NF-κB Dimers Research Articles

Biophysical characterization of RelA-p52 NF-κB dimer-A link between the canonical and the non-canonical NF-κB pathway.

The NF-κB family consists of the key transcription factors of the NF-κB signaling pathway, well known for its role in innate immune response, organogenesis, and a variety of cellular processes. The five NF-κB subunits-RelA, RelB, c-Rel, p50, and p52-are functional dimers, each of which share a conserved DNA binding domain which contains the dimerization domain (DD) as well. The NF-κB subunits can form 15 potential dimers among themselves of which, RelA-p50 is extensively studied and has largely become synonymous with NF-κB for transcription activation. While various reports have highlighted the importance of NF-κB subunit specificity in the transcription regulation of certain target genes, the dynamic nature of the NF-κB dimer composition is not well understood. In this study, we biophysically characterized six combinatorial dimers from three NF-κB subunits: RelA, p50, and p52, using NMR spectroscopy and differential scanning calorimetry. We show that the dimer composition is dynamic and can readily undergo exchange although at varied rates. Among the six dimers formed, RelA-p52 is found to be the most stable dimer with RelA-RelA being the least. Our results provide a plausible explanation as to why the RelA-p52 heterodimer is active during the later stages of the NF-κB activation and serve as a link between the canonical and non-canonical NF-κB pathway.

Polyomavirus ALTOs, but not MTs, downregulate viral early gene expression by activating the NF-κB pathway

Polyomaviruses are small, circular dsDNA viruses that can cause cancer. Alternative splicing of polyomavirus early transcripts generates large and small tumor antigens (LT, ST) that play essential roles in viral replication and tumorigenesis. Some polyomaviruses also express middle tumor antigens (MTs) or alternate LT open reading frames (ALTOs), which are evolutionarily related but have distinct gene structures. MTs are a splice variant of the early transcript whereas ALTOs are overprinted on the second exon of the LT transcript in an alternate reading frame and are translated via an alternative start codon. Merkel cell polyomavirus (MCPyV), the only human polyomavirus that causes cancer, encodes an ALTO but its role in the viral lifecycle and tumorigenesis has remained elusive. Here, we show MCPyV ALTO acts as a tumor suppressor and is silenced in Merkel cell carcinoma (MCC). Rescuing ALTO in MCC cells induces growth arrest and activates NF-κB signaling. ALTO activates NF-κB by binding SQSTM1 and TRAF2&3 via two N-Terminal Activating Regions (NTAR1+2), resembling Epstein-Barr virus (EBV) Latent Membrane Protein 1 (LMP1). Following activation, NF-κB dimers bind the MCPyV noncoding control region (NCCR) and downregulate early transcription. Beyond MCPyV, NTAR motifs are conserved in other polyomavirus ALTOs, which activate NF-κB signaling, but are lacking in MTs that do not. Furthermore, polyomavirus ALTOs downregulate their respective viral early transcription in an NF-κB- and NTAR-dependent manner. Our findings suggest that ALTOs evolved to suppress viral replication and promote viral latency and that MCPyV ALTO must be silenced for MCC to develop.

Selective regulation of a defined subset of inflammatory and immunoregulatory genes by an NF-κB p50-IκBζ pathway.

The five NF-κB family members and three nuclear IκB proteins play important biological roles, but the mechanisms by which distinct members of these protein families contribute to selective gene transcription remain poorly understood, especially at a genome-wide scale. Using nascent transcript RNA-seq, we observed considerable overlap between p50-dependent and IκBζ-dependent genes in Toll-like receptor 4 (TLR4)-activated macrophages. Key immunoregulatory genes, including Il6, Il1b, Nos2, Lcn2, and Batf, are among the p50-IκBζ-codependent genes. IκBζ-bound genomic sites are occupied at earlier time points by NF-κB dimers. However, p50-IκBζ codependence does not coincide with preferential binding of either p50 or IκBζ, as RelA co-occupies hundreds of genomic sites with the two proteins. A common feature of p50-IκBζ-codependent genes is a nearby p50/RelA/IκBζ-cobound site exhibiting p50-dependent binding of both RelA and IκBζ. This and other results suggest that IκBζ acts in concert with RelA:p50 heterodimers. Notably, p50-IκBζ-codependent genes comprise a high percentage of genes exhibiting the greatest differential expression between TLR4-stimulated and tumor necrosis factor receptor (TNFR)-stimulated macrophages. Thus, our genome-centric analysis reveals a defined p50-IκBζ pathway that selectively activates a set of key immunoregulatory genes and serves as an important contributor to differential TNFR and TLR4 responses.

