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Mammalian inflammatory signaling, for which NF-kappaB is a principal transcription factor, is an exquisite example of how cellular signaling pathways can be regulated to produce different yet specific responses to different inflammatory insults. Mathematical models, tightly linked to experiment, have been instrumental in unraveling the forms of regulation in NF-kappaB signaling and their underlying molecular mechanisms. Our initial model of the IkappaB-NF-kappaB signaling module highlighted the role of negative feedback in the control of NF-kappaB temporal dynamics and gene expression. Subsequent studies sparked by this work have helped to characterize additional feedback loops, the input-output behavior of the module, crosstalk between multiple NF-kappaB-activating pathways, and NF-kappaB oscillations. We anticipate that computational techniques will enable further progress in the NF-kappaB field, and the signal transduction field in general, and we discuss potential upcoming developments.

Mammalian inflammatory signaling, for which NF-kappaB is a principal transcription factor, is an exquisite example of how cellular signaling pathways can be regulated to produce different yet specific responses to different inflammatory insults. Mathematical models, tightly linked to experiment, have been instrumental in unraveling the forms of regulation in NF-kappaB signaling and their underlying molecular mechanisms. Our initial model of the IkappaB-NF-kappaB signaling module highlighted the role of negative feedback in the control of NF-kappaB temporal dynamics and gene expression. Subsequent studies sparked by this work have helped to characterize additional feedback loops, the input-output behavior of the module, crosstalk between multiple NF-kappaB-activating pathways, and NF-kappaB oscillations. We anticipate that computational techniques will enable further progress in the NF-kappaB field, and the signal transduction field in general, and we discuss potential upcoming developments.

Neurons in the central nervous system (CNS) fail to regenerate axons after injuries due to the diminished intrinsic axon growth capacity of mature neurons and the hostile extrinsic environment composed of a milieu of inhibitory factors. Recent studies revealed that targeting a particular group of extracellular inhibitory factors is insufficient to trigger long-distance axon regeneration. Instead of antagonizing the growing list of impediments, tackling a common target that mediates axon growth inhibition offers an alternative strategy to promote axon regeneration. Neuronal growth cone, the machinery that derives axon extension, is the final converging target of most, if not all, growth impediments in the CNS. In this study, we aim to promote axon growth by directly targeting the growth cone. Here we report that pharmacological inhibition or genetic silencing of nonmuscle myosin II (NMII) markedly accelerates axon growth over permissive and nonpermissive substrates, including major CNS inhibitors such as chondroitin sulfate proteoglycans and myelin-associated inhibitors. We find that NMII inhibition leads to the reorganization of both actin and microtubules (MTs) in the growth cone, resulting in MT reorganization that allows rapid axon extension over inhibitory substrates. In addition to enhancing axon extension, we show that local blockade of NMII activity in axons is sufficient to trigger axons to grow across the permissive-inhibitory border. Together, our study proposes NMII and growth cone cytoskeletal components as effective targets for promoting axon regeneration. [ABSTRACT FROM AUTHOR]

Loss of imprinting (LOl) of the insulin-like growth factor-Il gene (IGF2), leading to abnormal activation of the normally silent maternal allele, is a common human epigenetic population variant associated with a 5-fold increased frequency of colorectal neoplasia. Here, we show first that LOl leads specifically to increased expression of proliferation-related genes in mouse intestinal crypts. Surprisingly, LOl(+) mice also have enhanced sensitivity to IGF-ll signaling, not simply increased IGF-ll levels, because in vivo blockade with NVP-AEW541, a specific inhibitor of the IGF-ll signaling receptor, showed reduction of proliferation-related gene expression to levels half that seen in LOl(-) mice. Signal transduction assays in microfluidic chips confirmed this enhanced sensitivity with marked augmentation of Akt/PKB signaling in LOl(+) cells at low doses of IGF-ll, which was reduced in the presence of the inhibitor to levels below those found in LOl(-) cells, and was associated with increased expression of the IGF1 and insulin receptor genes. We exploited this increased IGF-ll sensitivity to develop an in vivo chemopreventive strategy using the azoxymethane (AOM) mutagenesis model. LOl(-i-) mice treated with AOM showed a 60% increase in premalignant aberrant crypt foci (ACF) formation over LOl(-) mice. In viva IGF-ll blockade with NVP-AEW541 abrogated this effect, reducing ACF to a level 30% lower even than found in exposed LOl(-) mice. Thus, LOl increases cancer risk in a counterintuitive way, by increasing the sensitivity of the IGF-ll signaling pathway itself, providing a previously undescribed epigenetic chemoprevention strategy in which cells with LOl are "IGF-ll addicted" and undergo reduced tumorigenesis in the colon upon IGF-ll pathway blockade. [ABSTRACT FROM AUTHOR]

