Evolutionary modelling of feed forward loops in gene regulatory networks

Feed forward loops (FFLs) are gene regulatory network motifs. They exist in different types, defined by the signs of the effects of genes in the motif on one another. We examine 36 feed forward loops in Escherichia coli, using evolutionary simulations to predict the forms of FFL expected to evolve to generate the pattern of expression of the output gene. These predictions are tested using likelihood ratios, comparing likelihoods of the observed FFL structures with their likelihoods under null models. The very high likelihood ratios generated, of over 10(11), suggest that evolutionary simulation is a valuable component in the explanation of FFL structure.     

Structural discrimination of robustness in transcriptional feedforward loops for pattern formation

Signaling pathways are interconnected to regulatory circuits for sensing the environment and expressing the appropriate genetic profile. In particular, gradients of diffusing molecules (morphogens) determine cell fate at a given position, dictating development and spatial organization. The feedforward loop (FFL) circuit is among the simplest genetic architectures able to generate one-stripe patterns by operating as an amplitude detection device, where high output levels are achieved at intermediate input ones. Here, using a heuristic optimization-based approach, we dissected the design space containing all possible topologies and parameter values of the FFL circuits. We explored the ability of being sensitive or adaptive to variations in the critical morphogen level where cell fate is switched. We found four different solutions for precision, corresponding to the four incoherent architectures, but remarkably only one mode for adaptiveness, the incoherent type 4 (I4-FFL). We further carried out a theoretical study to unveil the design principle for such structural discrimination, finding that the synergistic action and cooperative binding on the downstream promoter are instrumental to achieve absolute adaptive responses. Subsequently, we analyzed the robustness of these optimal circuits against perturbations in the kinetic parameters and molecular noise, which has allowed us to depict a scenario where adaptiveness, parameter sensitivity and noise tolerance are different, correlated facets of the robustness of the I4-FFL circuit. Strikingly, we showed a strong correlation between the input (environment-related) and the intrinsic (mutation-related) susceptibilities. Finally, we discussed the evolution of incoherent regulations in terms of multifunctionality and robustness.     

Design and implementation of three incoherent feed-forward motif based biological concentration sensors

Synthetic biology is a useful tool to investigate the dynamics of small biological networks and to assess our capacity to predict their behavior from computational models. In this work we report the construction of three different synthetic networks in Escherichia coli based upon the incoherent feed-forward loop architecture. The steady state behavior of the networks was investigated experimentally and computationally under different mutational regimes in a population based assay. Our data shows that the three incoherent feed-forward networks, using three different macromolecular inhibitory elements, reproduce the behavior predicted from our computational model. We also demonstrate that specific biological motifs can be designed to generate similar behavior using different components. In addition we show how it is possible to tune the behavior of the networks in a predicable manner by applying suitable mutations to the inhibitory elements. Brian Aufderheide provided material support.     

A DPP-mediated feed-forward loop canalizes morphogenesis during Drosophila dorsal closure

Development is robust because nature has selected various mechanisms to buffer the deleterious effects of environmental and genetic variations to deliver phenotypic stability. Robustness relies on smart network motifs such as feed-forward loops (FFLs) that ensure the reliable interpretation of developmental signals. In this paper, we show that Decapentaplegic (DPP) and JNK form a coherent FFL that controls the specification and differentiation of leading edge cells during Drosophila melanogaster dorsal closure (DC). We provide molecular evidence that through repression by Brinker (Brk), the DPP branch of the FFL filters unwanted JNK activity. High-throughput live imaging revealed that this DPP/Brk branch is dispensable for DC under normal conditions but is required when embryos are subjected to thermal stress. Our results indicate that the wiring of DPP signaling buffers against environmental challenges and canalizes cell identity. We propose that the main function of DPP pathway during Drosophila DC is to ensure robust morphogenesis, a distinct function from its well-established ability to spread spatial information.     

The incoherent feed‐forward loop can generate non‐monotonic input functions for genes

Gene regulation networks contain recurring circuit patterns called network motifs. One of the most common network motif is the incoherent type 1 feed‐forward loop (I1‐FFL), in which an activator controls both gene and repressor of that gene. This motif was shown to act as a pulse generator and response accelerator of gene expression. Here we consider an additional function of this motif: the I1‐FFL can generate a non‐monotonic dependence of gene expression on the input signal. Here, we study this experimentally in the galactose system of Escherichia coli, which is regulated by an I1‐FFL. The promoter activity of two of the gal operons, galETK and galP, peaks at intermediate levels of the signal cAMP. We find that mutants in which the I1‐FFL is disrupted lose this non‐monotonic behavior, and instead display monotonic input functions. Theoretical analysis suggests that non‐monotonic input functions can be achieved for a wide range of parameters by the I1‐FFL. The models also suggest regimes where a monotonic input‐function can occur, as observed in the mglBAC operon regulated by the same I1‐FFL. The present study thus experimentally demonstrates how upstream circuitry can affect gene input functions and how an I1‐FFL functions within its natural context in the cell.