@inproceedings{7147, abstract = {The expression of a gene is characterised by its transcription factors and the function processing them. If the transcription factors are not affected by gene products, the regulating function is often represented as a combinational logic circuit, where the outputs (product) are determined by current input values (transcription factors) only, and are hence independent on their relative arrival times. However, the simultaneous arrival of transcription factors (TFs) in genetic circuits is a strong assumption, given that the processes of transcription and translation of a gene into a protein introduce intrinsic time delays and that there is no global synchronisation among the arrival times of different molecular species at molecular targets. In this paper, we construct an experimentally implementable genetic circuit with two inputs and a single output, such that, in presence of small delays in input arrival, the circuit exhibits qualitatively distinct observable phenotypes. In particular, these phenotypes are long lived transients: they all converge to a single value, but so slowly, that they seem stable for an extended time period, longer than typical experiment duration. We used rule-based language to prototype our circuit, and we implemented a search for finding the parameter combinations raising the phenotypes of interest. The behaviour of our prototype circuit has wide implications. First, it suggests that GRNs can exploit event timing to create phenotypes. Second, it opens the possibility that GRNs are using event timing to react to stimuli and memorise events, without explicit feedback in regulation. From the modelling perspective, our prototype circuit demonstrates the critical importance of analysing the transient dynamics at the promoter binding sites of the DNA, before applying rapid equilibrium assumptions.}, author = {Guet, Calin C and Henzinger, Thomas A and Igler, Claudia and Petrov, Tatjana and Sezgin, Ali}, booktitle = {17th International Conference on Computational Methods in Systems Biology}, isbn = {9783030313036}, issn = {1611-3349}, location = {Trieste, Italy}, pages = {155--187}, publisher = {Springer Nature}, title = {{Transient memory in gene regulation}}, doi = {10.1007/978-3-030-31304-3_9}, volume = {11773}, year = {2019}, } @article{138, abstract = {Autoregulation is the direct modulation of gene expression by the product of the corresponding gene. Autoregulation of bacterial gene expression has been mostly studied at the transcriptional level, when a protein acts as the cognate transcriptional repressor. A recent study investigating dynamics of the bacterial toxin–antitoxin MazEF system has shown how autoregulation at both the transcriptional and post-transcriptional levels affects the heterogeneity of Escherichia coli populations. Toxin–antitoxin systems hold a crucial but still elusive part in bacterial response to stress. This perspective highlights how these modules can also serve as a great model system for investigating basic concepts in gene regulation. However, as the genomic background and environmental conditions substantially influence toxin activation, it is important to study (auto)regulation of toxin–antitoxin systems in well-defined setups as well as in conditions that resemble the environmental niche.}, author = {Nikolic, Nela}, journal = {Current Genetics}, number = {1}, pages = {133--138}, publisher = {Springer}, title = {{Autoregulation of bacterial gene expression: lessons from the MazEF toxin–antitoxin system}}, doi = {10.1007/s00294-018-0879-8}, volume = {65}, year = {2019}, } @article{196, abstract = {The abelian sandpile serves as a model to study self-organized criticality, a phenomenon occurring in biological, physical and social processes. The identity of the abelian group is a fractal composed of self-similar patches, and its limit is subject of extensive collaborative research. Here, we analyze the evolution of the sandpile identity under harmonic fields of different orders. We show that this evolution corresponds to periodic cycles through the abelian group characterized by the smooth transformation and apparent conservation of the patches constituting the identity. The dynamics induced by second and third order harmonics resemble smooth stretchings, respectively translations, of the identity, while the ones induced by fourth order harmonics resemble magnifications and rotations. Starting with order three, the dynamics pass through extended regions of seemingly random configurations which spontaneously reassemble into accentuated patterns. We show that the space of harmonic functions projects to the extended analogue of the sandpile group, thus providing a set of universal coordinates identifying configurations between different domains. Since the original sandpile group is a subgroup of the extended one, this directly implies that it admits a natural renormalization. Furthermore, we show that the harmonic fields can be induced by simple Markov processes, and that the corresponding stochastic dynamics show remarkable robustness over hundreds of periods. Finally, we encode information into seemingly random configurations, and decode this information with an algorithm requiring minimal prior knowledge. Our results suggest that harmonic fields might split the sandpile group into sub-sets showing different critical coefficients, and that it might be possible to extend the fractal structure of the identity beyond the boundaries of its domain. }, author = {Lang, Moritz and Shkolnikov, Mikhail}, issn = {1091-6490}, journal = {Proceedings of the National Academy of Sciences}, number = {8}, pages = {2821--2830}, publisher = {National Academy of Sciences}, title = {{Harmonic dynamics of the Abelian sandpile}}, doi = {10.1073/pnas.1812015116}, volume = {116}, year = {2019}, } @misc{7016, abstract = {Organisms cope with change by employing transcriptional regulators. However, when faced with rare environments, the evolution of transcriptional regulators and their promoters may be too slow. We ask whether the intrinsic instability of gene duplication and amplification provides a generic alternative to canonical gene regulation. By real-time monitoring of gene copy number mutations in E. coli, we show that gene duplications and amplifications enable adaptation to fluctuating environments by rapidly generating copy number, and hence expression level, polymorphism. This ‘amplification-mediated gene expression tuning’ occurs on timescales similar to canonical gene regulation and can deal with rapid environmental changes. Mathematical modeling shows that amplifications also tune gene expression in stochastic environments where transcription factor-based schemes are hard to evolve or maintain. The fleeting nature of gene amplifications gives rise to a generic population-level mechanism that relies on genetic heterogeneity to rapidly tune expression of any gene, without leaving any genomic signature.}, author = {Tomanek, Isabella}, keywords = {Escherichia coli, gene amplification, galactose, DOG, experimental evolution, Illumina sequence data, FACS data, microfluidics data}, publisher = {Institute of Science and Technology Austria}, title = {{Data for the paper "Gene amplification as a form of population-level gene expression regulation"}}, doi = {10.15479/AT:ISTA:7016}, year = {2019}, } @phdthesis{6371, abstract = {Decades of studies have revealed the mechanisms of gene regulation in molecular detail. We make use of such well-described regulatory systems to explore how the molecular mechanisms of protein-protein and protein-DNA interactions shape the dynamics and evolution of gene regulation. i) We uncover how the biophysics of protein-DNA binding determines the potential of regulatory networks to evolve and adapt, which can be captured using a simple mathematical model. ii) The evolution of regulatory connections can lead to a significant amount of crosstalk between binding proteins. We explore the effect of crosstalk on gene expression from a target promoter, which seems to be modulated through binding competition at non-specific DNA sites. iii) We investigate how the very same biophysical characteristics as in i) can generate significant fitness costs for cells through global crosstalk, meaning non-specific DNA binding across the genomic background. iv) Binding competition between proteins at a target promoter is a prevailing regulatory feature due to the prevalence of co-regulation at bacterial promoters. However, the dynamics of these systems are not always straightforward to determine even if the molecular mechanisms of regulation are known. A detailed model of the biophysical interactions reveals that interference between the regulatory proteins can constitute a new, generic form of system memory that records the history of the input signals at the promoter. We demonstrate how the biophysics of protein-DNA binding can be harnessed to investigate the principles that shape and ultimately limit cellular gene regulation. These results provide a basis for studies of higher-level functionality, which arises from the underlying regulation. }, author = {Igler, Claudia}, issn = {2663-337X}, keywords = {gene regulation, biophysics, transcription factor binding, bacteria}, pages = {152}, publisher = {Institute of Science and Technology Austria}, title = {{On the nature of gene regulatory design - The biophysics of transcription factor binding shapes gene regulation}}, doi = {10.15479/AT:ISTA:6371}, year = {2019}, } @article{305, abstract = {The hanging-drop network (HDN) is a technology platform based on a completely open microfluidic network at the bottom of an inverted, surface-patterned substrate. The platform is predominantly used for the formation, culturing, and interaction of self-assembled spherical microtissues (spheroids) under precisely controlled flow conditions. Here, we describe design, fabrication, and operation of microfluidic hanging-drop networks.}, author = {Misun, Patrick and Birchler, Axel and Lang, Moritz and Hierlemann, Andreas and Frey, Olivier}, journal = {Methods in Molecular Biology}, pages = {183 -- 202}, publisher = {Springer}, title = {{Fabrication and operation of microfluidic hanging drop networks}}, doi = {10.1007/978-1-4939-7792-5_15}, volume = {1771}, year = {2018}, } @article{723, abstract = {Escaping local optima is one of the major obstacles to function optimisation. Using the metaphor of a fitness landscape, local optima correspond to hills separated by fitness valleys that have to be overcome. We define a class of fitness valleys of tunable difficulty by considering their length, representing the Hamming path between the two optima and their depth, the drop in fitness. For this function class we present a runtime comparison between stochastic search algorithms using different search strategies. The (1+1) EA is a simple and well-studied evolutionary algorithm that has to jump across the valley to a point of higher fitness because it does not accept worsening moves (elitism). In contrast, the Metropolis algorithm and the Strong Selection Weak Mutation (SSWM) algorithm, a famous process in population genetics, are both able to cross the fitness valley by accepting worsening moves. We show that the runtime of the (1+1) EA depends critically on the length of the valley while the runtimes of the non-elitist algorithms depend crucially on the depth of the valley. Moreover, we show that both SSWM and Metropolis can also efficiently optimise a rugged function consisting of consecutive valleys.