UNLOCKING THE THERAPEUTIC POTENTIAL: EXPLORING NF-κB AS A VIABLE TARGET FOR DIVERSE PHARMACOLOGICAL APPROACHES

NF-κB is a vital transcription factor that responds to diverse stimuli like cytokines, infections, and stress. It forms different dimers, binds to specific DNA sequences, and regulates gene expression. It operates through two pathways: canonical (for inflammation and immunity) and non-canonical (for specific processes). These pathways tightly control activity of NF-κB and impacting gene expression. Aberrant NF-κB activation is linked to cancer and other diseases, making it a potential therapeutic target. This review explores the role of NF-κB in disease and its therapeutic potential in various conditions. Intricate signal transduction processes lead to NF-κB activation by phosphorylating IκB proteins, allowing NF-κB dimers to enter the nucleus and influence gene expression. This dynamic regulation involves co-activators and interactions with other transcription factors, shaping complex gene expression programs. Understanding the multifaceted functions off NF-κB is crucial as its deregulation is associated with a range of diseases, including cancer, autoimmune disorders, and inflammatory conditions. Exploring recent studies offers insights into potential therapeutic strategies aimed at modulating NF-κB activity to restore health and combat various pathological conditions. This Comprehensive review is based on the role of NF-κB in disease pathogenesis and therapeutic implications.

Polyomavirus ALTOs, but not MTs, downregulate viral early gene expression by activating the NF-κB pathway.

Polyomaviruses are small, circular dsDNA viruses that can cause cancer. Alternative splicing of polyomavirus early transcripts generates large and small tumor antigens (LT, ST) that play essential roles in viral replication and tumorigenesis. Some polyomaviruses also express middle tumor antigens (MTs) or Alternate LT ORFs (ALTOs), which are evolutionarily related but have distinct gene structures. MTs are a splice variant of the early transcript whereas ALTOs are overprinted on the second exon of the LT transcript in an alternate reading frame and are translated via an alternative start codon. Merkel cell polyomavirus (MCPyV), the only human polyomavirus that causes cancer, encodes an ALTO but its role in the viral lifecycle and tumorigenesis has remained elusive. Here, we show MCPyV ALTO acts as a tumor suppressor and is silenced in Merkel cell carcinoma (MCC). Rescuing ALTO in MCC cells induces growth arrest and activates NF-κB signaling. ALTO activates NF-κB by binding SQSTM1 and TRAF2&3 via two N-Terminal Activating Regions (NTAR1+2), resembling Epstein-Barr virus (EBV) Latent Membrane Protein 1 (LMP1).. Following activation, NF-κB dimers bind the MCPyV non-coding control region (NCCR) and downregulate early transcription. Beyond MCPyV, NTAR motifs are conserved in other polyomavirus ALTOs, which activate NF-κB signaling, but are lacking in MTs that do not. Furthermore, polyomavirus ALTOs downregulate their respective viral early transcription in an NF-κB and NTAR dependent manner. Our findings suggest that ALTOs evolved to suppress viral replication and promote viral latency and that MCPyV ALTO must be silenced for MCC to develop.

