Genomic and biochemical insights into the specificity of ETS transcription factors - PubMed (original) (raw)
Review
Genomic and biochemical insights into the specificity of ETS transcription factors
Peter C Hollenhorst et al. Annu Rev Biochem. 2011.
Abstract
ETS proteins are a group of evolutionarily related, DNA-binding transcriptional factors. These proteins direct gene expression in diverse normal and disease states by binding to specific promoters and enhancers and facilitating assembly of other components of the transcriptional machinery. The highly conserved DNA-binding ETS domain defines the family and is responsible for specific recognition of a common sequence motif, 5'-GGA(A/T)-3'. Attaining specificity for biological regulation in such a family is thus a conundrum. We present the current knowledge of routes to functional diversity and DNA binding specificity, including divergent properties of the conserved ETS and PNT domains, the involvement of flanking structured and unstructured regions appended to these dynamic domains, posttranslational modifications, and protein partnerships with other DNA-binding proteins and coregulators. The review emphasizes recent advances from biochemical and biophysical approaches, as well as insights from genomic studies that detect ETS-factor occupancy in living cells.
Figures
Figure 1
Structural and functional domains of the ETS family of transcription factors. (a) Nomenclature and domain organization of the 28 paralogous human ETS proteins (grouped according to the dendogram of Figure 2). We use HUGO nomenclature (191) for all ETS genes and proteins, with alternative names provided. Oncogenic ETS proteins, which are created by human chromosome rearrangements, are illustrated by two examples. Many ETS genes produce multiple protein products via alternative splicing/start sites, and in these cases, a single polypeptide was chosen arbitrarily. Boxes identify the DNA-binding ETS domain (red), PNT domain (green), OST domain (blue), and B-box (magenta) of the ETS factors as well as the C-terminal portion of RUNX1 (light brown) and the N-terminal portion of EWS (dark brown). The circled P symbolizes a phosphorylated region discussed in the text. Not identified are additional regions involved in transcriptional activation/repression, posttranslational modifications (including kinase docking and sumoylation), nuclear import/export, turnover, and so forth. (b) Ribbon diagram of the ETS1 ETS domain bound to a DNA duplex (1k79.pdb with the inhibitory helices deleted for clarity). The side chains of R391, R394, and Y395, which provide base-specific contacts to core GGAA, are also shown in stick format. Ribbon diagrams of (c) the ERG PNT domain (1sxe.pdb) and (d) the GABPA OST domain (1juo.pdb). See Supplemental Tables 2 and 3 for a referenced summary of all published ETS and PNT domain structures. Molecular diagrams were rendered with PyMOL (
) with coordinate files from the Protein Data Bank (
).
Figure 2
Diversity in function of ETS family. (left) A dendogram shows the degree of relatedness of 28 human ETS domain sequences, built by ClustalW (192) using the neighbor-joining method (193). The length of each horizontal line indicates estimated evolutionary distance. Branches that separate an individual subfamily are labeled in red. Classes defined by differences in the in vitro derived binding site (62) are indicated by background colors. (right) A list of selected tissues or functions that are defective in each mouse gene deletion, as cataloged at
. See also citations 159 and 197–234, which are linked to defects in mouse deletion in figure. For the sake of brevity, some phenotypes are not listed. N/A indicates the gene deletion has not been reported. The asterisk indicates that this phenotype results from a point mutation of ERG that affects transactivation ability rather than a gene deletion (194). Mice with deletions or mutations of two members of a subfamily have phenotypes that indicate overlapping roles in endothelial cells (ETS1 and ETS2) (30), hematopoietic stem cells (ERG and FLI1) (28), thymocyte development (ELK1 and ELK4) (33), B-cell receptor signal transduction (SPI1 and SPIB) (195), and limb-bud development (ETV1 and ETV5) (196).
Figure 3
Appended structures to the conserved ETS and PNT domains expand regulatory potential. (a) The core ETS domain (red) contains three α-helices (rectangles) on a four-stranded, antiparallel β-sheet (arrows). Appended N- and C-terminal helices (cyan), as well as an ordered C-terminal coil region in ELF3 (not shown) and dynamic sequences such as the serine-rich region (SRR) of ETS1, provide additional functional diversity. Shown (right) is a ribbon diagram of residues 301–441 of ETS1 (1r36.pdb), with the appended helices (cyan) packed against helix H1 of the core ETS domain. Helices HI-1, HI-2, H4, and H5 inhibit DNA binding of ETS1. Helix HI-1, which is distal from the recognition helix H3, unfolds upon DNA binding as part of the autoinhibition mechanism (Figures 4_c,d_ and 5_a_). (b) The core PNT domain (green) consists of four α-helices and a small α- or 310-helix (H2′) with a fold similar to that of the SAM domain. Although not shown for clarity, residues preceding H2 form a conserved helical-like turn. Again, appended N-terminal (H0, H1) helices (orange) provide additional routes to functional diversity. Shown (right) is the ribbon diagram of residues 29–138 of ETS1 (2jv3.pdb), including the PNT domain, which constitute a docking site for both MAP kinase and the TAZ1 domain of an ETS1 cofactor, CBP (Figure 5_b_). Two conserved phosphoacceptors (asterisk) preceding the dynamic helix H0 of ETS1 are indicated. See Supplemental Tables 2 and 3 for a referenced summary of all published ETS and PNT domain structures. The codes shown are from the Protein Data Bank (
).
