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Antisense expression increases gene expression variability and locus interdependency

Genome-wide transcription profiling has revealed extensive expression of non-coding RNAs antisense to genes, yet their functions, if any, remain to be understood. In this study, we perform a systematic analysis of sense-antisense expression in response to genetic and environmental changes in yeast. We find that antisense expression is associated with genes of larger expression variability. This is characterized by more 'switching off' at low levels of expression for genes with antisense compared to genes without, yet similar expression at maximal induction. By disrupting antisense transcription, we demonstrate that antisense expression confers an on-off switch on gene regulation for the SUR7 gene. Consistent with this, genes that must respond in a switch-like manner, such as stress-response and environment-specific genes, are enriched for antisense expression. In addition, our data provide evidence that antisense expression initiated from bidirectional promoters enables the spreading of regulatory signals from one locus to neighbouring genes. These results indicate a general regulatory effect of antisense expression on sense genes and emphasize the importance of antisense-initiating regions downstream of genes in models of gene regulation.

Numerical modelling of microRNA-mediated mRNA decay identifies novel mechanism of microRNA controlled mRNA downregulation

Post-transcriptional control of mRNA by micro-RNAs (miRNAs) represents an important mechanism of gene regulation. miRNAs act by binding to the 3′ untranslated region (3′UTR) of an mRNA, affecting the stability and translation of the target mRNA. Here, we present a numerical model of miRNA-mediated mRNA downregulation and its application to analysis of temporal microarray data of HepG2 cells transfected with miRNA-124a. Using the model our analysis revealed a novel mechanism of mRNA accumulation control by miRNA, predicting that specific mRNAs are controlled in a digital, switch-like manner. Specifically, the contribution of miRNAs to mRNA degradation is switched from maximum to zero in a very short period of time. Such behaviour suggests a model of control in which mRNA is at a certain moment protected from binding of miRNA and further accumulates with a basal rate. Genes associated with this process were identified and parameters of the model for all miRNA-124a affected mRNAs were computed.

Information processing without brains – the power of intercellular regulators in plants

Plants exhibit different developmental strategies than animals; these are characterized by a tight linkage between environmental conditions and development. As plants have neither specialized sensory organs nor a nervous system, intercellular regulators are essential for their development. Recently, major advances have been made in understanding how intercellular regulation is achieved in plants on a molecular level. Plants use a variety of molecules for intercellular regulation: hormones are used as systemic signals that are interpreted at the individual-cell level; receptor peptide-ligand systems regulate local homeostasis; moving transcriptional regulators act in a switch-like manner over small and large distances. Together, these mechanisms coherently coordinate developmental decisions with resource allocation and growth.

Reversible Phosphorylation Subserves Robust Circadian Rhythms by Creating a Switch in Inactivating the Positive Element

Reversible phosphorylation of proteins is ubiquitous in circadian systems, but the role it plays in generating rhythmicity is not completely understood. A common mechanism for most circadian rhythms involves a negative feedback loop between the positive and negative elements. Here, we built a minimal model for the Neurospora crassa circadian clock based on the core negative feedback loop and the protein FREQUENCY (FRQ)-dependent phosphorylation of the White Collar Complex (WCC). The model can reproduce basic features of the clock, such as the period length, phase relationship, and entrainment to light/dark cycles. We found that the activity of WCC can be controlled by FRQ in a switchlike manner owing to zero-order ultrasensitivity. WCC is inactivated when FRQ level crosses a threshold from below. As a result, low cooperativity in transcriptional activation is sufficient for circadian rhythms, and the level of active WCC exhibits spiky oscillations. Such oscillations are robust to molecular noise and may subserve controlling circadian output. Therefore, the core negative feedback together with phosphorylation of the positive element can ensure robust circadian rhythms. Our work provides insights into the critical roles of posttranslational modification in circadian clocks.

Computational Modeling of the Hematopoietic Erythroid-Myeloid Switch Reveals Insights into Cooperativity, Priming, and Irreversibility

Hematopoietic stem cell lineage choices are decided by genetic networks that are turned ON/OFF in a switch-like manner. However, prior to lineage commitment, genes are primed at low expression levels. Understanding the underlying molecular circuitry in terms of how it governs both a primed state and, at the other extreme, a committed state is of relevance not only to hematopoiesis but also to developmental systems in general. We develop a computational model for the hematopoietic erythroid-myeloid lineage decision, which is determined by a genetic switch involving the genes PU.1 and GATA-1. Dynamical models based upon known interactions between these master genes, such as mutual antagonism and autoregulation, fail to make the system bistable, a desired feature for robust lineage determination. We therefore suggest a new mechanism involving a cofactor that is regulated as well as recruited by one of the master genes to bind to the antagonistic partner that is necessary for bistability and hence switch-like behavior. An interesting fallout from this architecture is that suppression of the cofactor through external means can lead to a loss of cooperativity, and hence to a primed state for PU.1 and GATA-1. The PU.1–GATA-1 switch also interacts with another mutually antagonistic pair, –FOG-1. The latter pair inherits the state of its upstream master genes and further reinforces the decision due to several feedback loops, thereby leading to irreversible commitment. The genetic switch, which handles the erythroid-myeloid lineage decision, is an example of a network that implements both a primed and a committed state by regulating cooperativity through recruitment of cofactors. Perturbing the feedback between the master regulators and downstream targets suggests potential reprogramming strategies. The approach points to a framework for lineage commitment studies in general and could aid the search for lineage-determining genes.