}, author = {Oliveto, Pietro and Paixao, Tiago and Pérez Heredia, Jorge and Sudholt, Dirk and Trubenova, Barbora}, journal = {Algorithmica}, number = {5}, pages = {1604 -- 1633}, publisher = {Springer}, title = {{How to escape local optima in black box optimisation when non elitism outperforms elitism}}, doi = {10.1007/s00453-017-0369-2}, volume = {80}, year = {2018}, } @article{503, abstract = {Buffers are essential for diluting bacterial cultures for flow cytometry analysis in order to study bacterial physiology and gene expression parameters based on fluorescence signals. Using a variety of constitutively expressed fluorescent proteins in Escherichia coli K-12 strain MG1655, we found strong artifactual changes in fluorescence levels after dilution into the commonly used flow cytometry buffer phosphate-buffered saline (PBS) and two other buffer solutions, Tris-HCl and M9 salts. These changes appeared very rapidly after dilution, and were linked to increased membrane permeability and loss in cell viability. We observed buffer-related effects in several different E. coli strains, K-12, C and W, but not E. coli B, which can be partially explained by differences in lipopolysaccharide (LPS) and outer membrane composition. Supplementing the buffers with divalent cations responsible for outer membrane stability, Mg2+ and Ca2+, preserved fluorescence signals, membrane integrity and viability of E. coli. Thus, stabilizing the bacterial outer membrane is essential for precise and unbiased measurements of fluorescence parameters using flow cytometry.}, author = {Tomasek, Kathrin and Bergmiller, Tobias and Guet, Calin C}, journal = {Journal of Biotechnology}, pages = {40 -- 52}, publisher = {Elsevier}, title = {{Lack of cations in flow cytometry buffers affect fluorescence signals by reducing membrane stability and viability of Escherichia coli strains}}, doi = {10.1016/j.jbiotec.2018.01.008}, volume = {268}, year = {2018}, } @article{82, abstract = {In experimental cultures, when bacteria are mixed with lytic (virulent) bacteriophage, bacterial cells resistant to the phage commonly emerge and become the dominant population of bacteria. Following the ascent of resistant mutants, the densities of bacteria in these simple communities become limited by resources rather than the phage. Despite the evolution of resistant hosts, upon which the phage cannot replicate, the lytic phage population is most commonly maintained in an apparently stable state with the resistant bacteria. Several mechanisms have been put forward to account for this result. Here we report the results of population dynamic/evolution experiments with a virulent mutant of phage Lambda, λVIR, and Escherichia coli in serial transfer cultures. We show that, following the ascent of λVIR-resistant bacteria, λVIRis maintained in the majority of cases in maltose-limited minimal media and in all cases in nutrient-rich broth. Using mathematical models and experiments, we show that the dominant mechanism responsible for maintenance of λVIRin these resource-limited populations dominated by resistant E. coli is a high rate of either phenotypic or genetic transition from resistance to susceptibility—a hitherto undemonstrated mechanism we term "leaky resistance." We discuss the implications of leaky resistance to our understanding of the conditions for the maintenance of phage in populations of bacteria—their “existence conditions.”.}, author = {Chaudhry, Waqas and Pleska, Maros and Shah, Nilang and Weiss, Howard and Mccall, Ingrid and Meyer, Justin and Gupta, Animesh and Guet, Calin C and Levin, Bruce}, journal = {PLoS Biology}, number = {8}, publisher = {Public Library of Science}, title = {{Leaky resistance and the conditions for the existence of lytic bacteriophage}}, doi = {10.1371/journal.pbio.2005971}, volume = {16}, year = {2018}, } @misc{9810, author = {Chaudhry, Waqas and Pleska, Maros and Shah, Nilang and Weiss, Howard and Mccall, Ingrid and Meyer, Justin and Gupta, Animesh and Guet, Calin C and Levin, Bruce}, publisher = {Public Library of Science}, title = {{Numerical data used in figures}}, doi = {10.1371/journal.pbio.2005971.s008}, year = {2018}, } @article{457, abstract = {Temperate bacteriophages integrate in bacterial genomes as prophages and represent an important source of genetic variation for bacterial evolution, frequently transmitting fitness-augmenting genes such as toxins responsible for virulence of major pathogens. However, only a fraction of bacteriophage infections are lysogenic and lead to prophage acquisition, whereas the majority are lytic and kill the infected bacteria. Unless able to discriminate lytic from lysogenic infections, mechanisms of immunity to bacteriophages are expected to act as a double-edged sword and increase the odds of survival at the cost of depriving bacteria of potentially beneficial prophages. We show that although restriction-modification systems as mechanisms of innate immunity prevent both lytic and lysogenic infections indiscriminately in individual bacteria, they increase the number of prophage-acquiring individuals at the population level. We find that this counterintuitive result is a consequence of phage-host population dynamics, in which restriction-modification systems delay infection onset until bacteria reach densities at which the probability of lysogeny increases. These results underscore the importance of population-level dynamics as a key factor modulating costs and benefits of immunity to temperate bacteriophages}, author = {Pleska, Maros and Lang, Moritz and Refardt, Dominik and Levin, Bruce and Guet, Calin C}, journal = {Nature Ecology and Evolution}, number = {2}, pages = {359 -- 366}, publisher = {Springer Nature}, title = {{Phage-host population dynamics promotes prophage acquisition in bacteria with innate immunity}}, doi = {10.1038/s41559-017-0424-z}, volume = {2}, year = {2018}, } @article{5984, abstract = {G-protein-coupled receptors (GPCRs) form the largest receptor family, relay environmental stimuli to changes in cell behavior and represent prime drug targets. Many GPCRs are classified as orphan receptors because of the limited knowledge on their ligands and coupling to cellular signaling machineries. Here, we engineer a library of 63 chimeric receptors that contain the signaling domains of human orphan and understudied GPCRs functionally linked to the light-sensing domain of rhodopsin. Upon stimulation with visible light, we identify activation of canonical cell signaling pathways, including cAMP-, Ca2+-, MAPK/ERK-, and Rho-dependent pathways, downstream of the engineered receptors. For the human pseudogene GPR33, we resurrect a signaling function that supports its hypothesized role as a pathogen entry site. These results demonstrate that substituting unknown chemical activators with a light switch can reveal information about protein function and provide an optically controlled protein library for exploring the physiology and therapeutic potential of understudied GPCRs.}, author = {Morri, Maurizio and Sanchez-Romero, Inmaculada and Tichy, Alexandra-Madelaine and Kainrath, Stephanie and Gerrard, Elliot J. and Hirschfeld, Priscila and Schwarz, Jan and Janovjak, Harald L}, issn = {2041-1723}, journal = {Nature Communications}, number = {1}, publisher = {Springer Nature}, title = {{Optical functionalization of human class A orphan G-protein-coupled receptors}}, doi = {10.1038/s41467-018-04342-1}, volume = {9}, year = {2018}, } @article{19, abstract = {Bacteria regulate genes to survive antibiotic stress, but regulation can be far from perfect. When regulation is not optimal, mutations that change gene expression can contribute to antibiotic resistance. It is not systematically understood to what extent natural gene regulation is or is not optimal for distinct antibiotics, and how changes in expression of specific genes quantitatively affect antibiotic resistance. Here we discover a simple quantitative relation between fitness, gene expression, and antibiotic potency, which rationalizes our observation that a multitude of genes and even innate antibiotic defense mechanisms have expression that is critically nonoptimal under antibiotic treatment. First, we developed a pooled-strain drug-diffusion assay and screened Escherichia coli overexpression and knockout libraries, finding that resistance to a range of 31 antibiotics could result from changing expression of a large and functionally diverse set of genes, in a primarily but not exclusively drug-specific manner. Second, by synthetically controlling the expression of single-drug and multidrug resistance genes, we observed that their fitness-expression functions changed dramatically under antibiotic treatment in accordance with a log-sensitivity relation. Thus, because many genes are nonoptimally expressed under antibiotic treatment, many regulatory mutations can contribute to resistance by altering expression and by activating latent defenses.}, author = {Palmer, Adam and Chait, Remy P and Kishony, Roy}, issn = {0737-4038}, journal = {Molecular Biology and Evolution}, number = {11}, pages = {2669 -- 2684}, publisher = {Oxford University Press}, title = {{Nonoptimal gene expression creates latent potential for antibiotic resistance}}, doi = {10.1093/molbev/msy163}, volume = {35}, year = {2018}, } @article{438, abstract = {The MazF toxin sequence-specifically cleaves single-stranded RNA upon various stressful conditions, and it is activated as a part of the mazEF toxin–antitoxin module in Escherichia coli. Although autoregulation of mazEF expression through the MazE antitoxin-dependent transcriptional repression has been biochemically characterized, less is known about post-transcriptional autoregulation, as well as how both of these autoregulatory features affect growth of single cells during conditions that promote MazF production. Here, we demonstrate post-transcriptional autoregulation of mazF expression dynamics by MazF cleaving its own transcript. Single-cell analyses of bacterial populations during ectopic MazF production indicated that two-level autoregulation of mazEF expression influences cell-to-cell growth rate heterogeneity. The increase in growth rate heterogeneity is governed by the MazE antitoxin, and tuned by the MazF-dependent mazF mRNA cleavage. Also, both autoregulatory features grant rapid exit from the stress caused by mazF overexpression. Time-lapse microscopy revealed that MazF-mediated cleavage of mazF mRNA leads to increased temporal variability in length of individual cells during ectopic mazF overexpression, as explained by a stochastic model indicating that mazEF mRNA cleavage underlies temporal fluctuations in MazF levels during stress.}, author = {Nikolic, Nela and Bergmiller, Tobias and Vandervelde, Alexandra and Albanese, Tanino and Gelens, Lendert and Moll, Isabella}, journal = {Nucleic Acids Research}, number = {6}, pages = {2918--2931}, publisher = {Oxford University Press}, title = {{Autoregulation of mazEF expression underlies growth heterogeneity in bacterial populations}}, doi = {10.1093/nar/gky079}, volume = {46}, year = {2018}, } @misc{5569, abstract = {Nela Nikolic, Tobias Bergmiller, Alexandra Vandervelde, Tanino G. Albanese, Lendert Gelens, and Isabella Moll (2018) “Autoregulation of mazEF expression underlies growth heterogeneity in bacterial populations” Nucleic Acids Research, doi: 10.15479/AT:ISTA:74; microscopy experiments by Tobias Bergmiller; image and data analysis by Nela Nikolic.