Agent-Based Modeling of Complex Molecular Systems.

The seamless integration of laboratory experiments and detailed computational modeling provides an exciting route to uncovering many new insights into complex biological processes. In particular, the development of agent-based modeling using supercomputers has provided new opportunities for highly detailed, validated simulations that provide the researcher with greater understanding of these processes and new directions for investigation. This chapter examines some of the principles behind the powerful computational framework FLAME and its application in a number of different areas with a more detailed look at a particular signaling example involving the NF-κB cascade.

Distinct Binding Interactions of αβ-Integrin and Proteoglycans with Fibronectin.

Dynamic single-molecule force spectroscopy was performed to monitor the unbinding of fibronectin with the proteoglycans syndecan-4 (SDC4) and decorin and to compare this with the unbinding characteristics of αβ-integrin. A single energy barrier was sufficient to describe the unbinding of both SDC4 and decorin from fibronectin, whereas two barriers were observed for the dissociation of αβ-integrin from fibronectin. The outer (high-affinity) barriers in the interactions of fibronectin with αβ-integrin and SDC4 are characterized by larger barrier heights and widths and slower dissociation rates than those of the inner (low-affinity) barriers in the interactions of fibronectin with αβ-integrin and decorin.

The IL-1RI Co-Receptor TILRR ( Isoform 2) Controls Aberrant Inflammatory Responses and Development of Vascular Disease.

Expression of the interleukin-1 receptor type I (IL-1RI) co-receptor Toll-like and interleukin-1 receptor regulator (TILRR) is significantly increased in blood monocytes following myocardial infarction and in the atherosclerotic plaque, whereas levels in healthy tissue are low. TILRR association with IL-1RI at these sites causes aberrant activation of inflammatory genes, which underlie progression of cardiovascular disease. The authors show that genetic deletion of TILRR or antibody blocking of TILRR function reduces development of atherosclerotic plaques.

Statistical Techniques Complement UML When Developing Domain Models of Complex Dynamical Biosystems.

Computational modelling and simulation is increasingly being used to complement traditional wet-lab techniques when investigating the mechanistic behaviours of complex biological systems. In order to ensure computational models are fit for purpose, it is essential that the abstracted view of biology captured in the computational model, is clearly and unambiguously defined within a conceptual model of the biological domain (a domain model), that acts to accurately represent the biological system and to document the functional requirements for the resultant computational model. We present a domain model of the IL-1 stimulated NF-κB signalling pathway, which unambiguously defines the spatial, temporal and stochastic requirements for our future computational model.

Reducing complexity in an agent based reaction model-Benefits and limitations of simplifications in relation to run time and system level output.

Agent based modelling is a methodology for simulating a variety of systems across a broad spectrum of fields. However, due to the complexity of the systems it is often impossible or impractical to model them at a one to one scale. In this paper we use a simple reaction rate model implemented using the FLAME framework to test the impact of common methods for reducing model complexity such as reducing scale, increasing iteration duration and reducing message overheads.

The rise in computational systems biology approaches for understanding NF-κB signaling dynamics.

A study by Cheng et al. in this issue of Science Signaling highlights the distinct single-cell signaling characteristics conferred by pathways mediated by the adaptor proteins MyD88 and TRIF in the TLR4-dependent activation of the transcription factor nuclear factor κB (NF-κB).

Computational Modelling of NF-κB Activation by IL-1RI and Its Co-Receptor TILRR, Predicts a Role for Cytoskeletal Sequestration of IκBα in Inflammatory Signalling.

The transcription factor NF-κB (nuclear factor kappa B) is activated by Toll-like receptors and controlled by mechanotransduction and changes in the cytoskeleton. In this study we combine 3-D predictive protein modelling and in vitro experiments with in silico simulations to determine the role of the cytoskeleton in regulation of NF-κB. Simulations used a comprehensive agent-based model of the NF-κB pathway, which includes the type 1 IL-1 receptor (IL-1R1) complex and signalling intermediates, as well as cytoskeletal components.

Distinct control of MyD88 adapter-dependent and Akt kinase-regulated responses by the interleukin (IL)-1RI co-receptor, TILRR.

