Floppy SOX: Mutual Induced Fit in HMG (High-Mobility Group) Box-DNA Recognition
Michael A. Weiss
Department of Biochemistry Case Western Reserve University
Cleveland, Ohio 44106
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ABSTRACT
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The high-mobility group (HMG) box defines a
DNA-bending motif of broad interest in relation to human development
and disease. Major and minor wings of an L-shaped structure provide a
template for DNA bending. As in the TATA-binding protein and a diverse
family of factors, insertion of one or more side chains between base
pairs induces a DNA kink. The HMG box binds in the DNA minor groove and
may be specific for DNA sequence or distorted DNA architecture. Whereas
the angular structures of non-sequence-specific domains are well
ordered, free SRY and related autosomal SOX domains are in part
disordered. Observations suggesting that the minor wing lacks a fixed
tertiary structure motivate the hypothesis that DNA bending and
stabilization of protein structure define a coupled process. We further
propose that mutual induced fit in SOX-DNA recognition underlies the
sequence dependence of DNA bending and enables the induction of
promoter-specific architectures.
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INTRODUCTION
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The high-mobility group (HMG) box defines a superfamily of
eukaryotic DNA-binding proteins of central importance in mammalian gene
regulation (1). This approximately 80-residue domain, originally
described in abundant nonhistone chromosomal proteins HMG1 and HMG2,
exhibits an unusual L-shaped structure (Fig. 1A
and Ref. 2). Three
-helices and an
N-terminal ß-strand pack to form major and minor wings (3, 4, 5, 6). Each
wing is formed by distinct elements of secondary structure. The major
wing comprises
-helix 1,
-helix 2, the first turn of
-helix 3,
and connecting loops; the minor wing comprises the N-terminal
ß-strand, the remainder of
-helix 3, and the C-terminal segment.
Together, the L-shaped structure presents an angular inner surface as a
template for DNA bending (asterisk in Fig. 1A
; Refs. 7, 8, 9, 10, 11).
A side view of the HMG box illustrates its flat architecture (Fig. 1B
).
Two groups of HMG boxes are distinguished by their DNA-binding
properties. Whereas HMG1 and related proteins typically contain two or
more HMG boxes that recognize distorted DNA structures with weak or
absent sequence specificity, specific architectural transcription
factors contain one HMG box that recognizes both distorted DNA
structures and specific DNA sequences (2). Each group docks within a
widened minor groove, directs the sharp bending of an underwound double
helix, and can enhance binding of unrelated DNA-binding motifs to
neighboring DNA sites (7, 8, 9, 10, 11). The extent of DNA bending varies among
HMG boxes but in each case the protein binds on the outside of the DNA
bend to compress the major groove. The dramatic effects of HMG boxes on
DNA structure are proposed to contribute to the assembly of specific
transcriptional preinitiation complexes and, in turn, to the regulation
of gene expression (12, 13).
This minireview focuses on the unusual conformational repertoire of Sox
proteins (14, 15), a subgroup of specific HMG-box factors defined by
similarity to Sry (16), the mammalian testis-determining factor encoded
by the Y chromosome (17). Designated Sox in relation to the
Sry box, this subgroup is ubiquitous in the animal kingdom and involved
in diverse developmental processes, including germ layer formation,
cell type specification, and organogenesis (14, 15). More than 20 Sox
genes have been identified based on greater than 50% sequence identity
with the HMG box of Sry. Genetic analyses of Sox genes in
humans, mice, and Drosophila melanogaster have demonstrated
essential roles in specific cell fate decisions (18, 19, 20, 21). Mutations or
deletions in human SRY are a cause of Swyers syndrome, in which
failure of testicular differentiation in a 46,XY embryo leads to a
female somatic phenotype and sterility (17). XY sex reversal can also
occur with variable penetrance due to mutations in SOX9, a
gene on human chromosome 19 (18, 19). Such mutations cause campomelic
dysplasia, a syndrome of bony abnormalities associated with XY gonadal
dysgenesis. Whereas clinical mutations in SRY cluster in its
HMG box (7), SOX9 mutations are widely distributed in its
coding region (22).