}, author = {Bergmiller, Tobias and Nikolic, Nela}, keywords = {microscopy, microfluidics}, publisher = {Institute of Science and Technology Austria}, title = {{Time-lapse microscopy data}}, doi = {10.15479/AT:ISTA:74}, year = {2018}, } @article{161, abstract = {Which properties of metabolic networks can be derived solely from stoichiometry? Predictive results have been obtained by flux balance analysis (FBA), by postulating that cells set metabolic fluxes to maximize growth rate. Here we consider a generalization of FBA to single-cell level using maximum entropy modeling, which we extend and test experimentally. Specifically, we define for Escherichia coli metabolism a flux distribution that yields the experimental growth rate: the model, containing FBA as a limit, provides a better match to measured fluxes and it makes a wide range of predictions: on flux variability, regulation, and correlations; on the relative importance of stoichiometry vs. optimization; on scaling relations for growth rate distributions. We validate the latter here with single-cell data at different sub-inhibitory antibiotic concentrations. The model quantifies growth optimization as emerging from the interplay of competitive dynamics in the population and regulation of metabolism at the level of single cells.}, author = {De Martino, Daniele and Mc, Andersson Anna and Bergmiller, Tobias and Guet, Calin C and Tkacik, Gasper}, journal = {Nature Communications}, number = {1}, publisher = {Springer Nature}, title = {{Statistical mechanics for metabolic networks during steady state growth}}, doi = {10.1038/s41467-018-05417-9}, volume = {9}, year = {2018}, } @phdthesis{26, abstract = {Expression of genes is a fundamental molecular phenotype that is subject to evolution by different types of mutations. Both the rate and the effect of mutations may depend on the DNA sequence context of a particular gene or a particular promoter sequence. In this thesis I investigate the nature of this dependence using simple genetic systems in Escherichia coli. With these systems I explore the evolution of constitutive gene expression from random starting sequences at different loci on the chromosome and at different locations in sequence space. First, I dissect chromosomal neighborhood effects that underlie locus-dependent differences in the potential of a gene under selection to become more highly expressed. Next, I find that the effects of point mutations in promoter sequences are dependent on sequence context, and that an existing energy matrix model performs poorly in predicting relative expression of unrelated sequences. Finally, I show that a substantial fraction of random sequences contain functional promoters and I present an extended thermodynamic model that predicts promoter strength in full sequence space. Taken together, these results provide new insights and guides on how to integrate information on sequence context to improve our qualitative and quantitative understanding of bacterial gene expression, with implications for rapid evolution of drug resistance, de novo evolution of genes, and horizontal gene transfer.}, author = {Steinrück, Magdalena}, issn = {2663-337X}, pages = {109}, publisher = {Institute of Science and Technology Austria}, title = {{The influence of sequence context on the evolution of bacterial gene expression}}, doi = {10.15479/AT:ISTA:th1059}, year = {2018}, } @article{67, abstract = {Gene regulatory networks evolve through rewiring of individual components—that is, through changes in regulatory connections. However, the mechanistic basis of regulatory rewiring is poorly understood. Using a canonical gene regulatory system, we quantify the properties of transcription factors that determine the evolutionary potential for rewiring of regulatory connections: robustness, tunability and evolvability. In vivo repression measurements of two repressors at mutated operator sites reveal their contrasting evolutionary potential: while robustness and evolvability were positively correlated, both were in trade-off with tunability. Epistatic interactions between adjacent operators alleviated this trade-off. A thermodynamic model explains how the differences in robustness, tunability and evolvability arise from biophysical characteristics of repressor–DNA binding. The model also uncovers that the energy matrix, which describes how mutations affect repressor–DNA binding, encodes crucial information about the evolutionary potential of a repressor. The biophysical determinants of evolutionary potential for regulatory rewiring constitute a mechanistic framework for understanding network evolution.}, author = {Igler, Claudia and Lagator, Mato and Tkacik, Gasper and Bollback, Jonathan P and Guet, Calin C}, journal = {Nature Ecology and Evolution}, number = {10}, pages = {1633 -- 1643}, publisher = {Nature Publishing Group}, title = {{Evolutionary potential of transcription factors for gene regulatory rewiring}}, doi = {10.1038/s41559-018-0651-y}, volume = {2}, year = {2018}, } @misc{5585, abstract = {Mean repression values and standard error of the mean are given for all operator mutant libraries.}, author = {Igler, Claudia and Lagator, Mato and Tkacik, Gasper and Bollback, Jonathan P and Guet, Calin C}, publisher = {Institute of Science and Technology Austria}, title = {{Data for the paper Evolutionary potential of transcription factors for gene regulatory rewiring}}, doi = {10.15479/AT:ISTA:108}, year = {2018}, } @article{538, abstract = {Optogenetik und Photopharmakologie ermöglichen präzise räumliche und zeitliche Kontrolle von Proteinwechselwirkung und -funktion in Zellen und Tieren. Optogenetische Methoden, die auf grünes Licht ansprechen und zum Trennen von Proteinkomplexen geeignet sind, sind nichtweitläufig verfügbar, würden jedoch mehrfarbige Experimente zur Beantwortung von biologischen Fragestellungen ermöglichen. Hier demonstrieren wir die Verwendung von Cobalamin(Vitamin B12)-bindenden Domänen von bakteriellen CarH-Transkriptionsfaktoren zur Grünlicht-induzierten Dissoziation