Ets-1 transcription factor
The Ets-1 transcription factor regulates various genes, and is involved in stem cell development and tumorigenesis.
Domain organization/sequence features
Ets-1–DNA binding is regulated by an autoinhibitory region and requires a HI-1 helix to unfold. Interactions with DNA are further
attenuated by a serine-arginine rich region (SRR). Ets-1 autoinhibitory module is composed of four a helices (HI-1, HI-2, H4,
and H5) that flank the DNA- binding ETS domain. In the native protein, these helices pack cooperatively on a surface of the
ETS domain that is opposite to the DNA binding interface, reducing the affinity of Ets-1 for DNA 10-fold, as compared with the
affinity of the minimal ETS domain.
NMR data shows helix HI-1 is labile, even in the absence of DNA, and could serve as a control point for modulating DNA
The SRR is disordered in both the free and complex form. Truncation of the SRR region gradually increases the mobility of the
interface and facilitates HI-1 unfolding; a process that is required for DNA binding. Phosphorylation has minimal impact on the
secondary structure of the SRR region. Upon phosphorylation she most pronounced differences are in the dynamic properties
of the HI-1 autoinhibitory helix and recognition helices H1 and H3. These units form a hydrophobic network, whose motions are
dampened by SRR phosphorylation.
DNA binding affinity for ΔN301; ΔN280, a minimal fragment that recapitulates un- modified autoinhibited Ets-1 binding; and
ΔN2445P is KD 10-11, 10-10, and 10-8 M, indicating a gradual
dependence on length of the SRR region and its phosphorylation stage.
The distant fuzzy region in Ets-1 perturbs the dynamics of the protein–DNA interface and modulates the conformational
transition that leads to the tight, specific complex.
A striking linear pattern of change was observed from ΔN301 to ΔN280 to ΔN2445P in the amide
1H and 15N chemical shifts of almost all of the affected residues. This progressive colinear pattern
is a signature of an allosterically regulated molecule that is in conformational equilibrium between at least two states, with the
intermediate chemical shifts representing a population-weighted average of these states. Based on the correlation between
the linear pattern of amide chemical shifts with DNA binding affinity, the free Ets-1 has been proposed to be in equilibrium
between an active state (represented most closely by ΔN301), which is poised to bind DNA, and an inactive state
(represented by ΔN2445P).
Phosphorylation of five sites within the SRR region gradually reduces binding affinity up to 1000-fold. Phosphorylation interferes
with the formation of transient intraprotein contacts.
Alternative splicing of ETS1 removes the entire disordered SRR region; phosphorylation of this region reduces affinity for DNA
via modulating the flexibility of the interface. Hence, the activity of human Ets-1 is differentially regulated by two distinct
mechanisms: phosphorylation and alternative splicing.
The dynamic character of Ets-1 is likely essential for sequence-specific DNA binding andalso provides an opportunity for
gradual regulation of DNA binding (rheostat) depending on length and phosphorylation stage.