Sox genes are classified in seven families (designated AG) based on
extent of homology (>80% within a family). The families exhibit
similar DNA-binding and DNA-bending properties (14, 15). Random binding
site selection in vitro has revealed a shared specificity
for a core consensus sequence (5'-ACAAT-3'; Table 1
) with subtle distinctions in
preferences for flanking nucleotides (23, 24, 25, 26). Although such chemical
specificity is less stringent than that of classical major-groove
DNA-binding motifs, functional specificity is enhanced by
lineage-specific gene expression. The sequence specificity of Sox-9 and
Sox-10 can also be made more stringent by cooperative binding of
protein dimers to neighboring DNA target sites (27). Dimerization is
DNA dependent and mediated by a conserved N-terminal protein segment.
Although the structural basis of cooperativity is not understood,
analysis of the interaction of Sox10 with a Sox10 response
element active in neural crest-derived lineages [the protein
zero P(0) gene] has shown that DNA-dependent dimerization
markedly enhances specific DNA affinity and extent of induced DNA
bending (27). It is not known whether changes in DNA structure can, by
themselves, contribute to cooperativity, i.e. independently
of putative DNA-dependent protein-protein interactions. It is possible,
for example, that initial DNA bending and unwinding induced by one HMG
box can facilitate binding of a second HMG box to an adjoining DNA
site.
Specific DNA bends (typically in the range
7090o) may disallow or facilitate DNA binding
by unrelated transcription factors (i.e. proteins of other
structural classes) to adjoining DNA sites or facilitate
protein-protein interactions by flanking DNA-bound factors. HMG
box-induced DNA bends may also recruit trithorax or polycomb group
proteins, proposed to alter the higher-order chromatin structure
leading to the long-range regulation of gene expression. The
identification of target sites for Sox2 in murine and chicken
- and
-crystallin genes (28) and in human fgf4 (29) has led to
recognition of possible protein-protein interactions between Sox2 and
adjacent DNA-bound factors, including the Oct-3/4 POU domain. Analysis
of the murine fgf-4 enhancer in an embryonal carcinoma cell
line has demonstrated that synergistic activation of the enhancer
by Sox2 and Oct-3/4 requires a specific arrangement of factor-binding
sites (30). Synergy is mediated by at least two mechanisms, 1)
cooperative DNA binding by the HMG box and POU domain (30) and 2)
reciprocal conformational changes extending to regions outside of the
respective DNA-binding domains and leading to enhanced transcriptional
activation function (31). Analogous mechanisms may underlie functional
synergy between Sox10 and the classical Zn finger protein Sp1 in
transcriptional activation of genes encoding subunits of the neuronal
nicotinic acetylcholine receptor (32). The nature of these interactions
has not been defined. Individual Sox families often exhibit
conservation outside of the HMG box (i.e. within other
recognizable sequence motifs such as a leucine zipper or
serine-threonine-rich region). These regions include classical domains
of transcriptional activation or repression; therefore, Sox proteins
can, in principle, function as specific transcription factors (14, 15).
Functional evidence in one case is provided by the association between
campomelic dysplasia and truncation of or mutations within SOX9s
transactivation domain (18, 19). In contrast, almost all mammalian Sry
proteins lack discrete transactivation or repression domains. Although
sequences outside of the Sry HMG box are generally divergent (22, 33),
its extreme C terminus contains a PDZ-binding sequence. A candidate
SRY-interacting PDZ protein (SRY interacting protein 1; SIP1) has been
identified by the yeast two-hybrid assay (34). Involvement of
C-terminal sequences in SRY-mediated gene regulation would rationalize
the case report of 46,XY sex reversal associated with an SRY mutation
causing deletion of the C-terminal 41 residues but sparing the HMG box
(35). Target genes for human SRY or other mammalian Sry proteins are
not presently known.