Inflammatory responses are controlled through members of the interleukin-1 receptor (IL-1R)/Toll-like receptor superfamily. Our earlier work demonstrates that the IL-1 receptor type 1 (IL-1RI) co-receptor, Toll-like and IL-1 receptor regulator (TILRR), amplifies IL-1 activation of NF-κB and inflammatory genes. Here we show that TILRR similarly promotes IL-1-induced anti-apoptotic signals and reduces caspase-3 activity.

Bioinformatics Analysis of the FREM1 Gene-Evolutionary Development of the IL-1R1 Co-Receptor, TILRR.

The TLRs and IL-1 receptors have evolved to coordinate the innate immune response following pathogen invasion. Receptors and signalling intermediates of these systems are generally characterised by a high level of evolutionary conservation. The recently described IL-1R1 co-receptor TILRR is a transcriptional variant of the FREM1 gene.

Modelling complex biological systems using an agent-based approach.

Many of the complex systems found in biology are comprised of numerous components, where interactions between individual agents result in the emergence of structures and function, typically in a highly dynamic manner. Often these entities have limited lifetimes but their interactions both with each other and their environment can have profound biological consequences. We will demonstrate how modelling these entities, and their interactions, can lead to a new approach to experimental biology bringing new insights and a deeper understanding of biological systems.

TILRR, a novel IL-1RI co-receptor, potentiates MyD88 recruitment to control Ras-dependent amplification of NF-kappaB.

Host defense against infection is induced by Toll-like and interleukin (IL)-1 receptors, and controlled by the transcription factor NF-kappaB. Our earlier studies have shown that IL-1 activation impacts cytoskeletal structure and that IL-1 receptor (IL-1RI) function is substrate-dependent. Here we identify a novel regulatory component, TILRR, which amplifies activation of IL-1RI and coordinates IL-1-induced control with mechanotransduction.

NIK and IKKbeta interdependence in NF-kappaB signalling--flux analysis of regulation through metabolites.

Activation of the transcription factor NF-kappaB is central to control of immune and inflammatory responses. Cytokine induced activation through the classical or canonical pathway relies on degradation of the inhibitor, IkappaBalpha and regulation by the IKKbeta kinase. In addition, the NF-kappaB is activated through the NF-kappaB-inducing kinase, NIK.

Introducing spatial information into predictive NF-kappaB modelling--an agent-based approach.

Nature is governed by local interactions among lower-level sub-units, whether at the cell, organ, organism, or colony level. Adaptive system behaviour emerges via these interactions, which integrate the activity of the sub-units. To understand the system level it is necessary to understand the underlying local interactions.

Fluorescent protein reporter systems for single-cell measurements.

The unraveling of the complex dynamic networks that underlie cellular (and, by extension, tissue, organ, and organism) function requires sophisticated mathematical models and, in order to test those models, rich data sets. In addition, even in clonal populations of cells, there is a wide range of variability in cellular function at any given time, even in simple parameters such as the concentration of critical signaling components such as receptors or transcription factors. It remains a matter of conjecture as to whether this is noise, to which the system is inherently robust, or whether the cellular control network can exist in multiple discrete internal states, with indistinguishable input/output characteristics.

Feedback loops in intracellular signal processing and their potential for identifying novel signalling proteins.

Mapping of intracellular signalling networks has attracted a lot of attention during recent years. Approaches which have been used to characterise functional and physical relationships within signalling systems and to identify novel components have included bioinformatics, genomics, proteomics and genetics. However, the importance of feedback loops in the regulation of signal processing systems has often been ignored.

Formal agent-based modelling of intracellular chemical interactions.

Individual-based or agent-based models have proved useful in a variety of different biological contexts. This paper presents an agent-based model using a formal computational modelling approach to model a crucial biological system--the intracellular NF-kappaB signalling pathway. The pathway is vital to immune response regulation, and is fundamental to basic survival in a range of species.

Synergistic effect of Avemar on proinflammatory cytokine production and Ras-mediated cell activation.

Macrophages activated by lipopolysaccharide and/or phorbol esters exhibited high sensitivity to Avemar, a fermented wheat germ extract. Avemar synergized with lipopolysaccharide and PMA in the induction of the transcription of cytokine genes and release of inflammatory cytokines. At higher concentrations the preparation had a significant negative effect on the proliferation and survival of activated myeloid cell types.

Functional mapping and identification of novel regulators for the Toll/Interleukin-1 signalling network by transcription expression cloning.

