ATF-2/c-Jun and IRF-3 complexed with the interferon-beta enhancer
Oliver DeBarros '15 and Nate Petrou '15
Enhanceosomes are complexes of proteins that bind cooperatively
to a gene's enhancer region. Enhanceosomes are necessary for
transcriptional activation and important in the recruitment of basal
transcription factors to the promoter. One of the best understood
enhanceosome complexes is the virus-inducible enhancer of the
interferon-beta (IFN-beta) gene. In the presence of a virus,
interferon proteins are made by the IFN-beta gene, which is activated
by enhanceosome assembly. The assembly of the enhanceosome of
the IFN-beta gene involves the group of transcription factors
including ATF-3, c-Jun, IRF-3/IRF-7, NF-kB and HMGA1. Through DNA
binding to 4 different positive regulatory domains (PRDs) I-IV of the
IFN-beta enhancer these transcription factors can activate the gene. A
high level of specificity in the activation of the gene is achieved
through the cooperative binding of these proteins.
II. General Structure
This structure focuses on
the transcription factors ATF-2, c-Jun, IRF-3A and
IRF-3B in complex with 31 bp of the PRDIV-PRDIII region
of the IFN-beta enhancer.
ATF-2 and c-Jun
are two long alpha helices that form a leucine zipper heterodimer.
Both proteins contain a DNA-binding region containing several
basic amino-acid residues; as well as a dimerization domain
following a coiled-coil motif with heptad repeats.
The heterodimer interacts in a sequence specific manner
with the PRDIV enhancer within the promoter element of the gene.
The IRF-3 domains are composed of a
four-stranded antiparallel beta-sheets,
three alpha helices, and 3
The IRF-3 domains interact with DNA and cooperatively recruit
the ATF-2/c-Jun heterodimer; creating a complex that recognizes a
continuous sequence of approximately 24 base pairs within the PRDIV and PRDIII
enhancer elements of the IFN-beta promoter.
The enhanceosome's interaction with DNA is mostly sequence
III. ATF-2 and c-Jun DNA Binding
the C-terminus of the leucine zipper is involved in the
dimerization interface between ATF-2
and c-Jun, the N-terminus
interacts with DNA. Basic amino
acid residues, primarily Lysine and Arginine, make
extensive contact with the major groove of the DNA.
The ATF-2/c-Jun complex binds in the PRDIV region to an
8 bp sequence 5'-TGACATAG-3', similar to the high-affinity CRE
recognition sequence 5'-TGACGTCA-3'. ATF-2 binds the consensus half site (TGAC) and c-Jun
binds the nonconsensus half site
(ATAG), resulting in asymmetric binding favoring ATF-2.
c-Jun both bind DNA through hydrogen bonding to either the
bases or phosphate backbone. Van der Waals forces are
also present between DNA and both bZIP proteins. One
important interaction of ATF-2 involves the conserved Asn 344
residue. Asn 344 hydrogen bonds with O4 of base
T5 as well as N4 of base C27*
(*refers to c-Jun) on the opposite strand.
of c-Jun critical for DNA interaction is R270,
which engages in three hydrogen bonds; two with the phosphate
backbone and one with the base adenine.
IV. IRF-3 DNA Binding
The interferon regulatory factor (IRF) domains are
characterized by a conserved N-terminal DNA-binding domain of
around 120 amino acids that recognizes conserved DNA
sequences, named IFN-stimulated response elements (ISRE), in
the PRDIII region.
IRF-3A and IRF-3B have essentially the same
structure and make many similar DNA contacts. IRF-3A and
IRF-3B molecules are bound flanking the PRDIII region on
opposite faces of DNA, with the DNA duplex bending slightly
around the IRF recognition alpha helix.
The alpha3 recognition helix makes major groove contacts along the
DNA backbone, such as hydrogen bonds between Arg
81 and the conserved G.
also interacts with the minor groove, primarily by the L1
Loop. His 40, which is conserved in all IRF
domains, participates in two water-mediated
hydrogen bonds, both as an acceptor and donator.
42, specific to IRF-3, provides van der Waals
Also, after viral infection, IRF-3 is phosphorylated at several
serine and threonine residues which stimulates DNA binding and
increases transcriptional activation.
V. Cooperative Binding
Similar to other bZIP structures, the C-terminal leucine zipper of
the ATF-2/c-Jun heterodimer has a parallel coiled-coil
dimerization interface extending perpendicularly to the DNA. The leucine
zipper structural motif promotes dimerization by interaction of
at the d
position of the heptad repeat.
long, charged residues at the e and g positions of the heptad
repeat are important for dimerization, exemplified by pairs E379-R302*
salt bridge between E363 and R276*
also contributes to stabilization.
of ATF-2/c-Jun and IRF-3 to DNA is a cooperative process, with the
proteins making contacts with each other as well. IRF-3
preferentially binds to ATF-2/c-Jun bound DNA, evidenced by the
fact that IRF-3 exhibits decreased binding with free DNA. ATF-2
is contacted by the L1 and L3 loops of IRF-3A,
through hydrogen bonds between amino acids.
Extensive hydrogen bonding also occurs at the
interface of ATF-2 and IRF-3A.
bending of the DNA favors IRF-3A interaction, rather than IRF-3B,
with ATF-2/c-Jun due to the lack of binding of c-Jun to the
binds cooperatively to IRF-3B despite the fact that there are no
protein-protein contacts between the two. They
both bind DNA simultaneously and the position of one IRF
influences binding of the other.
CR, E Nistal-Villan, L Shen, A Garcia-Sastre, AK Aggarwal. 2007.
Structure of IRF-3 bound to the PRDIII-I regulatory
element of the human interferon-beta enhancer.
Cell 26: 703-16.
Daniel, Tom Maniatis, and Stephen C Harrison.
2004. Crystal structure of ATF-2/c-Jun and IRF-3
bound to the interferon-beta enhancer.
Journal 23: 4384-4393.
Daniel, Tom Maniatis, and Stephen C Harrison.
atomic model of the interferon-beta enhanceosome.
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