Now showing 1 - 10 of 16
  • Publication
    Impact of measurement noise, experimental design, and estimation methods on Modular Response Analysis based network reconstruction
    Modular Response Analysis (MRA) is a method to reconstruct signalling networks from steady-state perturbation data which has frequently been used in different settings. Since these data are usually noisy due to multi-step measurement procedures and biological variability, it is important to investigate the effect of this noise onto network reconstruction. Here we present a systematic study to investigate propagation of noise from concentration measurements to network structures. Therefore, we design an in silico study of the MAPK and the p53 signalling pathways with realistic noise settings. We make use of statistical concepts and measures to evaluate accuracy and precision of individual inferred interactions and resulting network structures. Our results allow to derive clear recommendations to optimize the performance of MRA based network reconstruction: First, large perturbations are favorable in terms of accuracy even for models with non-linear steady-state response curves. Second, a single control measurement for different perturbation experiments seems to be sufficient for network reconstruction, and third, we recommend to execute the MRA workflow with the mean of different replicates for concentration measurements rather than using computationally more involved regression strategies.
      387Scopus© Citations 9
  • Publication
    PP1 initiates the dephosphorylation of MASTL, triggering mitotic exit and bistability in human cells
    (The Company of Biologists, 2016-04-01) ; ; ;
    Entry into mitosis is driven by the phosphorylation of thousands of substrates, under the master control of Cdk1. During entry into mitosis, Cdk1, in collaboration with MASTL kinase, represses the activity of the major mitotic protein phosphatases, PP1 and PP2A, thereby ensuring mitotic substrates remain phosphorylated. For cells to complete and exit mitosis, these phosphorylation events must be removed, and hence, phosphatase activity must be reactivated. This reactivation of phosphatase activity presumably requires the inhibition of MASTL; however, it is not currently understood what deactivates MASTL and how this is achieved. In this study, we identified that PP1 is associated with, and capable of partially dephosphorylating and deactivating, MASTL during mitotic exit. Using mathematical modelling, we were able to confirm that deactivation of MASTL is essential for mitotic exit. Furthermore, small decreases in Cdk1 activity during metaphase are sufficient to initiate the reactivation of PP1, which in turn partially deactivates MASTL to release inhibition of PP2A and, hence, create a feedback loop. This feedback loop drives complete deactivation of MASTL, ensuring a strong switch-like activation of phosphatase activity during mitotic exit.
      320Scopus© Citations 40
  • Publication
    Pseudophosphatase STYX modulates cell-fate decisions and cell migration by spatiotemporal regulation of ERK1/2
    (National Academy of Sciences, 2013-07-11) ; ; ;
    Serine/threonine/tyrosine-interacting protein (STYX) is a catalytically inactive member of the dual-specificity phosphatases (DUSPs) family. Whereas the role of DUSPs in cellular signaling is well explored, the function of STYX is still unknown. Here, we identify STYX as a spatial regulator of ERK signaling. We used predictive-model simulation to test several hypotheses for possible modes of STYX action. We show that STYX localizes to the nucleus, competes with nuclear DUSP4 for binding to ERK, and acts as a nuclear anchor that regulates ERK nuclear export. Depletion of STYX increases ERK activity in both cytosol and nucleus. Importantly, depletion of STYX causes an ERK-dependent fragmentation of the Golgi apparatus and inhibits Golgi polarization and directional cell migration. Finally, we show that overexpression of STYX reduces ERK1/2 activation, thereby blocking PC12 cell differentiation. Overall, our results identify STYX as an important regulator of ERK1/2 signaling critical for cell migration and PC12 cell differentiation.
    Scopus© Citations 45  523
  • Publication
    Hippocampal contribution to vector model hypothesis during cue-dependent navigation
    (Cold Spring Harbor Laboratory Press, 2013-06-17) ; ;
    Learning to navigate toward a goal is an essential skill. Place learning is thought to rely on the ability of animals to associate the location of a goal with surrounding environmental cues. Using the Morris water maze, a task popularly used to examine place learning, we demonstrate that distal cues provide animals with distance and directional information. We show how animals use the cues in a visually dependent guidance manner to find the goal. Further, we demonstrate how hippocampal lesions disrupt this learning mechanism. Our results can be explained through the vector model of navigation built on associative learning principles rather than evoking a cognitive map.