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STRUCTURE OF THE NONSPECIFIC HMG BOX AND DNA COMPLEXES
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The purpose of this mini-review is to highlight the unusual
dynamics of the Sox HMG box (36, 37) and its possible implications for
function. An important foundation is provided by extensive nuclear
magnetic resonance (NMR) and crystallographic analyses of nonspecific
HMG boxes. Solution structures of four free nonspecific HMG boxes (the
A and B domains of HMG-1, Drosophila chromosomal protein
HMG-D, and yeast protein HNP6A) have been determined (3, 4, 5, 6).
Structures of these domains are well defined in the absence of DNA.
Unlike conventional globular domains, the two wings of the HMG box
contain discrete hydrophobic cores. The primary core, located between
helix 1 and helix 2, stabilizes the confluence of the major wing. Its
organization is dominated by conserved aromatic-aromatic interactions
as illustrated in the bottom panel of Fig. 2
. Such interactions not only contribute
to the hydrophobic character of the core but may also impose geometric
constraints on packing. Possible packing arrangements of side chains is
constrained by the size and planarity of the aromatic rings and by
weakly polar electrostatic interactions. The latter involve the
-electrons circulating above and below the faces of the aromatic
rings and the partial positive charges of C-H groups at ring edges. A
second and less extensive mini-core occurs in the minor wing between
-helix 3 and the N-terminal ß-strand (upper panel of
Fig. 2
). Both wings contribute to the motifs angular DNA-binding
surface (7, 8, 9, 10, 11). HMG-D (4, 10) and NHP6A (11) are similar to the B
domain of HMG1 (5, 6) whereas the A domain (3) differs in the structure
of helix 1 and length of the loop between helix 1 and 2. The functional
implications of these differences are not well understood.
Investigation of main-chain dynamics by analysis of
15N heteronuclear relaxation times suggests that
the nonspecific HMG box can be appropriately described as a rigid,
axially symmetric ellipsoid (38).

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Figure 2. Overview of Side-Chain Packing in the SRY HMG Box
Upper panel, A comparison of the bound SRY box
(red; Ref. 7) and nonspecific boxes (HMG-D, blue;
and HMG-1B, gold) is shown at left. Details of
side-chain packing in the minor wing of the bound SRY HMG box are shown
at right. Lower panel, Structure of the major
wing of SRY in a specific complex showing packing of multiple aromatic
rings and aliphatic side chains. The details of such packing differ in
Lef-1 (8 ) and nonspecific HMG boxes (3 4 5 6 ).
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Crystal structures have been obtained for two nonspecific complexes, 1)
the A domain of HMG1 with a 20-bp DNA site containing a
cis-platin adduct (9); and 2) HMG-D bound to a an unmodified
decamer DNA site (10). In addition, an NMR-derived model of a
nonspecific NHP6A-DNA complex has also been described (11). The
structures of bound and free nonspecific HMG boxes are similar. Small
structural adjustments occur, presumably to accommodate the details of
the distorted DNA surface. The protein-DNA interface is remarkable for
nonpolar contacts between the motifs angular protein surface and the
DNAs expanded, underwound, and bent minor groove. A conserved wedge
of aliphatic and aromatic side chains inserts between successive base
pairs to disrupt base stacking as observed in SRY (39, 40). Such
partial intercalation is similar to that observed in DNA complexes of
the TATA-binding protein (TBP) (41, 42), LacI/PurR family of repressors
(43, 44), and hyperthermophilic archaeal chromosomal proteins
(45). Although these proteins are unrelated in overall structure, use
of a cantilever side chain provides a common mechanism by which base
stacking is disrupted at a DNA kink. This side chain inserts
between base pairs, unlike conventional contacts between
side chains and edges of bases. At the site of insertion
base pairing is, in general, maintained. Of HMG boxes, the structure of
the HMG-D complex (10) is particularly remarkable for its three
distinct sites of partial intercalation, yielding an overall bend angle
of 111o. The interface also contains three
water-mediated hydrogen bonds between bases and polar or charged side
chains. Water is proposed to function as an adaptor between a given
protein side chain and variable target bases, making possible alternate
interactions by a common DNA-binding surface. Together, multiple and
distributed protein-DNA contacts in the HMG-D complex give rise to a
smooth overall DNA bend. A contrasting mode of binding is observed in
the complex between HMG-1A and the cis-platin-DNA adduct
(9). Binding and bending occur predominantly at the site of chemical
modification, which induces a preexisting kink in the DNA. The
structural basis of recognition of distorted DNA structures by
nonspecific HMG domains has recently been reviewed (46, 47).
