Encoding NF-κB temporal control in response to TNF: distinct roles for the negative regulators IκBα and A20

TNF-induced NF-κB activity shows complex temporal regulation whose different phases lead to distinct gene expression programs. Combining experimental studies and mathematical modeling, we identify two temporal amplification steps—one determined by the obligate negative feedback regulator IκBα—that define the duration of the first phase of NF-κB activity. The second phase is defined by A20, whose inducible expression provides for a rheostat function by which other inflammatory stimuli can regulate TNF responses. Our results delineate the nonredundant functions implied by the knockout phenotypes of iκbα and a20, and identify the latter as a signaling cross-talk mediator controlling inflammatory and developmental responses

Altered inflammasome activation in neonatal encephalopathy persists in childhood

Neonatal encephalopathy (NE) is characterized by altered neurological function in term infants and inflammation plays an important pathophysiological role. Inflammatory cytokines interleukin (IL)-1 beta, IL-1ra and IL-18 are activated by the nucleotide-binding and oligomerization domain (NOD)-, leucine-rich repeat domain (LRR)- and NOD-like receptor protein 3 (NLRP3) inflammasome; furthermore, we aimed to examine the role of the inflammasome multiprotein complex involved in proinflammatory responses from the newborn period to childhood in NE. Cytokine concentrations were measured by multiplex enzyme-linked immunosorbent assay (ELISA) in neonates and children with NE in the absence or presence of lipopolysaccharide (LPS) endotoxin. We then investigated expression of the NLRP3 inflammasome genes, NLRP3, IL-1 beta and ASC by polymerase chain reaction (PCR). Serum samples from 40 NE patients at days 1 and 3 of the first week of life and in 37 patients at age 4-7 years were analysed. An increase in serum IL-1ra and IL-18 in neonates with NE on days 1 and 3 was observed compared to neonatal controls. IL-1ra in NE was decreased to normal levels at school age, whereas serum IL-18 in NE was even higher at school age compared to school age controls and NE in the first week of life. Percentage of LPS response was higher in newborns compared to school-age NE. NLRP3 and IL-1 beta gene expression were up-regulated in the presence of LPS in NE neonates and NLRP3 gene expression remained up-regulated at school age in NE patients compared to controls. Increased inflammasome activation in the first day of life in NE persists in childhood, and may increase the window for therapeutic intervention

Homeostatic control of IkappaB metabolism determines NF- kappaB responsiveness

Cellular signal transduction begins with an input signal that activates a pathway or network of signaling proteins to exert a specific output or change in cellular behavior. A useful approach to understanding signal transduction and mapping signaling pathways has been to examine changes to specific proteins, either in their expression, interaction with other proteins, post-translation modifications, or enzymatic activities, in cells after application of the input. As useful as this approach may be, it does not answer how the pathway may be regulated in the absence of an input signal, nor does it explain how the output may be more or less responsive to an input depending on the homeostatic regulation within the particular network. Tight regulation of the NF-[kappa]B transcription factor is mediated by the coordinated control of the synthesis and degradation of the inhibitor of NF-[kappa]B proteins, the I[kappa]Bs. My dissertation takes a mechanistic approach to understanding how steady state I[kappa]B synthesis and degradation affect constitutive NF-[kappa]B activity and NF-[kappa]B responses to inflammatory and metabolic stress signals. Biochemistry, genetics, and molecular biology, are used in conjunction with mathematical modeling to explore how homeostatic mechanisms determine UV-induced NF-[kappa]B activity and how type I and type II interferons alter NF-[kappa]B responsiveness to inflammatory signals. This work suggests that the targeting of the steady-state reactions of a network may be important consideration when designing therapeutics to specifically target certain aspects of a signaling pathwa