NF-κB signaling activation and roles in thyroid cancers: implication of MAP3K14/NIK

Among follicular-derived thyroid cancers (TC), those with aggressive behavior and resistance to current treatments display poor prognosis. NF-κB signaling pathways are involved in tumor progression of various cancers. Here, we finely characterize the NF-κB pathways and their involvement in TC. By using immunoblot and gel shift assays, we demonstrated that both classical and alternative NF-κB pathways are activated in ten TC-derived cell lines, leading to activated RelA/p50 and RelB/p50 NF-κB dimers. By analyzing the RNAseq data of the large papillary thyroid carcinoma (PTC) cohort from The Cancer Genome Atlas (TCGA) project, we identified a tumor progression-related NF-κB signature in BRAFV600E mutated-PTCs. That corroborated with the role of RelA and RelB in cell migration and invasion processes that we demonstrated specifically in BRAFV600E mutated-cell lines, together with their role in the control of expression of genes implicated in invasiveness (MMP1, PLAU, LCN2 and LGALS3). We also identified NF-κB-inducing kinase (NIK) as a novel actor of the constitutive activation of the NF-κB pathways in TC-derived cell lines. Finally, its implication in invasiveness and its overexpression in PTC samples make NIK a potential therapeutic target for advanced TC treatment.

Constitutive photomorphogenic protein 1 ubiquitinates interleukin-1 receptor accessory protein in human liver cancer.

Constitutive photomorphogenic protein 1 (COP1) plays a pivotal role in the development and progression of several human cancers and is reported to be upregulated in liver cancer. However, the role of COP1 in human liver cancer is unclear. We analyzed the COP1 expression in normal liver and liver cancer tissue samples using western blot and immunohistochemical analysis. We overexpressed and silenced COP1 in HepG2 and Huh7 cells and analyzed the effect on liver cancer cell proliferation. Additionally, COP1 was used as a bait to screen COP1-interacting proteins in a human cDNA library in a yeast two-hybrid screen and the results were confirmed with co-immunoprecipitation (co-IP) assays. Moreover, immunofluorescence staining was performed to assess co-localization. The protein levels of COP1 and mIL1RAcP were determined in clinical samples. COP1 was upregulated in liver cancer samples compared to that in normal tissue samples. COP1 overexpression promoted proliferation of liver cancer cells, while COP1 knockdown exerted the opposite effect. Yeast two-hybrid screen identified interleukin-1 receptor accessory protein (IL1RAP) as a potential COP1-interacting protein. Co-IP assays further confirmed that COP1 interacts with both preIL1RAP and membrane-bound form of IL1RAP (mIL1RAP). Furthermore, COP1 upregulated mIL1RAP protein levels and promoted nuclear translocation and activation of the nuclear factor kappa B (NF-κB) (p50/p65) dimer. Additionally, we demonstrated that COP1 regulated mIL1RAP expression through K63-linked polyubiquitination, suggesting that COP1 plays a role in stabilizing mIL1RAP. Finally, the protein levels of COP1 and mIL1RAcP were found to be positively correlated in clinical samples. COP1 regulates IL1RAP, which in turn results in activation of the NF-κB signaling. Our findings suggest that the COP1/IL1RAP/NF-κB axis promotes proliferation of liver cancer cells and is a potential target for the treatment of liver cancer.

The Effect of Simvastatin on the Dynamics of NF-κB-Regulated Neurodegenerative and Neuroprotective Processes in the Acute Phase of Ischemic Stroke

Statins are lipid-lowering drugs that act by inhibiting 3-hydroxy-3-methylglutaryl coenzyme A reductase, a rate-limiting enzyme in cholesterol biosynthesis. Animal studies have shown neuroprotective effects of statins in cerebral stroke. However, the underlying mechanisms are not fully understood. The nuclear factor-kappa B (NF-κB) transcription factor is involved in the regulation of apoptosis in stroke. Different dimers of NF-κB regulate the gene expression of proteins involved in both neurodegeneration and neuroprotection. We aimed to determine whether simvastatin improves stroke outcome via inhibition of the RelA/p65-containing subunit and downregulation of stroke-induced pro-apoptotic genes or via activation of NF-κB dimers containing the c-Rel subunit and upregulation of anti-apoptotic genes during the acute stroke phase. Eighteen-month-old Wistar rats, subjected to permanent MCAO or sham surgery, were administered simvastatin (20 mg/kg b.w.) or saline for 5 days before the procedure. Stroke outcome was determined by measuring cerebral infarct and assessing motor functions. The expression of NF-κB subunits in various cell populations was investigated using immunofluorescence/confocal microscopy. RelA and c-Rel were detected by WB. The NF-κB-DNA binding activity was investigated using EMSA, and expression of Noxa, Puma, Bcl-2, and Bcl-x genes was analyzed by qRT-PCR. Results showed a 50% infarct size reduction and significant motor function improvement in the simvastatin-treated animals which correlated with a decrease in RelA and a transient increase in the c-Rel level in the nucleus, normalization of the NF-κB-DNA binding activity, and downregulation of the NF-κB-regulated genes. Our results provide new insights into the statin-mediated neuroprotective action against stroke based on NF-κB pathway inhibition.