    Scopus© Citations 10  340
  • Publication
    Wnt signalling is a bi-directional vulnerability of cancer cells
    Wnt signalling is involved in the formation, metastasis and relapse of a wide array of cancers. However, there is ongoing debate as to whether activation or inhibition of the pathway holds the most promise as a therapeutic treatment for cancer, with conflicting evidence from a variety of tumour types. We show that Wnt/β-catenin signalling is a bi-directional vulnerability of neuroblastoma, malignant melanoma and colorectal cancer, with hyper-activation or repression of the pathway both representing a promising therapeutic strategy, even within the same cancer type. Hyper-activation directs cancer cells to undergo apoptosis, even in cells oncogenically driven by β-catenin. Wnt inhibition blocks proliferation of cancer cells and promotes neuroblastoma differentiation. Wnt and retinoic acid co-treatments synergise, representing a promising combination treatment for MYCN-amplified neuroblastoma. Additionally, we report novel cross-talks between MYCN and β-catenin signalling, which repress normal β-catenin mediated transcriptional regulation. A β-catenin target gene signature could predict patient outcome, as could the expression level of its DNA binding partners, the TCF/LEFs. This β-catenin signature provides a tool to identify neuroblastoma patients likely to benefit from Wnt-directed therapy. Taken together, we show that Wnt/β-catenin signalling is a bi-directional vulnerability of a number of cancer entities, and potentially a more broadly conserved feature of malignant cells.
    Scopus© Citations 27  360
  • Publication
    BAX and SMAC regulate bistable properties of the apoptotic caspase system
    The initiation of apoptosis is a core mechanism in cellular biology by which organisms control the removal of damaged or unnecessary cells. The irreversible activation of caspases is essential for apoptosis, and mathematical models have demonstrated that the process is tightly regulated by positive feedback and a bistable switch. BAX and SMAC are often dysregulated in diseases such as cancer or neurodegeneration and are two key regulators that interact with the caspase system generating the apoptotic switch. Here we present a mathematical model of how BAX and SMAC control the apoptotic switch. Formulated as a system of ordinary differential equations, the model summarises experimental and computational evidence from the literature and incorporates the biochemical mechanisms of how BAX and SMAC interact with the components of the caspase system. Using simulations and bifurcation analysis, we find that both BAX and SMAC regulate the time-delay and activation threshold of the apoptotic switch. Interestingly, the model predicted that BAX (not SMAC) controls the amplitude of the apoptotic switch. Cell culture experiments using siRNA mediated BAX and SMAC knockdowns validated this model prediction. We further validated the model using data of the NCI-60 cell line panel using BAX protein expression as a cell-line specific parameter and show that model simulations correlated with the cellular response to DNA damaging drugs and established a defined threshold for caspase activation that could distinguish between sensitive and resistant melanoma cells. In summary, we present an experimentally validated dynamic model that summarises our current knowledge of how BAX and SMAC regulate the bistable properties of irreversible caspase activation during apoptosis.
      16Scopus© Citations 11
  • Publication
    Integrative omics reveals MYCN as a global suppressor of cellular signalling and enables network-based therapeutic target discovery in neuroblastoma
    Despite intensive study, many mysteries remain about the MYCN oncogene's functions. Here we focus on MYCN's role in neuroblastoma, the most common extracranial childhood cancer. MYCN gene amplification occurs in 20% of cases, but other recurrent somatic mutations are rare. This scarcity of tractable targets has hampered efforts to develop new therapeutic options. We employed a multi-level omics approach to examine MYCN functioning and identify novel therapeutic targets for this largely un-druggable oncogene. We used systems medicine based computational network reconstruction and analysis to integrate a range of omic techniques: sequencing-based transcriptomics, genome-wide chromatin immunoprecipitation, siRNA screening and interaction proteomics, revealing that MYCN controls highly connected networks, with MYCN primarily supressing the activity of network components. MYCN's oncogenic functions are likely independent of its classical heterodimerisation partner, MAX. In particular, MYCN controls its own protein interaction network by transcriptionally regulating its binding partners.Our network-based approach identified vulnerable therapeutically targetable nodes that function as critical regulators or effectors of MYCN in neuroblastoma. These were validated by siRNA knockdown screens, functional studies and patient data. We identified β-estradiol and MAPK/ERK as having functional cross-talk with MYCN and being novel targetable vulnerabilities of MYCN-amplified neuroblastoma. These results reveal surprising differences between the functioning of endogenous, overexpressed and amplified MYCN, and rationalise how different MYCN dosages can orchestrate cell fate decisions and cancerous outcomes. Importantly, this work describes a systems-level approach to systematically uncovering network based vulnerabilities and therapeutic targets for multifactorial diseases by integrating disparate omic data types.