Control and signal processing by transcriptional interference

A transcriptional activator can suppress gene expression by interfering with transcription initiated by another activator. Transcriptional interference has been increasingly recognized as a regulatory mechanism of gene expression. The signals received by the two antagonistically acting activators are combined by the polymerase trafficking along the DNA. We have designed a dual-control genetic system in yeast to explore this antagonism systematically. Antagonism by an upstream activator bears the hallmarks of competitive inhibition, whereas a downstream activator inhibits gene expression non-competitively. When gene expression is induced weakly, the antagonistic activator can have a positive effect and can even trigger paradoxical activation. Equilibrium and non-equilibrium models of transcription shed light on the mechanism by which interference converts signals, and reveals that self-antagonism of activators imitates the behavior of feed-forward loops. Indeed, a synthetic circuit generates a bell-shaped response, so that the induction of expression is limited to a narrow range of the input signal. The identification of conserved regulatory principles of interference will help to predict the transcriptional response of genes in their genomic context.

Excitatory and Feed-Forward Inhibitory Hippocampal Synapses Work Synergistically as an Adaptive Filter of Natural Spike Trains

Short-term synaptic plasticity (STP) is an important mechanism for modifying neural circuits during computation. Although STP is much studied, its role in the processing of complex natural spike patterns is unknown. Here we analyze the responses of excitatory and inhibitory hippocampal synapses to natural spike trains at near-physiological temperatures. Our results show that excitatory and inhibitory synapses express complementary sets of STP components that selectively change synaptic strength during epochs of high-frequency discharge associated with hippocampal place fields. In both types of synapses, synaptic strength rapidly alternates between a near-constant level during low activity and another near-constant, but elevated (for excitatory synapses) or reduced (for inhibitory synapses) level during high-frequency epochs. These history-dependent changes in synaptic strength are largely independent of the particular temporal pattern within the discharges, and occur concomitantly in the two types of synapses. When excitatory and feed-forward inhibitory synapses are co-activated within the hippocampal feed-forward circuit unit, the net effect of their complementary STP is an additional increase in the gain of excitatory synapses during high-frequency discharges via selective disinhibition. Thus, excitatory and feed-forward inhibitory hippocampal synapses in vitro act synergistically as an adaptive filter that operates in a switch-like manner and is selective for high-frequency epochs.

A Polybasic Motif Allows N-WASP to Act as a Sensor of PIP 2 Density

Phosphatidylinositol 4,5-bisphosphate (PIP 2) activates the actin regulatory protein N-WASP by binding to a short polybasic region involved in N-WASP autoinhibition. Here, we show that unlike canonical lipid binding modules, such as PH domains, this polybasic motif binds PIP 2 in a multivalent, cooperative manner. As a result, PIP 2 activation of N-WASP-mediated actin polymerization in vitro and in extracts is ultrasensitive: above a certain threshold, N-WASP responds in a switch-like manner to a small increase in the density of PIP 2 (Hill coefficient n H = ∼20). We show that the sharpness of the PIP 2 activation threshold can be tuned by varying the length of the polybasic motif. This sharp activation threshold may help suppress N-WASP activation by quiescent PIP 2 levels yet leave it poised for activation upon subtle, signaling-induced perturbations in PIP 2 distribution.

Simulating genetic networks made easy: network construction with simple building blocks.

We present SIM-plex, a genetic network simulator with a very intuitive interface in which a user can easily specify interactions as simple 'if-then' statements. The simulator is based on the mathematical model of Piecewise Linear Differential Equations (PLDEs). With PLDEs, genetic interactions are approximated as acting in a switch-like manner. The Java program, examples and a tutorial are available at http://www.psb.ugent.be/cbd/ {stcru,makui}@psb.ugent.be

Activation of single heteromeric GABAA receptor ion channels by full and partial agonists

