vitro preferences is only partly explained: ETS-RUNX composites account for some ETS1-specific enhancers, but RUNX1 also co-occupies sites with FLI1 and ERG, and the contributions of partner choice, autoinhibition and relative expression level to the remaining specific sites have not been separated.
- Why ChIP-derived models converge on CAGGAAG while in vitro models give ACCGGAAGT is unresolved: one reading is an intrinsic ETS1 preference for the variant that confers specificity, the other is that the A simply removes a CpG whose methylation would block binding at enhancers outside protected CpG islands.
- Whether binding to GGAA microsatellites is an intrinsic property of the ETS domain or a neomorphic property of the EWSR1 fusion is contested: in vitro and reporter work found binding that scales with repeat number and reported that several wild-type ETS proteins bind repeats, while in cells wild-type FLI1 does not occupy repeats and EWS-FLI1 needs its prion-like-domain tyrosines to do so.
- The databases cluster SPDEF models with class I, whereas in vitro profiling makes SPDEF the sole class IV factor with a GGAT core; whether SPDEF also uses GGAA sites in cells, or the clustering reflects only the shared CCGGA flank, is not established.
A hit tells you the family, not the member: all class I ETS proteins select the same nine-mer in vitro and differ only in weak flanking preferences, so a "FLI1 site" and an "ERG site" are the same sequence. The core is short and frequent, so a match outside an accessible, conserved or composite context is expected by chance. Sixteen of the 32 JASPAR models here are CAP-SELEX heterodimers whose matrices are partly a bHLH, FOX, HOX or SREBF site; four models here are GGAA repeats rather than single sites and will hit any GGAA microsatellite; and the ChIP-derived HOCOMOCO models (evidence letter P) include RUNX-extended and CAGGAAG variants that reflect partners and indirect binding rather than intrinsic ETS specificity. SPDEF is placed in this family here but reads GGAT and is biochemically its own class.
References
- Hollenhorst PC, McIntosh LP, Graves BJ. Genomic and biochemical insights into the specificity of ETS transcription factors. Annu Rev Biochem 2011;80:437-71. PMID 21548782
- Wei GH, Badis G, Berger MF, et al. Genome-wide analysis of ETS-family DNA-binding in vitro and in vivo. EMBO J 2010;29:2147-60. PMID 20517297
- Sharrocks AD. The ETS-domain transcription factor family. Nat Rev Mol Cell Biol 2001;2:827-37. PMID 11715049
- Kodandapani R, Pio F, Ni CZ, et al. A new pattern for helix-turn-helix recognition revealed by the PU.1 ETS-domain-DNA complex. Nature 1996;380:456-60. PMID 8602247
- Batchelor AH, Piper DE, de la Brousse FC, et al. The structure of GABPalpha/beta: an ETS domain- ankyrin repeat heterodimer bound to DNA. Science 1998;279:1037-41. PMID 9461436
- Hollenhorst PC, Shah AA, Hopkins C, et al. Genome-wide analyses reveal properties of redundant and specific promoter occupancy within the ETS gene family. Genes Dev 2007;21:1882-94. PMID 17652178
- Hollenhorst PC, Chandler KJ, Poulsen RL, et al. DNA specificity determinants associate with distinct transcription factor functions. PLoS Genet 2009;5:e1000778. PMID 20019798