    Scopus© Citations 29  379
  • Publication
    Crosstalk and Signaling Switches in Mitogen-Activated Protein Kinase Cascades
    Mitogen-activated protein kinase (MAPK) cascades control cell fate decisions, such as proliferation, differentiation, and apoptosis by integrating and processing intra- and extracellular cues. However, similar MAPK kinetic profiles can be associated with opposing cellular decisions depending on cell type, signal strength, and dynamics. This implies that signaling by each individual MAPK cascade has to be considered in the context of the entire MAPK network. Here, we develop a dynamic model of feedback and crosstalk for the three major MAPK cascades; extracellular signal-regulated kinase (ERK), p38 mitogen-activated protein kinase (p38), c-Jun N-terminal kinase (JNK), and also include input from protein kinase B (AKT) signaling. Focusing on the bistable activation characteristics of the JNK pathway, this model explains how pathway crosstalk harmonizes different MAPK responses resulting in pivotal cell fate decisions. We show that JNK can switch from a transient to sustained activity due to multiple positive feedback loops. Once activated, positive feedback locks JNK in a highly active state and promotes cell death. The switch is modulated by the ERK, p38, and AKT pathways. ERK activation enhances the dual specificity phosphatase (DUSP) mediated dephosphorylation of JNK and shifts the threshold of the apoptotic switch to higher inputs. Activation of p38 restores the threshold by inhibiting ERK activity via the PP1 or PP2A phosphatases. Finally, AKT activation inhibits the JNK positive feedback, thus abrogating the apoptotic switch and allowing only proliferative signaling. Our model facilitates understanding of how cancerous deregulations disturb MAPK signal processing and provides explanations for certain drug resistances. We highlight a critical role of DUSP1 and DUSP2 expression patterns in facilitating the switching of JNK activity and show how oncogene induced ERK hyperactivity prevents the normal apoptotic switch explaining the failure of certain drugs to induce apoptosis.
    Scopus© Citations 135  388
  • Publication
    Polyubiquitin chain assembly and organization determine the dynamics of protein activation and degradation
    Protein degradation via ubiquitination is a major proteolytic mechanism in cells. Once a protein is destined for degradation, it is tagged by multiple ubiquitin (Ub) molecules. The synthesized polyubiquitin chains can be recognized by the 26S proteosome where proteins are degraded. These chains form through multiple ubiquitination cycles that are similar to multi-site phosphorylation cycles. As kinases and phosphatases, two opposing enzymes (E3 ligases and deubiquitinases DUBs) catalyze (de)ubiquitination cycles. Although multi-ubiquitination cycles are fundamental mechanisms of controlling protein concentrations within a cell, their dynamics have never been explored. Here, we fill this knowledge gap. We show that under permissive physiological conditions, the formation of polyubiquitin chain of length greater than two and subsequent degradation of the ubiquitinated protein, which is balanced by protein synthesis, can display bistable, switch-like responses. Interestingly, the occurrence of bistability becomes pronounced, as the chain grows, giving rise to "all-or-none" regulation at the protein levels. We give predictions of protein distributions under bistable regime awaiting experimental verification. Importantly, we show for the first time that sustained oscillations can robustly arise in the process of formation of ubiquitin chain, largely due to the degradation of the target protein. This new feature is opposite to the properties of multi-site phosphorylation cycles, which are incapable of generating oscillation if the total abundance of interconverted protein forms is conserved. We derive structural and kinetic constraints for the emergence of oscillations, indicating that a competition between different substrate forms and the E3 and DUB is critical for oscillation. Our work provides the first detailed elucidation of the dynamical features brought about by different molecular setups of the polyubiquitin chain assembly process responsible for protein degradation.
      323Scopus© Citations 24
  • Publication
    Signaling pathway models as biomarkers: Patient-specific simulations of JNK activity predict the survival of neuroblastoma patients
    Signaling pathways control cell fate decisions that ultimately determine the behavior of cancer cells. Therefore, the dynamics of pathway activity may contain prognostically relevant information different from that contained in the static nature of other types of biomarkers. To investigate this hypothesis, we characterized the network that regulated stress signaling by the c-Jun N-terminal kinase (JNK) pathway in neuroblastoma cells. We generated an experimentally calibrated and validated computational model of this network and used the model to extract prognostic information from neuroblastoma patient–specific simulations of JNK activation. Switch-like JNK activation mediates cell death by apoptosis. An inability to initiate switch-like JNK activation in the simulations was significantly associated with poor overall survival for patients with neuroblastoma with or without MYCN amplification, indicating that patient-specific simulations of JNK activation could stratify patients. Furthermore, our analysis demonstrated that extracting information about a signaling pathway to develop a prognostically useful model requires understanding of not only components and disease-associated changes in the abundance or activity of the components but also how those changes affect pathway dynamics.
      1349Scopus© Citations 109