Sustained inflammatory responses are central to the development and progression of chronic diseases, including atherosclerosis and rheumatoid arthritis. A large number of stimuli initiate inflammation by acting on Toll-Interleukin-1 related (TIR) domain containing receptors, producing multiple second messengers and thence large scale transcriptional changes. The mechanism by which this activation occurs is complex, and the continuing isolation of novel pathway components, mostly based on sequence similarities and protein-protein interaction studies, suggests that many elements of the TIR-initiated signalling network remain to be identified.

Distinct NF-kappaB regulation by shear stress through Ras-dependent IkappaBalpha oscillations: real-time analysis of flow-mediated activation in live cells.

NF-kappaB, a transcription factor central to inflammatory regulation during development of atherosclerosis, is activated by soluble mediators and through biomechanical inputs such as flow-mediated shear- stress. To investigate the molecular mechanisms underlying shear stress mediated signal transduction in vascular cells we have developed a system that applies flow-mediated shear stress in a controlled manner, while inserted in a confocal microscope. In combination with GFP-based methods, this allows continuous monitoring of flow induced signal transduction in live cells and in real time.

Human tribbles, a protein family controlling mitogen-activated protein kinase cascades.

Control of mitogen-activated protein kinase (MAPK) cascades is central to regulation of many cellular responses. We describe here human tribbles homologues (Htrbs) that control MAPK activity. MAPK kinases interact with Trbs and regulate their steady state levels.

Hunting for genes by functional screens.

Advances in high throughput sequencing technologies have led to an explosion of sequence information available for today's researchers. Efforts in the emerging next phase of the genomic era are focusing on the assignment of function to genes uncovered by genome sequencing programs. The main approaches include high throughput mutagenesis, predictions based on homology in primary sequence, microarray and proteomics.

Ultrasound-enhanced transgene expression in vascular cells is not dependent upon cavitation-induced free radicals.

Although acoustic cavitation is clearly important in ultrasound (US)-enhanced gene delivery (UEGD), the relative importance of mechanical and sonochemical (free radical) bioeffects remains unclear, as does the mechanism of gene delivery at the cellular level. Porcine vascular smooth muscle cells (VSMC) were transfected with luciferase or green fluorescent protein (GFP) plasmid +/- pulsed 956 kHz US (2.0 mechanical index (MI), 128 W cm(-2) spatial peak pulse average intensity, ISPPA) for 60 s, in the presence or absence of 20 mM cysteamine or N-acetyl-L-cysteine.

RelA control of IkappaBalpha phosphorylation: a positive feedback loop for high affinity NF-kappaB complexes.

NF-kappaB-IkappaB complex formation regulates the level and specificity of NF-kappaB activity. Quantitative analyses showed that RelA-NF-kappaB-induced IkappaBalpha binding is regulated through inhibitor retention and phosphorylation. RelA caused an increase in IkappaBalpha phosphorylation and in degradation, which was enhanced monotonically with inhibitor concentration.

Regulation of nuclear translocation of nuclear factor-kappaB relA: evidence for complex dynamics at the single-cell level.

We have analysed activation of nuclear factor-kappaB (NF-kappaB) in response to interleukin-1 (IL-1) in human fibroblasts by tracking intracellular distribution and levels of endogenous relA, NF-kappaB1 and inhibitor of kappaB (I-kappaB) alpha using semi-quantitative confocal microscopy. Nuclear translocation of endogenous relA correlated with I-kappaBalpha degradation during stimulation with IL-1, whereas no effects were seen on levels or localization of NF-kappaB1. During pathway activation, relA was transported up a concentration gradient, resulting in a 3-4-fold increase in nuclear levels, but without any significant decrease in cytoplasmic concentration.

PDGF enhancement of IL-1 receptor levels in smooth muscle cells involves induction of an attachment-regulated, heparan sulfate binding site (IL-1RIII).

This study shows that increase in IL-1 receptor levels by platelet derived growth factor (PDGF) involves an enhancement of a matrix-dependent, low-affinity receptor that constitutes a heparan sulfate. Fibronectin attachment caused pronounced alterations in IL-1 receptor function in smooth muscle cells, involving a pronounced increase in cell surface binding from an average of 2,000 up to approximately 8,000 receptors/cell and an increase in affinity (K(a)) of the type I receptor from 1.8 +/- 0.