The linkage between agonist binding and the activation of a GABA(A) receptor ion channel is yet to be resolved. This aspect was examined on human recombinant alpha1beta2gamma2S GABA(A) receptors expressed in human embryonic kidney cells using the following series of receptor agonists: GABA, isoguvacine, 4,5,6,7-tetrahydroisoxazolo[5,4-c]pyridin-3-ol (THIP), isonipecotic acid, piperidine-4-sulphonic acid (P4S), imidazole-4-acetic acid (IAA), 5-(4-piperidyl)-3-isothiazolol (thio-4-PIOL) and 5-(4-piperidyl)-3-isoxazolol (4-PIOL). Whole-cell concentration-response curves enabled the agonists to be categorized into four classes based upon their maximum responses. Single channel analyses revealed that the channel conductance of 25-27 pS was unaffected by the agonists. However, two open states were resolved from the open period distributions with mean open times reduced 5-fold by the weakest partial agonists. Using saturating agonist concentrations, estimates of the channel shutting rate, alpha, ranged from 200 to 600 s(-1). The shut period distributions were described by three or four components and for the weakest partial agonists, the interburst shut periods increased whilst the mean burst durations and longest burst lengths were reduced relative to the full agonists. From the burst analyses, the opening rates for channel activation, beta, and the total dissociation rates, k(-1), for the agonists leaving the receptor were estimated. The agonist efficacies were larger for the full agonists (E approximately 7-9) compared to the weak partial agonists ( approximately 0.4-0.6). Overall, changes in agonist efficacy largely determined the different agonist profiles with contributions from the agonist affinities and the degree of receptor desensitization. From this we conclude that GABA(A) receptor activation does not occur in a switch-like manner since the agonist recognition sites are flexible, accommodating diverse agonist structures which differentially influence the opening and shutting rates of the ion channel.

Targeted reduction of complex models with time scale hierarchy – a case study

With the increasing flow of biological data there is a growing demand for mathematical tools whereby essential aspects of complex causal dynamic models can be captured and detected by simpler mathematical models without sacrificing too much of the realism provided by the original ones. Given the presence of a time scale hierarchy, singular perturbation techniques represent an elegant method for making such minimised mathematical representations. Any reduction of a complex model by singular perturbation methods is a targeted reduction by the fact that one has to pick certain mechanisms, processes or aspects thought to be essential in a given explanatory context. Here we illustrate how such a targeted reduction of a complex model of melanogenesis in mammals recently developed by the authors provides a way to improve the understanding of how the melanogenic system may behave in a switch-like manner between production of the two major types of melanins. The reduced model is shown by numerical means to be in good quantitative agreement with the original model. Furthermore, it is shown how the reduced model discloses hidden robustness features of the full model, and how the making of a reduced model represents an efficient analytical sensitivity analysis. In addition to yielding new insights concerning the melanogenic system, the paper provides an illustration of a protocol that could be followed to make validated simplifications of complex biological models possessing time scale hierarchies.

Hot Feet: Erythromelalgia and Related Disorders

Erythromelalgia is an extraordinary pain syndrome first described by S. Weir Mitchell in 1878. Episodes of severe burning pain in the distal limbs, accompanied by striking redness and warmth of the skin, are precipitated by heat or activity and can be terminated only by cooling the affected part. Primary erythromelalgia is a sporadic or autosomal-dominant hereditary disorder whose symptoms begin in childhood. Secondary erythromelalgia occurs in association with thrombocythemia, collagen-vascular diseases, diabetes mellitus, peripheral neuropathy, and use of certain drugs. Aspirin is effective for patients with thrombocythemia, but most other cases are very resistant to treatment. The pathogenesis of erythromelalgia has remained puzzling, especially the peculiar switch-like manner in which symptoms are turned on by heat and turned off by cold. Following Ochoa's description of the ABC (angry backfiring C nociceptors) syndrome, it seems plausible to regard erythromelalgia as a problem of sensitized skin polymodal C-fiber receptors. C-fiber threshold to activation by heat would be lowered to 32 degrees C to 36 degrees C; activated C fibers would cause vasodilation via axon reflexes with redness, heat, and swelling. Cooling would bring the nociceptors below threshold. Secondary erythromelalgia may result from humoral factors released from platelets or ischemic tissues or from C-fiber injury in some cases of neuropathy, whereas primary erythromelalgia could be due to a mutation of the capsaicin receptor.

Emergence of donor control in patchy predator-prey systems

We study a general predator-prey system in a spatially heterogeneous environment. The predation process, which occurs on a behavioural time-scale, is much faster than the other processes (reproduction, natural mortality and migrations) occurring on the population dynamics time-scale. We show that, taking account of this difference in time-scales, and assuming that the prey have a refuge, the dynamics of the system on a slow time-scale become donor-controlled. Even though predators may control the prey density locally and on a behavioural fast time-scale, nevertheless, both globally and on a slow time-scale, the prey dynamics are independent of predator density: the presence of predators generates a constant prey mortality. In other words, in heterogeneous environments, the prey population dynamics depend in a switch-like manner on the presence or absence of predators, not on their actual density.