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STRUCTURES OF SPECIFIC COMPLEXES
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The structures of complexes between the specific HMG boxes of SRY
and Lef-1 (a non-Sox-specific domain with related specificity
5'-TTCAAA-3'; nonconsensus SRY nucleotides in
bold) and their cognate DNA sites have been determined by
NMR spectroscopy (Fig. 3
, A and B; Refs.
7, 8). The structures of the bound HMG boxes strongly resemble those
of nonspecific HMG boxes. As anticipated by homology, the SRY- and
Lef-1 HMG boxes are each L-shaped and contain discrete wing-specific
hydrophobic cores (Fig. 2
, A and B). Comparison of specific and
nonspecific complexes has provided insight into the origins of sequence
specificity and differences in the position or extent of partial
side-chain intercalation (10). Remarkably, such properties seem to
reflect sequence changes at only a handful of protein positions. Two
examples illustrate a common theme: 1) Lef-1, SRY, and Sox domains
contain an invariant Asn at position 10. The Asn carboxamide makes
sequence-specific bidentate hydrogen bonds to edges of base pairs at an
invariant 5'-TG-3' step in target DNA sites (7, 8). The corresponding
side chain in nonspecific domains is Ser10, which also contacts DNA but
without sequence specificity. Its interactions in the HMG-D complex,
described as sequence neutral, are water-mediated (10). 2) Residues 32,
33, and 36 are nonpolar and likewise sequence neutral in nonspecific
domains but polar and capable of specific contacts in specific domains.
An example is provided by Phe32 in HMG-1A, which inserts into the
cis-platin-induced DNA kink. Similarly, Val32 partially
intercalates in the HMG-D complex. The corresponding side chain in SRY
and Lef-1 is serine, the polarity of which apparently precludes partial
intercalation. In the future, the proposed relationship between protein
sequence and sequence specificity (10, 46) can be tested by
mutagenesis.

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Figure 3. Comparison of Sequence-Specific HMG Boxes
A, Ribbon model of specific SRY-DNA complex (7 ). The protein is shown
in white and DNA backbone in red. B, Ribbon model
of specific Lef1-DNA complex (8 ). The protein is shown in
blue and DNA backbone in green. C, Superposition
of SRY and Lef1 HMG boxes according to the main-chain atoms of
-helices 1 and 2 demonstrates relative displacement of the minor
wing (asterisk). D, NMR-derived ensemble of Sox-4 contains
well ordered major wing and disordered minor wing (37 ). Although the
N-terminal segment is locally disordered, helix 3 is locally ordered
but lacks a coherent orientation relative to the major wing. Details of
packing between helix 1 and helix 2 apparently differ from those of
other HMG boxes.
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Specific SRY and Lef-1 complexes exhibit overall similarities as well
as key apparent differences. Each exhibits a single side-chain
cantilever at corresponding positions: partial intercalation by Ile
(SRY; position 13 of the HMG box consensus, lower panel of
Fig. 2
) or Met (Lef-1) similarly disrupts base stacking but not base
pairing (7, 8, 39, 40). Additional sites of insertion as defined in
nonspecific complexes (9, 10, 11) are not observed. The reported
orientation of aromatic side chains in the major core of SRY differs in
detail from that of Lef-1, which is similar to nonspecific HMG boxes.
Specific SRY and Lef-1 complexes also differ in apparent bend angle.