Modeling the regulation of the Il12b locus by NF-κB dimers

Abstract The transcription factor NF-κB is composed of a family, which regulates the expression of various genes in immune and inflammatory responses. The NF-κB family consists of five members: RelA/p65, RelB, c-Rel, p50/p105, and p52/p100, which control the transcription of target genes by binding to regulatory κB sites in homo-or heterodimer forms. Unlike the widely known RelA, the mechanisms related to the regulation of target genes by c-Rel is less understood. Interleukin 12b(Il12b), which encodes the p40 subunit of IL12 and IL23, is one example of gene that depend on c-Rel for proper activation in macrophages. The differences in intrinsic binding properties of c-Rel- and RelA-containing dimers at various regulatory sites around Il12b can be a key to understand the mechanism of c-Rel-containing dimers’ role in Il12b regulation. Using c-Rel and Il12b reporter mice with fluorescence live-cell imaging, we observe the single-cell dynamics of nuclear c-Rel and Il12b gene expression in bone marrow-derived macrophages induced by NF-κB activating ligands. We analyze the live-cell imaging data and extract quantitative features of nuclear c-Rel and Il12b expression from the time-series data. For additional mechanistic insights of the Il12b gene regulation by NF-κB dimers, we propose a mathematical model of dynamic gene regulation which includes actions of RelA- and c-Rel-containing dimers at κB sites proximal and distal to the Il12b locus. Our combined experimental and mathematical modeling approach will reveal quantitative insights that may provide a more comprehensive understanding of regulatory mechanisms underlying Il12b gene expression employed by NF-κB dimers.

The NF-κB multidimer system model: A knowledge base to explore diverse biological contexts.

The nuclear factor κB (NF-κB) system is critical for various biological functions in numerous cell types, including the inflammatory response, cell proliferation, survival, differentiation, and pathogenic responses. Each cell type is characterized by a subset of 15 NF-κB dimers whose activity is regulated in a stimulus-responsive manner. Numerous studies have produced different mathematical models that account for cell type-specific NF-κB activities. However, whereas the concentrations or abundances of NF-κB subunits may differ between cell types, the biochemical interactions that constitute the NF-κB signaling system do not. Here, we synthesized a consensus mathematical model of the NF-κB multidimer system, which could account for the cell type-specific repertoires of NF-κB dimers and their cell type-specific activation and cross-talk. Our review demonstrates that these distinct cell type-specific properties of NF-κB signaling can be explained largely as emergent effects of the cell type-specific expression of NF-κB monomers. The consensus systems model represents a knowledge base that may be used to gain insights into the control and function of NF-κB in diverse physiological and pathological scenarios and that describes a path for generating similar regulatory knowledge bases for other pleiotropic signaling systems.

Dry tobacco leaves: an in vivo and in silico approach to the consequences of occupational exposure.

Exposure of tobacco workers handling dried tobacco leaves has been linked to an increased risk of toxicity and respiratory illness due to the presence of nicotine and other chemicals. This study aimed to evaluate the DNA damage caused by the exposure of tobacco growers during the dry leaf classification process and the relation to cellular mechanisms. A total of 86 individuals participated in the study, divided into a group exposed to dry tobacco (n = 44) and a control group (n = 42). Genotoxicity was evaluated using the alkaline comet assay and lymphocyte micronucleus (MN) assay (CBMN-Cyt), and measurement of telomere length. The levels of oxidative and nitrosative stress were evaluated through the formation of thiobarbituric acid reactive species, and nitric oxide levels, respectively. The inorganic elements were measured in the samples using particle-induced X-ray emission method. The combination of variables was demonstrated through principal component analysis and the interactions were expanded through systems biology. Comet assay, MN, death cells, thiobarbituric acid reactive species, and nitrosative stress showed a significant increase for all exposed groups in relation to the control. Telomere length showed a significant decrease for exposed women and total exposed group in relation to men and control groups, respectively. Bromine (Br) and rubidium (Rb) in the exposed group presented higher levels than control groups. Correlations between nitrate and apoptosis; Br and MN and necrosis; and Rb and telomeres; besides age and DNA damage and death cells were observed. The systems biology analysis demonstrated that tobacco elements can increase the nuclear translocation of NFKB dimers inducing HDAC2 expression, which, associated with BRCA1 protein, can potentially repress transcription of genes that promote DNA repair. Dry tobacco workers exposed to dry leaves and their different agents showed DNA damage by different mechanisms, including redox imbalance.

