NF-κB p65 (RelA) homodimer uses distinct mechanisms to recognize DNA targets

Chen, Yongqing; Sengchanthalangsy, Lei Lei; Hackett, Arthur; Ghosh, Gourisankar

doi:10.1016/s0969-2126(00)00123-4

Cited by 68 publications

(64 citation statements)

References 31 publications

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“…Together, these structures reveal that the p50 and p52 subunits bind to the 5 bp 5 0 -GGGRN-3 0 half-site of the consensus sequence and the RelA, c-Rel and RelB subunit prefer the 4 bp 5 0 -YYCC-3 0 half-site (Chen and Ghosh, 1999;Chen et al, 2000). Following this mode of binding, both the p50 and p52 homodimers appear to prefer an 11 bp kB site comprising of two 5 bp half-sites separated by a central A T bp.…”

Section: Dna Target Sites Of Nf-kb Dimersmentioning

confidence: 87%

Transcriptional regulation via the NF-κB signaling module

2006

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Stimulus-induced nuclear factor-jB (NF-jB) activity, the central mediator of inflammatory responses and immune function, comprises a family of dimeric transcription factors that regulate diverse gene expression programs consisting of hundreds of genes. A family of inhibitor of jB (IjB) proteins controls NF-jB DNA-binding activity and nuclear localization. IjB protein metabolism is intricately regulated through stimulus-induced degradation and feedback re-synthesis, which allows for dynamic control of NF-jB activity. This network of interactions has been termed the NF-jB signaling module. Here, we summarize the current understanding of the molecular structures and biochemical mechanisms that determine NF-jB dimer formation and the signal-processing characteristics of the signaling module. We identify NF-jB-jB site interaction specificities and dynamic control of NF-jB activity as mechanisms that generate specificity in transcriptional regulation. We discuss examples of gene regulation that illustrate how these mechanisms may interface with other transcription regulators and promoter-associated events, and how these mechanisms suggest regulatory principles for NF-jB-mediated gene activation.

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Section: Dna Target Sites Of Nf-kb Dimersmentioning

confidence: 87%

Transcriptional regulation via the NF-κB signaling module

2006

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“…Dimerization of p50 Is Not Affected by Mutating Ser 65 , Ser 337 , or Ser 342 -The x-ray crystal structures of the p50/p50 homodimer and p50/p65 heterodimer binding to DNA have been solved (12)(13)(14)(15)(16). Based on the structural information, Ser 65 resides in the DNA recognition loop of p50 but does not directly contact DNA (Fig.…”

Section: Sermentioning

confidence: 99%

“…and p50/p50 and p65/p65 homodimer binding to DNA have revealed a conformation often referred to as a "butterfly" (12)(13)(14)(15)(16). The DNA recognition loop (L1) in the N-terminal half of NF-B Rel homology domain mediates base-specific DNA contacts, whereas the C-terminal half is responsible for dimerization and nonspecific DNA contacts (17).…”

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confidence: 99%

Phosphorylation of Serine 337 of NF-κB p50 Is Critical for DNA Binding

Hou

Guan

Ricciardi

2003

Journal of Biological Chemistry

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It has been demonstrated that phosphorylation of the p50 subunit of NF-B is required for efficient DNA binding, yet the specific phospho-residues of p50 have not been determined. In this study, we substituted all of the serine and conserved threonine residues in the p50 Rel homology domain and identified three serine residues, Ser 65 , Ser 337 , and Ser 342 , as critical for DNA binding without affecting dimerization. Although substitution with negatively charged aspartic acid at each of these positions failed to restore DNA binding, substitution with threonine, a potential phospho-acceptor, retained DNA binding for residues 65 and 337. In particular, Ser 337 , in a consensus site for protein kinase A (PKA) and other kinases, was shown to be phosphorylated both in vitro and in vivo. Importantly, phosphorylation of Ser 337 by PKA in vitro dramatically increased DNA binding of p50. This study shows for the first time that the DNA binding ability of NF-B p50 subunit is regulated through phosphorylation of residue Ser 337 , which has implications for both positive and negative control of NF-B transcription.The transcription factor NF-B acts as a central regulator of inflammatory, immune, and stress responses by controlling gene expression of cytokines, chemokines, immunoreceptors, antigen-presenting proteins, growth factors, transcription factors, cell adhesion molecules, stress response proteins, and apoptotic regulators (1, 2). Members of Rel/NF-B transcription factor family include Dorsal, Relish, and Dif in Drosophila and p65 (RelA), RelB, c-Rel, p50/p105, and p52/p100 in vertebrates. All of these proteins have a highly conserved DNAbinding and dimerization domain called the Rel homology domain. The vertebrate Rel family proteins can form homodimers or heterodimers that bind to 10-basepair B sites in the promoters of NF-B target genes (1, 3, 4). The most common and important NF-B transactivating species is the p50/p65 heterodimer. The p65 subunit, which contains a transactivation domain in the C terminus of the Rel homology domain, is responsible for the ability of NF-B to stimulate transcription. By contrast, p50 subunit, which lacks a transactivation domain, functions mainly in NF-B DNA binding (5-9). Another important dimeric species, the p50/p50 homodimer, serves mainly as a negative regulator of NF-B activity through competing with p50/p65 for NF-B response elements on DNA and through its association with co-repressor histone deacetylase (10, 11). The x-ray crystal structures of p50/p65 heterodimer and p50/p50 and p65/p65 homodimer binding to DNA have revealed a conformation often referred to as a "butterfly" (12-16). The DNA recognition loop (L1) in the N-terminal half of NF-B Rel homology domain mediates base-specific DNA contacts, whereas the C-terminal half is responsible for dimerization and nonspecific DNA contacts (17).NF-B activity is regulated by nuclear translocation. In most cell types, p50/p65 heterodimers exist in the cytoplasm as an inactive form associated with the inhibitor protein, I B. A w...