The 15-bp DNA duplex employed in the Lef-1 complex is bent by
approximately 110o and exhibits a remarkable
similarity to the corresponding portion of the nonspecific HMG-D
complex. The 8-bp DNA duplex employed in the SRY complex is less bent
(40o-80o); however, its
limited length inhibits accurate assessment of the bend angle (J. Love
and P. E. Wright, personal communication). The marked difference
in extent of DNA bending is in part due to the influence of Lef-1s
basic tail, which binds across the major groove as an electrostatic
clamp (8). The different DNA bend angles are associated with a change
in the orientation between major and minor wings as illustrated by
molecular modeling. Superposition of
-helices 1 and 2 in SRY and
Lef-1 gives rise to a large relative displacement in the apparent
position of
-helix 3 (asterisk in Fig. 3C
). Because
structures of free SRY and Lef-1 have not independently been
determined, it is not know whether the apparent differences between
their bound structures result from differential induced fit or instead
preexist in the respective unbound proteins.
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STRUCTURE AND DYNAMICS OF SOX DOMAINS
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The solution structures of lymphocyte transcriptional activator
Sox-4 and testis-specific factor Sox5 in the absence of DNA have been
found to exhibit a novel combination of order and disorder (Fig. 3D
).
The three canonical
-helices of the HMG box are present and locally
well ordered (36, 37). Whereas the tertiary structure of the major wing
is well defined in Sox4, the minor wing is not. Similar features occur
in free SRY (E. Rivera, N. Phillips, and M. A. Weiss, unpublished
results). The major wings characteristic aromatic- aromatic
interactions are associated with dispersion of NMR chemical shifts (the
inequivalence of precise proton resonance frequencies due to
differences in local environments in a protein) and short-range
distances between neighboring side chains in space (nuclear Overhauser
enhancements; NOEs). These NMR features are similar in free domains and
in specific DNA complexes. The minor wings characteristic chemical
shifts and NOEs are, by contrast, absent in spectra of the free
domains. These include otherwise prominent interactions among the side
chains of Val5, His65, Tyr69, and Tyr72 (upper panel of Fig. 2
), residues conserved among Sox sequences (Fig. 4
). An illustrative example is provided
by ring currents generated by aromatic rings in
-helix 3 of SRY
(Fig. 5
). Ring currents, local magnet
fields arising from aromatic electrons, are readily estimated by a
parameterized dipole approximation (48). In the bound state, such ring
currents intersect with the N-terminal ß-strand due to folding of the
minor wing. In particular, the
-methyl groups of Val5 overlay the
aromatic ring of Tyr69, giving rise to a large up-field ring-current
shift (boldface values in Table 2
) associated with long-range NOEs. None
of these features are observed in spectra of the free SRY or Sox4
domains: instead minor-wing side chains, such as Val5 and Tyr72,
exhibit motional narrowing, near-random coil chemical shifts, and an
absence of long-range NOEs. Because chemical shifts of minor-wing
aromatic and methyl resonances can readily by obtained even in the
absence of exhaustive NMR analysis, we suggest that these features will
be of general value in screening other Sox domains for minor-wing
disorder and induced fit on DNA binding.

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Figure 4. Comparison of N-Terminal and C-Terminal Sequences
of Human and Murine SRY and Selected Sox Domains
The conservation of Val or Ile at position 5 and an aromatic side
chain at position 69 is highlighted (boxes). These side
chains provide valuable NMR markers for the dynamics and folding of the
minor wing (see upper panel of Fig. 2 ).
Asterisk indicates position 13 of HMG-box consensus,
which residue inserts between base pairs as a cantilever to disrupt
base stacking (7 36 39 40 ).
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Figure 5. Simulation of Aromatic Ring Currents in the Bound
Structure of the SRY HMG Box
Stereo representation of the protein backbone (white)
and selected side chains. Red balls represent contours
at an up-field ring current of 0.5 ppm; negative ring currents are not
displayed. At bottom a cluster of four aromatic ring currents from
C-terminal residues (His65, Tyr69, Tyr72, and Tyr74) is seen to impinge
upon the neighboring N-terminal ß-strand. The side chain of Val5 is
shown in white encased within the ring current of Tyr69
(see Table 2 ).