- Kim WY, Sieweke M, Ogawa E, et al. Mutual activation of Ets-1 and AML1 DNA binding by direct interaction of their autoinhibitory domains. EMBO J 1999;18:1609-20. PMID 10075931
- Garvie CW, Hagman J, Wolberger C. Structural studies of Ets-1/Pax5 complex formation on DNA. Mol Cell 2001;8:1267-76. PMID 11779502
- Pimanda JE, Ottersbach K, Knezevic K, et al. Gata2, Fli1, and Scl form a recursively wired gene-regulatory circuit during early hematopoietic development. Proc Natl Acad Sci U S A 2007;104:17692-7. PMID 17962413
- Tomlins SA, Rhodes DR, Perner S, et al. Recurrent fusion of TMPRSS2 and ETS transcription factor genes in prostate cancer. Science 2005;310:644-8. PMID 16254181
- Gangwal K, Sankar S, Hollenhorst PC, et al. Microsatellites as EWS/FLI response elements in Ewing's sarcoma. Proc Natl Acad Sci U S A 2008;105:10149-54. PMID 18626011
- Karim FD, Urness LD, Thummel CS, et al. The ETS-domain: a new DNA-binding motif that recognizes a purine-rich core DNA sequence. Genes Dev 1990;4:1451-3. PMID 2253872
- Ristevski S, O'Leary DA, Thornell AP, et al. The ETS transcription factor GABPalpha is essential for early embryogenesis. Mol Cell Biol 2004;24:5844-9. PMID 15199140
- Boulay G, Sandoval GJ, Riggi N, et al. Cancer-Specific Retargeting of BAF Complexes by a Prion-like Domain. Cell 2017;171:163-178.e19. PMID 28844694
81 motifs in this family — JASPAR 32 · HOCOMOCO 23 · CIS-BP 16 · Vierstra 10
| motif | database | consensus | bits |
|---|---|---|---|
| EWSR1-FLI1 (MA0149.1) | JASPAR | GGAAGGAAGGAAGGAAGG | 32.9 |
| M08111_2.00 | CIS-BP | GGAAGGAAGGAAGGAAGG | 32.9 |
| M04778_2.00 | CIS-BP | ACCGGAAGtaCTTCCGGT | 26.7 |
| M07950_2.00 | CIS-BP | GCcTkCTGGGAatTGTAGTtC | 25 |
| ERF::SREBF2 (MA1939.2)dimer | JASPAR | CGGAAATCACGTGAT | 20.8 |
| ERF::NHLH1 (MA1938.2)dimer | JASPAR | aGCAGCTGCCGGAAgT | 20.6 |
| AC0561:FLI/ZNF:Ets | Vierstra | TTCCTTCCtTCCTTCC | 19.9 |
| AC0079:ERF::NHLH:['Tryptophan_cluster_factors',_'Basic_helix-loop-helix_factors_(bHLH)'] | Vierstra | GCAGCTGCCGGAAg | 19.4 |
| AC0365:SPDEF:Ets | Vierstra | TaATCCGGGACCA | 19 |
| ELK1::HOXB13 (MA1932.2)dimer | JASPAR | ACCGGAAGThGTAAA | 18.9 |
| AC0643:ETS/ETV:Ets | Vierstra | cGGAAGTACTTCCg | 18.6 |
| AC0646:ERG/FLI:Ets | Vierstra | CCGGAtATCCGG | 18.4 |
| ELK1::SREBF2 (MA1933.2)dimer | JASPAR | CCGGAAGTggCGTGA | 18.3 |
| AC0105:SPDEF:Ets | Vierstra | CAGdAAGAAGTAaa | 18.1 |
| ERF::FIGLA (MA1934.2)dimer | JASPAR | CCGGAAaCAcCTG | 18 |
| ELK1::HOXA1 (MA1931.1)dimer | JASPAR | ACCGGAAGTAATT | 17.4 |
| ETV5::FIGLA (MA1945.2)dimer | JASPAR | cGGAAgCAGGTG | 17.4 |
| ETV4.H14CORE.1.SM.BSMB | HOCOMOCO | ACCGGAAGT | 15.7 |
| M04752_2.00 | CIS-BP | ACCGGAAGT | 15.2 |
| AC0076:ERF::FIGLA/ETV2::FIGLA:['Tryptophan_cluster_factors',_'Basic_helix-loop-helix_factors_(bHLH)'] | Vierstra | CGGAAgCAGgTG | 15.2 |
| AC0541:ETS/SIX:Ets | Vierstra | ACTACAacTCCCAG | 14.8 |
| SPDEF.H14CORE.1.SM.BSMB | HOCOMOCO | AaCCGGATGTdvnnhccg | 14.7 |
| M02973_2.00 | CIS-BP | AcCCGGATGT | 14.5 |
| FEV (MA0156.4) | JASPAR | ACCGGAAGT | 14.4 |
| SPDEF (MA0686.2) | JASPAR | AcCCGGATGT | 14.3 |
| GABPA.H14CORE.0.PSM.APSMA | HOCOMOCO | ACCGGAAGT | 14.3 |
| ELK1 (MA0028.3) | JASPAR | ACCGGAAGT | 14.2 |
| ELK1.H14CORE.0.PSM.APSMA | HOCOMOCO | ACCGGAAGT | 14.2 |
| ERG.H14CORE.1.SM.BSMB | HOCOMOCO | ACCGGAAGT | 14.2 |
| ETV5 (MA0765.4) | JASPAR | ACCGGAAGT | 14.1 |