The Regulatory Logic of the NF-κB Signaling System

NF-κB refers to multiple dimers of Rel homology domain (RHD) containing polypeptides, which are controlled by a stimulus-responsive signaling system that mediates the physiological responses to inflammatory intercellular cytokines, pathogen exposure, and developmental signals. The NF-κB signaling system operates on transient or short timescales, relevant to inflammation and immune responses, and on longer-term timescales relevant to cell differentiation and organ formation. Here, we summarize our current understanding of the kinetic mechanisms that allow for NF-κB regulation at these different timescales. We distinguish between the regulation of NF-κB dimer formation and the regulation of NF-κB activity. Given the number of regulators and reactions involved, the NF-κB signaling system is capable of integrating a multitude of signals to tune NF-κB activity, signal dose responsiveness, and dynamic control. We discuss the prevailing mechanisms that mediate signaling cross talk

NF-κB dictates the degradation pathway of IκBα

Correction to: The EMBO Journal, advance online publication 10 April 2008; doi:10.1038/emboj.2008.7

A homeostatic model of IkappaB metabolism to control constitutive NF-kappaB activity.

Cellular signal transduction pathways are usually studied following administration of an external stimulus. However, disease-associated aberrant activity of the pathway is often due to misregulation of the equilibrium state. The transcription factor NF-kappaB is typically described as being held inactive in the cytoplasm by binding its inhibitor, IkappaB, until an external stimulus triggers IkappaB degradation through an IkappaB kinase-dependent degradation pathway. Combining genetic, biochemical, and computational tools, we investigate steady-state regulation of the NF-kappaB signaling module and its impact on stimulus responsiveness. We present newly measured in vivo degradation rate constants for NF-kappaB-bound and -unbound IkappaB proteins that are critical for accurate computational predictions of steady-state IkappaB protein levels and basal NF-kappaB activity. Simulations reveal a homeostatic NF-kappaB signaling module in which differential degradation rates of free and bound pools of IkappaB represent a novel cross-regulation mechanism that imparts functional robustness to the signaling module

A homeostatic model of IκB metabolism to control constitutive NF‐κB activity

Cellular signal transduction pathways are usually studied following administration of an external stimulus. However, disease-associated aberrant activity of the pathway is often due to misregulation of the equilibrium state. The transcription factor NF-κB is typically described as being held inactive in the cytoplasm by binding its inhibitor, IκB, until an external stimulus triggers IκB degradation through an IκB kinase-dependent degradation pathway. Combining genetic, biochemical, and computational tools, we investigate steady-state regulation of the NF-κB signaling module and its impact on stimulus responsiveness. We present newly measured in vivo degradation rate constants for NF-κB-bound and -unbound IκB proteins that are critical for accurate computational predictions of steady-state IκB protein levels and basal NF-κB activity. Simulations reveal a homeostatic NF-κB signaling module in which differential degradation rates of free and bound pools of IκB represent a novel cross-regulation mechanism that imparts functional robustness to the signaling module

Youth mental health in deprived urban areas: A Delphi study on the role of the GP in early intervention.

Background: GPs, as healthcare professionals with whom young people commonly interact, have a central role in early intervention for mental health problems. However, successfully fulfilling this role is a challenge, and this is especially in deprived urban areas. Aims: To inform a complex intervention to support GPs in this important role, we aim to identify the key areas in which general practice can help address youth mental health and strategies to enhance implementation. Methods: We conducted a modified Delphi study which involved establishing an expert panel involving key stakeholders / service providers at two deprived urban areas. The group reviewed emerging literature on the topic at a series of meetings and consensus was facilitated by iterative surveys. Results: We identified 20 individual roles in which GPs could help address youth mental health address youth mental health, across five domains: 1. Prevention, Health Promotion and Access, 2. Assessment and Identification, 3. Treatment Strategies, 4.Interaction with Other Agencies/Referral, and 5. Ongoing Support. With regard to strategies to enhance implementation, we identified a further 19 interventions, across five domains: 1.Training, 2. Consultation Improvements, 3. Service-Level Changes, 4. Collaboration, and 5.Healthcare-system Changes. Conclusions: GPs have a key role in addressing youth mental health and this study highlights the key domains of this role and the key components of a complex intervention to support this role