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An ensemble of NMR-based models of Sox4 (37), obtained by
distance geometry and simulated annealing (DG/SA), in fact contains no
fixed relationship between
-helix 3 and the major wing (
-helices
1 and 2). The N-terminal strand is disordered and detached from
-
helix 3. The major hydrophobic core with its conserved aromatic
side chains is well organized whereas the minor hydrophobic core is
absent. Packing of the N-terminal strand of Sox4 against
-helix 3 is
induced on specific DNA binding (E. Rivera, N. Phillips, and M. A.
Weiss, unpublished results). The presence of ordered
-helical
segments with imprecise tertiary relationship is reminiscent of a
molten globule (49), an intermediate state of protein organization
observed in protein-folding pathways. A schematic model of an
equilibrium between open and closed minor wings is provided in Fig. 1B
.
We caution that the precision of DG/SA models reflects the number of
restraints and may or may not correspond to physical fluctuations. The
minor wings imprecision as seen in the Sox4 model (Fig. 3D
) thus
reflects a paucity of NMR-derived restraints in this region of the
protein. The short range and steep distance dependence of the NOE
(typically <5 Å and scaling with r-6)
implies that distances longer than this cut off cannot routinely be
measured. Thus, spatial separations of 8 Å may appear similar to
spatial separations of 20 Å as each would give rise to an unobserved
signal. Because absence of evidence does not necessarily imply evidence
of absence, the DG/SA calculation is underdetermined and hence
unphysical. The model shown in Fig. 3D
was thus proposed as a working
hypothesis rather than definitive characterization of the extent of
disorder in the minor wing.
Evidence for an equilibrium between open and closed conformations has
been obtained in studies of Sox5 by an elegant combination of
biophysical techniques. Although NMR studies likewise suggested that
its minor wing is largely unfolded at 37 C, decreasing temperature was
found to lead to progressive folding of this segment (36). Intensities
of key interresidue NOEs, chosen to reflect tertiary contacts, were
monitored as a function of temperature. Whereas the intensity of NOEs
diagnostic of the major wings tertiary structure was unaffected by
temperature in the range 1631 C, attenuation of minor wing-specific
NOEs was observed with increasing temperature. Although these NMR
observations in themselves could have multiple interpretations,
complementary evidence of discrete major and minor wing unfolding
transitions was obtained by fluorescence spectroscopy and differential
scanning calorimetry. The minor wing of Sox5 unfolds with a midpoint of
34 C whereas the major wing unfolds with a midpoint of 46 C (36).
Analogous studies of the free SRY HMG box suggests that its minor wing
is unfolded even at temperatures as low as 4 C (N. Phillips and M.
A. Weiss, unpublished results). Unfortunately, none of these methods
can provide a quantitative estimate of the extent of unfolding.
Although the unusual spectroscopic features of Sox domains
emphasize the distinction between the dynamics of the two wings,
NMR and fluorescent studies have not to date addressed the extent of
excursions between the domains N-terminal segment and helix 3,
i.e. how open is the "open" state? In particular,
because the NMR methods employed in these studies are based on the
short-range NOE interaction (48), it is possible, in principle, that
more long-range order is present in solution than is suggested by the
DG/SA model shown in Fig. 3D
. In the future it would be of interest to
investigate the extent of long-range correlation between the major wing
and
-helix 3 by use of residual dipolar couplings in
partially oriented samples in solution. This new NMR methodology (50)
circumvents the restriction of NMR parameters to local properties or
short-range interactions (48). It would be of complementary interest to
measure distributions of long-range distances by time-resolved
fluorescence resonance energy transfer (FRET) (51, 52). Although FRET
is formally a r-6
dipole-dipole interaction like the NOE, its distance range is
determined by the Förster distance (Ro)
governing resonance energy transfer between donor and acceptor probes.