von Rezeptoren. Fusioniert mit dem Fibroblasten-W achstumsfaktor-Rezeptor 1 führten diese im Dunkeln in kultivierten Zellen zu Signalaktivität durch Oligomerisierung, welche durch Beleuchten umgehend aufgehoben wurde. In Zebrafischembryonen, die einen derartigen Rezeptor exprimieren, ermöglichte grünes Licht die Kontrolle über abnormale Signalaktivität während der Embryonalentwicklung. }, author = {Kainrath, Stephanie and Stadler, Manuela and Gschaider-Reichhart, Eva and Distel, Martin and Janovjak, Harald L}, journal = {Angewandte Chemie}, number = {16}, pages = {4679 -- 4682}, publisher = {Wiley}, title = {{Grünlicht-induzierte Rezeptorinaktivierung durch Cobalamin-bindende Domänen}}, doi = {10.1002/ange.201611998}, volume = {129}, year = {2017}, } @article{570, abstract = {Most phenotypes are determined by molecular systems composed of specifically interacting molecules. However, unlike for individual components, little is known about the distributions of mutational effects of molecular systems as a whole. We ask how the distribution of mutational effects of a transcriptional regulatory system differs from the distributions of its components, by first independently, and then simultaneously, mutating a transcription factor and the associated promoter it represses. We find that the system distribution exhibits increased phenotypic variation compared to individual component distributions - an effect arising from intermolecular epistasis between the transcription factor and its DNA-binding site. In large part, this epistasis can be qualitatively attributed to the structure of the transcriptional regulatory system and could therefore be a common feature in prokaryotes. Counter-intuitively, intermolecular epistasis can alleviate the constraints of individual components, thereby increasing phenotypic variation that selection could act on and facilitating adaptive evolution. }, author = {Lagator, Mato and Sarikas, Srdjan and Acar, Hande and Bollback, Jonathan P and Guet, Calin C}, issn = {2050084X}, journal = {eLife}, publisher = {eLife Sciences Publications}, title = {{Regulatory network structure determines patterns of intermolecular epistasis}}, doi = {10.7554/eLife.28921}, volume = {6}, year = {2017}, } @article{613, abstract = {Bacteria in groups vary individually, and interact with other bacteria and the environment to produce population-level patterns of gene expression. Investigating such behavior in detail requires measuring and controlling populations at the single-cell level alongside precisely specified interactions and environmental characteristics. Here we present an automated, programmable platform that combines image-based gene expression and growth measurements with on-line optogenetic expression control for hundreds of individual Escherichia coli cells over days, in a dynamically adjustable environment. This integrated platform broadly enables experiments that bridge individual and population behaviors. We demonstrate: (i) population structuring by independent closed-loop control of gene expression in many individual cells, (ii) cell-cell variation control during antibiotic perturbation, (iii) hybrid bio-digital circuits in single cells, and freely specifiable digital communication between individual bacteria. These examples showcase the potential for real-time integration of theoretical models with measurement and control of many individual cells to investigate and engineer microbial population behavior.}, author = {Chait, Remy P and Ruess, Jakob and Bergmiller, Tobias and Tkacik, Gasper and Guet, Calin C}, issn = {20411723}, journal = {Nature Communications}, number = {1}, publisher = {Nature Publishing Group}, title = {{Shaping bacterial population behavior through computer interfaced control of individual cells}}, doi = {10.1038/s41467-017-01683-1}, volume = {8}, year = {2017}, } @article{624, abstract = {Bacteria adapt to adverse environmental conditions by altering gene expression patterns. Recently, a novel stress adaptation mechanism has been described that allows Escherichia coli to alter gene expression at the post-transcriptional level. The key player in this regulatory pathway is the endoribonuclease MazF, the toxin component of the toxin-antitoxin module mazEF that is triggered by various stressful conditions. In general, MazF degrades the majority of transcripts by cleaving at ACA sites, which results in the retardation of bacterial growth. Furthermore, MazF can process a small subset of mRNAs and render them leaderless by removing their ribosome binding site. MazF concomitantly modifies ribosomes, making them selective for the translation of leaderless mRNAs. In this study, we employed fluorescent reporter-systems to investigate mazEF expression during stressful conditions, and to infer consequences of the mRNA processing mediated by MazF on gene expression at the single-cell level. Our results suggest that mazEF transcription is maintained at low levels in single cells encountering adverse conditions, such as antibiotic stress or amino acid starvation. Moreover, using the grcA mRNA as a model for MazF-mediated mRNA processing, we found that MazF activation promotes heterogeneity in the grcA reporter expression, resulting in a subpopulation of cells with increased levels of GrcA reporter protein.