| ETV5::HOXA2 (MA1948.2)dimer | JASPAR | CGGAaGTaATTA | 14.1 |
| FLI1::DRGX (MA1949.2)dimer | JASPAR | aCCGGAAGTAATTA | 14.1 |
| ELK3.H14CORE.0.PSM.APSMA | HOCOMOCO | aCCGGAAGT | 14.1 |
| ELK3 (MA0759.3) | JASPAR | ACCGGAAGT | 14 |
| ETV5::FOXI1 (MA1946.2)dimer | JASPAR | GTAAACAGGAaG | 14 |
| M02734_2.00 | CIS-BP | CGGAtaTCCG | 14 |
| AC0054:ELK/ETV:Ets | Vierstra | TCGTAAATGC | 13.9 |
| FEV.H14CORE.0.S.BSB | HOCOMOCO | aCCGGAAGT | 13.8 |
| ETS2.H14CORE.0.S.CSC | HOCOMOCO | aCCGGAAGT | 13.5 |
| ETS2 (MA1484.2) | JASPAR | ACCGGAAGt | 13.3 |
| FLI1 (MA0475.3) | JASPAR | ACCGGAAgT | 13.2 |
| ETV5::DRGX (MA1944.2)dimer | JASPAR | CGGAAGtAATTA | 13.2 |
| ETV1.H14CORE.1.PM.APMA | HOCOMOCO | gCAGGAAGTg | 13.2 |
| ETS1.H14CORE.1.P.BPB | HOCOMOCO | aCaGGAAGTGG | 13.1 |
| FLI1::FOXI1 (MA1950.2)dimer | JASPAR | tAAACAGGAAg | 13 |
| FLI1.H14CORE.1.P.BPB | HOCOMOCO | GgcAGGAAGbgg | 13 |
| M07952_2.00 | CIS-BP | CCGGAAGT | 13 |
| ERF::FOXO1 (MA1936.2)dimer | JASPAR | gtaAACAGGAAG | 12.9 |
| ERG.H14CORE.0.P.BPB | HOCOMOCO | aCCGGAAGTg | 12.9 |
| M02968_2.00 | CIS-BP | ACCGGAAGT | 12.9 |
| ELK4 (MA0076.3) | JASPAR | aCTTCCGG | 12.7 |
| ERF (MA0760.2) | JASPAR | ACCGGAAGT | 12.7 |
| FLI1.H14CORE.0.PSM.APSMA | HOCOMOCO | aCCGGAAGTg | 12.7 |
| M04730_2.00 | CIS-BP | aCCGGAAGT | 12.6 |
| ETV1.H14CORE.0.PSM.APSMA | HOCOMOCO | ACCGGAAGT | 12.5 |
| SPDEF.H14CORE.0.PSM.APSMA | HOCOMOCO | amCCGGATgt | 12.5 |
| M04791_2.00 | CIS-BP | aCCGGAAGT | 12.5 |
| ETS1.H14CORE.0.S.BSB | HOCOMOCO | ACCGGAAGT | 12.4 |
| M07957_2.00 | CIS-BP | CCGGAAGt | 12.2 |
| Erg (MA0474.4) | JASPAR | ACAGGAAGTg | 12.1 |
| ETV3.H14CORE.0.SM.BSMB | HOCOMOCO | ACCGGAAGT | 12.1 |
| ETS1 (MA0098.4) | JASPAR | ACCGGAAgT | 12 |
| ERF::HOXB13 (MA1937.2)dimer | JASPAR | CGGAAgtngTaAA | 11.9 |
| GABPA (MA0062.4) | JASPAR | CACTTCCTGT | 11.7 |
| ETV1 (MA0761.3) | JASPAR | ACAGGAAGT | 11.6 |
| ETV5::FOXO1 (MA1947.2)dimer | JASPAR | GtAAACAGGA | 11.6 |
| M02970_2.00 | CIS-BP | ACCGGAAGT | 11.6 |
| ETV3 (MA0763.2) | JASPAR | ACCGGAAGT | 11.5 |
| ETV4 (MA0764.4) | JASPAR | aCCGGAAGT | 11.5 |
| ELK4.H14CORE.0.PSM.APSMA | HOCOMOCO | CCGGAAGt | 11.4 |
| ETV5.H14CORE.0.PS.APSA | HOCOMOCO | CCGGAAGt | 11.4 |
| ETS2.H14CORE.1.P.BPB | HOCOMOCO | ccGGAAGtG | 11.2 |
| ETV4.H14CORE.0.P.BPB | HOCOMOCO | CaGGAAG | 11 |
| ETV5.H14CORE.1.PM.APMA | HOCOMOCO | CCGGAAGt | 10.9 |
| M03006_2.00 | CIS-BP | aCCGGAAGT | 10.6 |
| ERF.H14CORE.0.PS.APSA | HOCOMOCO | AGGAAgT | 9.8 |
| M04800_2.00 | CIS-BP | cCGGAaG | 9.6 |
| ERF::FOXI1 (MA1935.2)dimer | JASPAR | ACaGGAAg | 9.5 |
| M04708_2.00 | CIS-BP | CGGAaG | 8.8 |
| AC0641:ETV/ETS:Ets | Vierstra | GGaAGGAAG | 7.8 |
| M01477_2.00Mus musculus 1 | CIS-BP | GGAa | 5.9 |
HOCOMOCO tags read as assay + quality grade: P ChIP-seq, S HT-SELEX, M methyl-HT-SELEX, G GHT-SELEX, I SMiLE-seq, B PBM; grade A means two or more assay types agreed. A P-only matrix is a record of where the protein was crosslinked, which need not be a sequence it binds itself.
The factors, elsewhere
These entries are written from the primary literature and every citation is checked against PubMed, but this one can still be out of date, too confident, or simply wrong — especially where a motif’s published site and its database matrices disagree. Corrections and additions are welcome, and they are the fastest way to make this page better for everyone.