This distance is probe dependent and typically lies in the range 1080
Å. Attachment of suitable probes to the N-terminal segment and helix 3
would thus enable direct characterization of long-range distances and
fluctuations. It is likely that time-resolved FRET analysis of Sox5
would permit a definitive test of the hypothesis that the free domain
exists in an equilibrium between open and closed conformations.
The DNA-dependent order-disorder transition of Sox domains
differs in kind from those of basic zipper (bZIP) and basic
helix-loop-helix (bHLH) major-groove DNA-binding motifs (53, 54, 55, 56, 57). The
latter contain disordered N-terminal basic arms, which form divergent
pairs of recognition
-helices on specific DNA binding. Induced fit
in the protein thus occurs at the level of secondary structure.
Although small conformational adjustments occur in DNA structure,
including limited DNA bending (typically
<20o), the DNA remains in the B family, and its
major groove acts essentially as a preformed template for protein
folding (58, 59). The
-helical structure of Sox domains is by
contrast preorganized: it is tertiary structure that is
specified on DNA binding (36, 37, 58). Sox-induced DNA bending thus
reflects a bidirectional induced fit wherein the Sox domain
instructs the DNA how to bend as the DNA instructs the protein to
complete its tertiary fold. Because the angular surface of the Sox HMG
box is not fixed, a given domain may be compatible with a range of DNA
bend angles rather than a single value. This hypothesis suggests that
the precise DNA bend adopted in a specific complex could depend on the
exact DNA target sequence and presence of neighboring protein-DNA
complexes. Indeed, sequence-dependent DNA bending has been inferred
from electrophoretic measurements of sequence-specific HMG boxes,
including human and murine Sry and LEF1 (Table 3
; Refs. 60, 61). Induced bend angles
can differ by as much as 30o, implying a
substantial difference in underlying DNA and protein structures.
Accordingly, it would be of future interest to obtain crystal or NMR
structures of the same Sox HMG box bound to variant DNA sites
associated with different electrophoretic bend angles. Analogous
crystallographic studies of the TBP in variant DNA complexes revealed
no changes in bend angle, suggesting that TBP (unlike a Sox domain)
functions as a robust and preformed template for DNA bending (62).
Why are specific HMG boxes floppy? We propose that adaptability
of the motifs angular surface enables a Sox protein to induce
different architectures in different functional contexts. We imagine
that target genes for a given factor will differ, for example,
in the precise sequence of Sox binding sites and its
combinatorial relation to other factor-binding sites in the same
promoter or enhancer. Because context-dependent changes in overall
architecture may differentially affect transcription, a single factor
may exert fine control over relative levels of expression within a set
of target genes. Were the specific HMG box a rigid platform directing a
preset DNA bend angle, then the cell might need a very large collection
of factors, each calibrated to a different angle, to effect such fine
control. Use of a single flexible motif with an adjustable set point
for DNA bending would represent a striking economy of protein design.
Testing this hypothesis will require the in vitro
reconstitution and structural characterization of Sox-specific
enhanceosomes.
Note Added in Proof.
Recent studies of the murine Sox-5 HMG box by multidimensional NMR and
15N relaxation measurements have demonstrated anomalous
mobility of the minor wing (63).
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ACKNOWLEDGMENTS
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We thank A. S. Stern for ring-current shift calculations;
D. N. Jones and N. Narayana for assistance with figures; G. Chen,
E. Haas, A. Jancso, J. Radek, E. Rivera, and N. Phillips for
communication of results before publication; and P. Donahoe, C.-Y.
King, J. Love, and P. E. Wright for discussion.
 |
FOOTNOTES
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Address requests for reprints to: Dr. Michael A. Weiss, Department of Biochemistry, Case Western Reserve University School of Medicine, 2109 Adelbert Road, Cleveland, Ohio 44106-4935.
This work was supported in part by NIH Grant CA-63485 to
M.A.W.
Received for publication October 13, 2000.
Revision received December 1, 2000.
Accepted for publication December 20, 2000.
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