Resvega, a Nutraceutical Preparation, Affects NFκB Pathway and Prolongs the Anti-VEGF Effect of Bevacizumab in Undifferentiated ARPE-19 Retina Cells.

Age-related macular degeneration (AMD) is an irreversible chronic degenerative pathology that affects the retina. Despite therapeutic advances thanks to the use of anti-vascular endothelial growth factor (VEGF) agents, resistance mechanisms have been found to accentuate the visual deficit. In the present study, we explored whether a nutraceutical formulation composed of omega-3 fatty acids and resveratrol, called Resvega®, was able to disrupt VEGF-A secretion in human ARPE-19 retina cells. We found that Resvega® inhibits VEGF-A secretion through decreases in both the PI3K-AKT-mTOR and NFκB signaling pathways. In NFκB signaling pathways, Resvega® inhibits the phosphorylation of the inhibitor of NFκB, IκB, which can bind NFκB dimers and sequester them in the cytoplasm. Thus, the NFκB subunits cannot migrate to the nucleus where they normally bind and stimulate the transcription of target genes such as VEGF-A. The IκB kinase complex (IKK) is also affected by Resvega® since the nutraceutical formulation decreases both IKKα and IKKβ subunits and the IKKγ subunit which is required for the stimulation of IKK. Very interestingly, we highlight that Resvega® could prolong the anti-angiogenic effect of Avastin®, which is an anti-VEGF agent typically used in clinical practice. Our results suggest that Resvega® may have potential interest as nutritional supplementation against AMD.

Atypical IκB Bcl3 enhances the generation of the NF-κB p52 homodimer

The NF-κB family of dimeric transcription factors regulate diverse biological functions. Their cellular expression profiles differ, which lead to different concentrations in different cell/tissue types. Although the activation mechanisms of different NF-κB dimers have been widely investigated, there is limited information on specific NF-κB dimers’ formation. The NF-κB p52:p52 homodimer regulates an important subset of target genes in cancer cells; however, the molecular mechanism of the generation of this specific homodimer remains unclear. Our study has revealed that the atypical IκB protein, Bcl3, plays an essential role in enhancing the p52:p52 homodimer population which is a unique mechanism to p52 within the NF-κB family. p52 was shown to heterodimerize with four other NF-κB subunits (RelA, RelB, cRel, and p50); all heterodimers, except p52:p50, are significantly more stable than the p52:p52 homodimer. Bcl3 is able to compete with all other NF-κB subunits in cells for efficient p52:p52 homodimer formation which consequently leads to the upregulation of target genes that are involved in cell proliferation, migration, and inflammation, which explain why aberrant activation of Bcl3 and p52 leads to cancer.

An intrinsically disordered transcription activation domain increases the DNA binding affinity and reduces the specificity of NFκB p50/RelA