Many cellular behaviors cannot be completely captured or appropriately described at the cell population level. Noise induced by stochastic chemical reactions, spatially polarized signaling networks, and heterogeneous cell-cell communication are among the many phenomena that require fine-grained analysis. Accordingly, the mathematical models used to describe such systems must be capable of single cell or subcellular resolution. Here, we review techniques for modeling single cells, including models of stochastic chemical kinetics, spatially heterogeneous intracellular signaling, and spatial stochastic systems. We also briefly discuss applications of each type of model.

Paradichlorobenzene (PDB) is a common household deodorant and pesticide found in room deodorizers, toilet bowl fresheners, and some mothballs. Although human exposure to the compound is generally limited and harmless, PDB in larger doses can produce neurotoxic effects, including a chemical "high" similar to that seen with inhalants such as toluene. Although rare, frank addiction to PDB has been reported, and, in such cases, has been associated with gait ataxia, tremor, dysarthria, limb weakness, and bradyphrenia, in various combinations. In such cases, the adverse neurologic consequences have been presumed to result from a direct toxic effect of this small, organic molecule. We report a case of chronic mothball ingestion where profound encephalopathy with cognitive, pyramidal, extrapyramidal, and cerebellar features appears to have been largely the result of PDB withdrawal, rather than direct toxicity. This case raises important questions about the mechanism of PDB neurotoxicity and possible treatment options for PDB-addicted patients. We propose that in cases with clear clinical deterioration after abstinence, readministration and gradual taper of PDB might be considered a therapeutic option.

Dynamic properties of signaling pathways control their behavior and function. We undertook an iterative computational and experimental investigation of the dynamic properties of tumor necrosis factor (TNF)alpha-mediated activation of the transcription factor NF-kappaB. Surprisingly, we found that the temporal profile of the NF-kappaB activity is invariant to the TNFalpha dose. We reverse engineered a computational model of the signaling pathway to identify mechanisms that impart this important response characteristic, thus predicting that the IKK activity profile must transiently peak at all TNFalpha doses to generate the observed NF-kappaB dynamics. Experimental confirmation of this prediction emphasizes the importance of mechanisms that rapidly down-regulate IKK following TNFalpha activation. A refined computational model further revealed signaling characteristics that ensure robust TNFalpha-mediated cell-cell communication over considerable distances, allowing for fidelity of cellular inflammatory responses in infected tissue.

Lac repressor (LacI) binds two operator DNA sites, looping the intervening DNA. DNA molecules containing two lac operators bracketing a sequence-directed bend were previously shown to form hyperstable LacI-looped complexes. Biochemical studies suggested that orienting the operators outward relative to the bend direction (in construct 9C14) stabilizes a positively supercoiled closed form, with a V-shaped LacI, but that the most stable loop construct (11C12) is a more open form. Here, fluorescence resonance energy transfer (FRET) is measured on DNA loops, between fluorescein and TAMRA attached near the two operators, approximately 130 basepairs apart. For 9C14, efficient LacI-induced energy transfer ( approximately 74% based on donor quenching) confirms that the designed DNA shape can force the looped complex into a closed form. From enhanced acceptor emission, correcting for observed donor-dependent quenching of acceptor fluorescence, approximately 52% transfer was observed. Time-resolved FRET suggests that this complex exists in both closed- and open form populations. Less efficient transfer, approximately 10%, was detected for DNA-LacI sandwiches and 11C12-LacI, consistent with an open form loop. This demonstration of long-range FRET in large DNA loops confirms that appropriate DNA design can control loop geometry. LacI flexibility may allow it to maintain looping with other proteins bound or under different intracellular conditions.