}, author = {Nikolic, Nela and Didara, Zrinka and Moll, Isabella}, issn = {21678359}, journal = {PeerJ}, number = {9}, publisher = {PeerJ}, title = {{MazF activation promotes translational heterogeneity of the grcA mRNA in Escherichia coli populations}}, doi = {10.7717/peerj.3830}, volume = {2017}, year = {2017}, } @article{655, abstract = {The bacterial flagellum is a self-assembling nanomachine. The external flagellar filament, several times longer than a bacterial cell body, is made of a few tens of thousands subunits of a single protein: flagellin. A fundamental problem concerns the molecular mechanism of how the flagellum grows outside the cell, where no discernible energy source is available. Here, we monitored the dynamic assembly of individual flagella using in situ labelling and real-time immunostaining of elongating flagellar filaments. We report that the rate of flagellum growth, initially ~1,700 amino acids per second, decreases with length and that the previously proposed chain mechanism does not contribute to the filament elongation dynamics. Inhibition of the proton motive force-dependent export apparatus revealed a major contribution of substrate injection in driving filament elongation. The combination of experimental and mathematical evidence demonstrates that a simple, injection-diffusion mechanism controls bacterial flagella growth outside the cell.}, author = {Renault, Thibaud and Abraham, Anthony and Bergmiller, Tobias and Paradis, Guillaume and Rainville, Simon and Charpentier, Emmanuelle and Guet, Calin C and Tu, Yuhai and Namba, Keiichi and Keener, James and Minamino, Tohru and Erhardt, Marc}, issn = {2050084X}, journal = {eLife}, publisher = {eLife Sciences Publications}, title = {{Bacterial flagella grow through an injection diffusion mechanism}}, doi = {10.7554/eLife.23136}, volume = {6}, year = {2017}, } @article{541, abstract = {While we have good understanding of bacterial metabolism at the population level, we know little about the metabolic behavior of individual cells: do single cells in clonal populations sometimes specialize on different metabolic pathways? Such metabolic specialization could be driven by stochastic gene expression and could provide individual cells with growth benefits of specialization. We measured the degree of phenotypic specialization in two parallel metabolic pathways, the assimilation of glucose and arabinose. We grew Escherichia coli in chemostats, and used isotope-labeled sugars in combination with nanometer-scale secondary ion mass spectrometry and mathematical modeling to quantify sugar assimilation at the single-cell level. We found large variation in metabolic activities between single cells, both in absolute assimilation and in the degree to which individual cells specialize in the assimilation of different sugars. Analysis of transcriptional reporters indicated that this variation was at least partially based on cell-to-cell variation in gene expression. Metabolic differences between cells in clonal populations could potentially reduce metabolic incompatibilities between different pathways, and increase the rate at which parallel reactions can be performed.}, author = {Nikolic, Nela and Schreiber, Frank and Dal Co, Alma and Kiviet, Daniel and Bergmiller, Tobias and Littmann, Sten and Kuypers, Marcel and Ackermann, Martin}, issn = {15537390}, journal = {PLoS Genetics}, number = {12}, publisher = {Public Library of Science}, title = {{Cell-to-cell variation and specialization in sugar metabolism in clonal bacterial populations}}, doi = {10.1371/journal.pgen.1007122}, volume = {13}, year = {2017}, } @misc{9847, abstract = {information on culture conditions, phage mutagenesis, verification and lysate preparation; Raw data}, author = {Pleska, Maros and Guet, Calin C}, publisher = {The Royal Society}, title = {{Supplementary materials and methods; Full data set from effects of mutations in phage restriction sites during escape from restriction–modification}}, doi = {10.6084/m9.figshare.5633917.v1}, year = {2017}, } @misc{9845, abstract = {Estimates of 13 C-arabinose and 2 H-glucose uptake from the fractions of heavy isotopes measured in single cells}, author = {Nikolic, Nela and Schreiber, Frank and Dal Co, Alma and Kiviet, Daniel and Bergmiller, Tobias and Littmann, Sten and Kuypers, Marcel and Ackermann, Martin}, publisher = {Public Library of Science}, title = {{Mathematical model}}, doi = {10.1371/journal.pgen.1007122.s017}, year = {2017}, } @misc{9849, abstract = {This text provides additional information about the model, a derivation of the analytic results in Eq (4), and details about simulations of an additional parameter set.}, author = {Lukacisinova, Marta and Novak, Sebastian and Paixao, Tiago}, publisher = {Public Library of Science}, title = {{Modelling and simulation details}}, doi = {10.1371/journal.pcbi.1005609.s001}, year = {2017}, } @misc{9850, abstract = {In this text, we discuss how a cost of resistance and the possibility of lethal mutations impact our model.}, author = {Lukacisinova, Marta and Novak, Sebastian and Paixao, Tiago}, publisher = {Public Library of Science}, title = {{Extensions of the model}}, doi = {10.1371/journal.pcbi.1005609.s002}, year = {2017}, } @misc{9846, author = {Nikolic, Nela and Schreiber, Frank and Dal Co, Alma and Kiviet, Daniel and Bergmiller, Tobias and Littmann, Sten and Kuypers, Marcel and Ackermann, Martin}, publisher = {Public Library of Science}, title = {{Supplementary methods}}, doi = {10.1371/journal.pgen.1007122.s016}, year = {2017}, } @misc{9851, abstract = {Based on the intuitive derivation of the dynamics of SIM allele frequency pM in the main text, we present a heuristic prediction for the long-term SIM allele frequencies with χ > 1 stresses and compare it to numerical simulations.}, author = {Lukacisinova, Marta and Novak, Sebastian and Paixao, Tiago}, publisher = {Public Library of Science}, title = {{Heuristic prediction for multiple stresses}}, doi = {10.1371/journal.pcbi.1005609.s003}, year = {2017}, } @misc{9852, abstract = {We show how different combination strategies affect the fraction of individuals that are multi-resistant.