Many transcription factors contain intrinsically disordered transcription activation domains (TADs), which mediate interactions with coactivators to activate transcription. Historically, DNA-binding domains and TADs have been considered as modular units, but recent studies have shown that TADs can influence DNA binding. Whether these results can be generalized to more TADs is not clear. Here, we biophysically characterized the NFκB p50/RelA heterodimer including the RelA TAD and investigated the TAD’s influence on NFκB–DNA interactions. In solution, we show the RelA TAD is disordered but compact, with helical tendency in two regions that interact with coactivators. We determined that the presence of the TAD increased the stoichiometry of NFκB–DNA complexes containing promoter DNA sequences with tandem κB recognition motifs by promoting the binding of NFκB dimers in excess of the number of κB sites. In addition, we measured the binding affinity of p50/RelA for DNA containing tandem κB sites and single κB sites. While the presence of the TAD enhanced the binding affinity of p50/RelA for all κB sequences tested, it also increased the affinity for nonspecific DNA sequences by over 10-fold, leading to an overall decrease in specificity for κB DNA sequences. In contrast, previous studies have generally reported that TADs decrease DNA-binding affinity and increase sequence specificity. Our results reveal a novel function of the RelA TAD in promoting binding to nonconsensus DNA, which sheds light on previous observations of extensive nonconsensus DNA binding by NFκB in vivo in response to strong inflammatory signals.

Development of (4-Phenylamino)quinazoline Alkylthiourea Derivatives as Novel NF-κB Inhibitors.

For many inflammatory diseases, new effective drugs with fewer side effects are needed. While it appears promising to target the activation of the central pro-inflammatory transcription factor NF-κB, many previously discovered agents suffered from cytotoxicity. In this study, new alkylthiourea quinazoline derivatives were developed that selectively inhibit the activation of NF-κB in macrophage-like THP−1 cells while showing low general cytotoxicity. One of the best compounds, 19, strongly inhibited the production of IL-6 (IC50 = 0.84 µM) and, less potently, of TNFα (IC50 = 4.0 µM); in comparison, the reference compound, caffeic acid phenethyl ester (CAPE), showed IC50s of 1.1 and 11.4 µM, respectively. Interestingly, 19 was found to block the translocation of the NF-κB dimer to the nucleus, although its release from the IκB complex was unaffected. Furthermore, 19 suppressed the phosphorylation of NF-κB-p65 at Ser468 but not at Ser536; however, 19 did not inhibit any kinase involved in NF-κB activation. The only partial suppression of p65 phosphorylation might be associated with fewer side effects. Since several compounds selectively induced cell death in activated macrophage-like THP−1 cells, they might be particularly effective in various inflammatory diseases that are exacerbated by excess activated macrophages, such as arteriosclerosis and autoimmune diseases.

The role of the NF-kappa B transcription activation domain in specific and nonspecific DNA interactions

The transcription factor NFκB activates genes that mediate inflammation, the immune response, and cell death and survival via interactions with both DNA and co-activator proteins. NFκB dimers (p50/RelA) contain a natively folded Rel-homology domain (RHD) and an intrinsically disordered transcription activation domain (TAD). Historically, the RHD was believed to mediate DNA binding whereas the TAD was responsible for the protein-protein interactions necessary for transcription activation. Nearly all previous in vitro studies have used truncated NFκB constructs and studied the roles of these domains in isolation.

The canonical and non-canonical NF-κB pathways and their crosstalk: A comparative study based on Petri nets

NF-κB is a protein complex that occurs in almost all animal cell types. It regulates the cellular immune responses to stimuli in the nucleus. Dysregulation of NF-κB can cause severe diseases like chronic inflammation, autoimmune diseases or cancer. We modeled the two major pathways leading from the external cellular stimulation of the CD40 receptor to the nuclear translocation of NF-κB dimers, the canonical and non-canonical pathway. Based on literature data, we developed two Petri net models describing these pathways. In a third Petri net, we combined the two models, introducing crosstalk specific in CD40L-stimulated B cells. In terms of structural properties, we checked the Petri nets for their consistency and correctness. To explore differences and similarities, we compared structural properties and the simulation behavior of the models. The non-canonical NF-κB pathway exhibited a more diverse regulation than the canonical pathway. Applying in silico knockout analyses, we were able to quantify the relevance of individual biochemical processes. We predicted interrelationships, e.g., between the synthesis of the protein NF-κB-inducing kinase and the processing of the precursor protein p100. The activation of the transcription factors, p50-RelA and p52-RelB, was affected by most of the knockouts. The results of the in silico knockout were in accordance with experimental studies. The Petri net models provide a basis for further analyses and could be extended to include gene expression, additional pathways, molecular processes, and kinetic data.

