Cell differentiation requires the ability to detect and respond appropriately to a variety of extracellular signals. Here we investigate a differentiation switch induced by changes in the concentration of a single stimulus. Yeast cells exposed to high doses of mating pheromone undergo cell division arrest. Cells at intermediate doses become elongated and divide in the direction of a pheromone gradient (chemotropic growth). Either of the pheromone-responsive MAP kinases, Fus3 and Kss1, promotes cell elongation, but only Fus3 promotes chemotropic growth. Whereas Kss1 is activated rapidly and with a graded dose-response profile, Fus3 is activated slowly and exhibits a steeper dose-response relationship (ultrasensitivity). Fus3 activity requires the scaffold protein Ste5; when binding to Ste5 is abrogated, Fus3 behaves like Kss1, and the cells no longer respond to a gradient or mate efficiently with distant partners. We propose that scaffold proteins serve to modulate the temporal and dose-response behavior of the MAP kinase.

SummaryThe health of organisms and cells depends on appropriate responses to diverse internal and external cues, stimuli, or challenges, such as changes in hormone or cytokine levels, or exposure to a pathogen. Cellular responses must be tailored to the identity and intensity of the stimulus and therefore intracellular signals must carry information about both. However, signaling mediators often form intricate networks that react to multiple stimuli yet manage to produce stimulus-specific responses. The multi-functionality ("functional pleiotropism") of signaling nodes suggests that biological networks have evolved ways of passing physiologically relevant stimulus information through shared channels. Increasing evidence supports the notion that this is achieved in part through temporal regulation of signaling mediators' activities. The present challenge is to identify the features of temporal activity profile that represent information about a given stimulus and understand how cells read the temporal codes to control their responses. What's in a signal?The striking temporal control of signaling mediators' activity revealed by recent studies suggest that dynamics (understood as the spatiotemporal patterns of activity) are an intrinsic part of a signal [1,2] that, together with the chemical identity of the mediator, carries information about the stimulus. Pronounced temporal control of signaling mediators is particularly pervasive in stress [3,4] and immune responses [5,6], triggered by stimuli that have a well-defined starting point, at least in cell culture studies.In general, signal dynamics often depend on one or more properties of the stimulus, such as its identity, its amplitude, rate of increase, duration, or rate of decrease. Particular stimulus features do not necessarily translate into equivalent signal features; for example, stimulus amplitude may determine signal duration or vice versa. Signals are often classified as amplitude or frequency modulated (AM or FM). Signals are thought to be amplitude modulated when their amplitude, duration, or a combination of both is modulated by the stimulus. Frequency modulation is ascribed to particular cases when the frequency of a periodic signal is a function of the stimulus. In this review, we advance the view that to understand temporal signaling codes, we must focus on not only on the encoding mechanism that relates stimulus to signal, but also on the decoding mechanism that relates signal to cellular response.

We perform a systematic analysis of mechanisms of feedback regulation that underlie short-term adaptation in intracellular signaling systems. Upon receiving an external cue, these systems generate a transient response that quickly returns to basal levels even if the stimulus persists. Signaling pathways capable of short-term adaptation are found in systems as diverse as the high osmolarity response of yeast, gradient sensing in Dictyostelium, and the cytokine response in vertebrates. Using mathematical analysis and computational experiments, we compare different feedback architectures in terms of response amplitude and duration, ability to adapt, and response to variable stimulus levels. Our analysis reveals three important features of these systems: 1), multiple step signaling cascades improve sensitivity to low doses by an effect distinct from signal amplification; 2), some feedback architectures act as signal transducers converting stimulus strength into response duration; and 3), feedback deactivation acts as a dose-dependent switch between transient and sustained responses. Finally, we present characteristic features for each form of feedback regulation that can aid in their identification.

Highly networked signaling hubs are often associated with disease, but targeting them pharmacologically has largely been unsuccessful in the clinic because of their functional pleiotropy. Motivated by the hypothesis that a dynamical signaling code confers functional specificity, we investigated whether dynamical features may be targeted pharmacologically to achieve therapeutic specificity. With a virtual screen we identified combinations of signaling hub topologies and dynamic signal profiles that are amenable to selective inhibition. Mathematical analysis revealed principles that may guide stimulus-specific inhibition of signaling hubs, even in the absence of detailed mathematical models. Using the NFκB signaling module as a test bed, we identified perturbations that selectively affect the response to cytokines or pathogen components. Together, our results demonstrate that the dynamics of signaling may serve as a pharmacological target, and we reveal principles that delineate the opportunities and constraints of developing stimulus-specific therapeutic agents aimed at pleiotropic signaling hubs.

A structural and functional study delineates how the interaction between NF-κB subunit RelA and co-activator CBP/p300 helps drive transcription of NF-κB target genes.

Summary:  Mathematical modeling has proved to be a critically important approach in the study of many complex networks and dynamic systems in physics, engineering, chemistry, and biology. The nuclear factor κB (NF‐κB) system consists of more than 50 proteins and protein complexes and is both a highly networked and dynamic system. To date, mathematical modeling has only addressed a small fraction of the molecular species and their regulation, but when employed in conjunction with experimental analysis has already led to important insights. Here, we provide a personal account of studying how the NF‐κB signaling system functions using mathematical descriptions of the molecular mechanisms. We focus on the insights gained about some of the key regulatory components: the control of the steady state, the signaling dynamics, and signaling crosstalk. We also discuss the biological relevance of these regulatory systems properties.

A switchlike response in nuclear factor-κB (NF-κB) activity implies the existence of a threshold in the NF-κB signaling module. We show that the CARD-containing MAGUK protein 1 (CARMA1, also called CARD11)-TAK1 (MAP3K7)-inhibitor of NF-κB (IκB) kinase-β (IKKβ) module is a switch mechanism for NF-κB activation in B cell receptor (BCR) signaling. Experimental and mathematical modeling analyses showed that IKK activity is regulated by positive feedback from IKKβ to TAK1, generating a steep dose response to BCR stimulation. Mutation of the scaffolding protein CARMA1 at serine-578, an IKKβ target, abrogated not only late TAK1 activity, but also the switchlike activation of NF-κB in single cells, suggesting that phosphorylation of this residue accounts for the feedback.

These findings reveal a novel phosphorylation-dependent feedback loop leading to diminished cellular responses to an osmotic-stress stimulus.