Figure 4
Structurally characterized ETS protein partnerships. Shown are molecular models of the ETS partnerships involving the ETS domains of (a) GABPA with GABPB1 (1awc.pdb), (b) ELK4 with serum response factor (SRF) (Ihbx.pdb), (c) ETS1 with PAX5 (1 mdm.pdb), (d) the ETS1 homodimer (2nny.pdb), and (e) SPI1 with IRF4 (46). In each case, the ribbon diagrams are colored as ETS domain (red), appended helices (cyan), B-box (magenta), and partner (yellow). (f) Three monomer units of an ETV6 PNT domain polymer associated head-to-tail (greens; 1ji7.pdb). See Supplemental Tables 2 and 3 for a referenced summary of all published ETS and PNT domain structures. The codes listed are from the Protein Data Bank (
).
Figure 5
Dynamic properties of the ETS and PNT domains regulate ETS1 function. (a) ETS1 autoinhibition involves a conformational equilibrium between a rigid inactive state and a flexible active state. Upon DNA binding, helix HI-1 unfolds. Transient phosphorylation-dependent interactions of the unstructured serine-rich region (SRR) with the regulatable unit, formed by the core ETS domain (red) and the appended inhibitory module (cyan), stabilize the inactive state (80). This stabilization increases progressively with increasing levels of CaM kinase II multisite phosphorylation of the SRR, leading to rheostatic control of ETS1 DNA binding (79). (b) The PNT domain of ETS1 interacts with the TAZ1 domain of CBP. Phosphorylation of ETS1 causes a conformational change and increases the affinity of the interaction (97). The ETS1 PNT domain consists of a core helical bundle (green) with appended helices H0 and H1 (orange). MAP kinase phosphorylation of Thr38 and Ser41 shifts a conformational equilibrium of the dynamic helix H0 toward a more open state. This conformational switch, along with increased complementary electrostatic interactions, favors binding of the TAZ1 domain (yellow). Note that the structure of the PNT-TAZ1 complex has not been determined, and though not shown in panel a or b, both the unmodified and phosphorylated forms of ETS1 can bind CBP and DNA, respectively.
Figure 6
Genomic analyses via ChIP-chip and ChIP-Seq detect specific and redundant ETS binding. (a) Redundantly occupied regions in cells have a DNA sequence similar to preferred ETS-binding sequences derived in vitro. Sequence frequencies are compiled (top) from all 27 in vitro–derived ETS-binding sites (62) or (bottom) from the most overrepresented sequence in regions co-occupied in cells by GABPA, ETS1, and ELF1 (62, 65, 136). Frequencies are shown in logo form, which depicts base pair (A:T, T:A, G:C, C:G) frequency as relative heights of A, T, G, and C, respectively. (b) Redundant occupancy is observed for even the most divergent family members and between species. The relative distribution of specific and overlapping bound regions are shown for GABPA (136) and ELF1 (62) in human Jurkat T cells and SPI1 in mouse macrophages (138) by numbers of bound regions within a Venn diagram display. Analysis of SPI1 mouse data was limited to promoter regions, which could be compared with orthologous human regions using the LiftOver tool at
(235). Data were reprocessed for this comparison, as previously reported (65). (c,d) Serum response factor (SRF) and RUNX preferentially co-occupy regions bound specifically by ELK1 and ETS1, respectively. Diagrams (c,d) illustrate data from Boros et al. (134) and Hollenhorst et al. (65), respectively. See Supplemental Table 4 for database sources of all genomics data.
Figure 7
Partnerships provide a route to specificity in normal and disease states. Models depict ETS protein interactions with other DNA-binding proteins at the promoters or enhancers of distinct target classes. ETS proteins exhibit redundant occupancy of a consensus ETS-binding site in housekeeping promoters (132, 134). In contrast, ETS proteins bind specifically near genes that agree with predicted biological functions in either normal or cancer cells (, –134, 139). Specific interactions are associated with activation (green arrows) or repression (red bar) of target genes. Specific targets are highlighted by co-occupancy of multiple transcription factors [RUNX, serum response factor (SRF), androgen receptor (AR)] implicating partnerships as drivers of specificity. EWS-FLI1 tandem occupancy is inferred from functional and biochemical analyses of binding to GGAA microsatellite repeats (133, 137, 187). See Supplemental Table 4 for database sources of all genomics data.
References
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