An epithelial Nfkb2 pathway exacerbates intestinal inflammation by supplementing latent RelA dimers to the canonical NF-κB module

Aberrant inflammation, such as that associated with inflammatory bowel disease (IBD), is fueled by the inordinate activity of RelA/NF-κB factors. As such, the canonical NF-κB module mediates controlled nuclear activation of RelA dimers from the latent cytoplasmic complexes. What provokes pathological RelA activity in the colitogenic gut remains unclear. The noncanonical NF-κB pathway typically promotes immune organogenesis involving Nfkb2 gene products. Because NF-κB pathways are intertwined, we asked whether noncanonical signaling aggravated inflammatory RelA activity. Our investigation revealed frequent engagement of the noncanonical pathway in human IBD. In a mouse model of experimental colitis, we established that Nfkb2-mediated regulations escalated the RelA-driven proinflammatory gene response in intestinal epithelial cells, exacerbating the infiltration of inflammatory cells and colon pathologies. Our mechanistic studies clarified that cell-autonomous Nfkb2 signaling supplemented latent NF-κB dimers, leading to a hyperactive canonical RelA response in the inflamed colon. In sum, the regulation of latent NF-κB dimers appears to link noncanonical Nfkb2 signaling to RelA-driven inflammatory pathologies and may provide for therapeutic targets.

Insights into the NF-κB-DNA Interaction through NMR Spectroscopy.

Transcription factors bind specifically to their target elements in the genome, eliciting specific gene expression programs. The nuclear factor-κB (NF-κB) system is a family of proteins comprising inducible transcription activators, which play a critical role in inflammation and cancer. The NF-κB members function as dimers with each monomeric unit binding the κB-DNA. Despite the available structures of the various NF-κB dimers in complex with the DNA, the structural features of these dimers in the nucleic acid-free form are not well-characterized. Using solution NMR spectroscopy, we characterize the structural features of 73.1 kDa p50 subunit of the NF-κB homodimer in the DNA-free form and compare it with the κB DNA-bound form of the protein. The study further reveals that in the nucleic acid-free form, the two constituent domains of p50, the N-terminal and the dimerization domains, are structurally independent of each other. However, in a complex with the κB DNA, both the domains of p50 act as a single unit. The study also provides insights into the mechanism of κB DNA recognition by the p50 subunit of NF-κB.

Attribution of NF-κB Activity to CHUK/IKKα-Involved Carcinogenesis.

Studies analyzing human cancer genome sequences and genetically modified mouse models have extensively expanded our understanding of human tumorigenesis, even challenging or reversing the dogma of certain genes as originally characterized by in vitro studies. Inhibitor-κB kinase α (IKKα), which is encoded by the conserved helix-loop-helix ubiquitous kinase (CHUK) gene, is first identified as a serine/threonine protein kinase in the inhibitor-κB kinase complex (IKK), which is composed of IKKα, IKKβ, and IKKγ (NEMO). IKK phosphorylates serine residues 32 and 36 of IκBα, a nuclear factor-κB (NF-κB) inhibitor, to induce IκBα protein degradation, resulting in the nuclear translocation of NF-κB dimers that function as transcriptional factors to regulate immunity, infection, lymphoid organ/cell development, cell death/growth, and tumorigenesis. NF-κB and IKK are broadly and differentially expressed in the cells of our body. For a long time, the idea that the IKK complex acts as a direct upstream activator of NF-κB in carcinogenesis has been predominately accepted in the field. Surprisingly, IKKα has emerged as a novel suppressor for skin, lung, esophageal, and nasopharyngeal squamous cell carcinoma, as well as lung and pancreatic adenocarcinoma (ADC). Thus, Ikkα loss is a tumor driver in mice. On the other hand, lacking the RANKL/RANK/IKKα pathway impairs mammary gland development and attenuates oncogene- and chemical carcinogen-induced breast and prostate tumorigenesis and metastasis. In general, NF-κB activation leads one of the major inflammatory pathways and stimulates tumorigenesis. Since IKKα and NF-κB play significant roles in human health, revealing the interplay between them greatly benefits the diagnosis, treatment, and prevention of human cancer. In this review, we discuss the intriguing attribution of NF-κB to CHUK/IKKα-involved carcinogenesis.