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“…Structures of carbon catabolite protein A (CcpA) and mammalian transcription factor NF-B p65 (RelA) have been determined when bound to three different operator sequences in each case (35)(36)(37). The two proteins bind their cognate operators with similar affinities but use different structural strategies for interactions.…”

Section: Discussionmentioning

confidence: 99%

Plasticity in Repressor-DNA Interactions Neutralizes Loss of Symmetry in Bipartite Operators

Jain

Narayanan

Nair

2016

Journal of Biological Chemistry

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Transcription factor-DNA interactions are central to gene regulation. Many transcription factors regulate multiple target genes and can bind sequences that do not conform strictly to the consensus. To understand the structural mechanism utilized by the transcription regulators to bind diverse target sequences, we have employed the repressor AraR from Bacillus subtilis as a model system. AraR is known to bind to eight different operator sites in the bacterial genome. Although there are differences in the sequences of four of these operators, ORE1, ORX1, ORA1, and ORR3, the AraR-DNA binding domain (AraR-DBD) as well as full-length AraR unexpectedly binds to each of these sequences with similar affinities as measured by fluorescence anisotropy experiments. We have determined crystal structures of AraR-DBD in complex with two different natural operators ORE1 and ORX1 up to 2.07 and 1.97 Å resolution, respectively. These structures were compared with the previously reported structures of AraR-DBD bound to two other natural operators (ORA1 and ORR3). Interactions of two molecules of AraR-DBD with the symmetric operator, ORE1, are identical, but their interaction with the non-symmetric operator ORX1 results in breakdown of the symmetry in protein-DNA interactions. The novel interactions observed are accompanied by local conformational change in the DNA. ChIP-sequencing (ChIP-Seq) data on other transcription factors has shown that they can bind to diverse targets, and hence the plasticity exhibited by AraR may be a general phenomenon. The ability of transcription factors to form alternate interactions may be important for employment in new functions and evolution of novel regulatory circuits.The binding of proteins to cognate DNA sequences with high specificity is critical for a number of cellular processes. Among such proteins, the ability to detect and bind target sequences present at different locations in the whole genome is especially crucial for the function of transcription factors. This ability enables the transcriptional regulatory pathways to function with remarkable precision and sensitivity. The specificity of transcription factor for its cognate DNA sequence is largely achieved due to non-covalent interactions between the amino acid side chains and the functional groups present on the DNA bases in the major and minor groove (1). In addition, the shape of the DNA also plays a significant role in specific protein-DNA recognition (2-5). The availability of transcription factor binding data spanning entire genomes has led to identification of new targets for transcription factors. It has been observed that a single transcription factor can bind to several different sites throughout the genome. Sequence deviations have been seen in target sites that are recognized by the same transcription factor. Examples include glp repressors of Escherichia coli and NdgR from Streptomyces coelicolor, which recognize 13 and 19 different operator sequences, respectively (6, 7). The sequence differences suggest the possibility o...

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NF-κB p65 (RelA) homodimer uses distinct mechanisms to recognize DNA targets

Abstract: Taken together, these structures reveal that p65 exhibits the unique capability to specifically bind DNA targets of variable lengths from four to ten base pairs. Also, the small protein segment Arg41-Ser42-Ala43 is at least partially responsible for flexibility in DNA-binding modes.

Cited by 68 publications

References 31 publications

Transcriptional regulation via the NF-κB signaling module

Transcriptional regulation via the NF-κB signaling module

Phosphorylation of Serine 337 of NF-κB p50 Is Critical for DNA Binding

Plasticity in Repressor-DNA Interactions Neutralizes Loss of Symmetry in Bipartite Operators

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