}, author = {Lukacisinova, Marta and Novak, Sebastian and Paixao, Tiago}, publisher = {Public Library of Science}, title = {{Resistance frequencies for different combination strategies}}, doi = {10.1371/journal.pcbi.1005609.s004}, year = {2017}, } @misc{9844, author = {Nikolic, Nela and Schreiber, Frank and Dal Co, Alma and Kiviet, Daniel and Bergmiller, Tobias and Littmann, Sten and Kuypers, Marcel and Ackermann, Martin}, publisher = {Public Library of Science}, title = {{Source data for figures and tables}}, doi = {10.1371/journal.pgen.1007122.s018}, year = {2017}, } @article{561, abstract = {Restriction–modification systems are widespread genetic elements that protect bacteria from bacteriophage infections by recognizing and cleaving heterologous DNA at short, well-defined sequences called restriction sites. Bioinformatic evidence shows that restriction sites are significantly underrepresented in bacteriophage genomes, presumably because bacteriophages with fewer restriction sites are more likely to escape cleavage by restriction–modification systems. However, how mutations in restriction sites affect the likelihood of bacteriophage escape is unknown. Using the bacteriophage l and the restriction–modification system EcoRI, we show that while mutation effects at different restriction sites are unequal, they are independent. As a result, the probability of bacteriophage escape increases with each mutated restriction site. Our results experimentally support the role of restriction site avoidance as a response to selection imposed by restriction–modification systems and offer an insight into the events underlying the process of bacteriophage escape.}, author = {Pleska, Maros and Guet, Calin C}, issn = {1744-9561}, journal = {Biology Letters}, number = {12}, publisher = {The Royal Society}, title = {{Effects of mutations in phage restriction sites during escape from restriction–modification}}, doi = {10.1098/rsbl.2017.0646}, volume = {13}, year = {2017}, } @phdthesis{202, abstract = {Restriction-modification (RM) represents the simplest and possibly the most widespread mechanism of self/non-self discrimination in nature. In order to provide bacteria with immunity against bacteriophages and other parasitic genetic elements, RM systems rely on a balance between two enzymes: the restriction enzyme, which cleaves non-self DNA at specific restriction sites, and the modification enzyme, which tags the host’s DNA as self and thus protects it from cleavage. In this thesis, I use population and single-cell level experiments in combination with mathematical modeling to study different aspects of the interplay between RM systems, bacteria and bacteriophages. First, I analyze how mutations in phage restriction sites affect the probability of phage escape – an inherently stochastic process, during which phages accidently get modified instead of restricted. Next, I use single-cell experiments to show that RM systems can, with a low probability, attack the genome of their bacterial host and that this primitive form of autoimmunity leads to a tradeoff between the evolutionary cost and benefit of RM systems. Finally, I investigate the nature of interactions between bacteria, RM systems and temperate bacteriophages to find that, as a consequence of phage escape and its impact on population dynamics, RM systems can promote acquisition of symbiotic bacteriophages, rather than limit it. The results presented here uncover new fundamental biological properties of RM systems and highlight their importance in the ecology and evolution of bacteria, bacteriophages and their interactions.}, author = {Pleska, Maros}, issn = {2663-337X}, pages = {126}, publisher = {Institute of Science and Technology Austria}, title = {{Biology of restriction-modification systems at the single-cell and population level}}, doi = {10.15479/AT:ISTA:th_916}, year = {2017}, } @article{1351, abstract = {The behaviour of gene regulatory networks (GRNs) is typically analysed using simulation-based statistical testing-like methods. In this paper, we demonstrate that we can replace this approach by a formal verification-like method that gives higher assurance and scalability. We focus on Wagner’s weighted GRN model with varying weights, which is used in evolutionary biology. In the model, weight parameters represent the gene interaction strength that may change due to genetic mutations. For a property of interest, we synthesise the constraints over the parameter space that represent the set of GRNs satisfying the property. We experimentally show that our parameter synthesis procedure computes the mutational robustness of GRNs—an important problem of interest in evolutionary biology—more efficiently than the classical simulation method. We specify the property in linear temporal logic. We employ symbolic bounded model checking and SMT solving to compute the space of GRNs that satisfy the property, which amounts to synthesizing a set of linear constraints on the weights.}, author = {Giacobbe, Mirco and Guet, Calin C and Gupta, Ashutosh and Henzinger, Thomas A and Paixao, Tiago and Petrov, Tatjana}, issn = {00015903}, journal = {Acta Informatica}, number = {8}, pages = {765 -- 787}, publisher = {Springer}, title = {{Model checking the evolution of gene regulatory networks}}, doi = {10.1007/s00236-016-0278-x}, volume = {54}, year = {2017}, } @article{1336, abstract = {Evolutionary algorithms (EAs) form a popular optimisation paradigm inspired by natural evolution. In recent years the field of evolutionary computation has developed a rigorous analytical theory to analyse the runtimes of EAs on many illustrative problems. Here we apply this theory to a simple model of natural evolution. In the Strong Selection Weak Mutation (SSWM) evolutionary regime the time between occurrences of new mutations is much longer than the time it takes for a mutated genotype to take over the population. In this situation, the population only contains copies of one genotype and evolution can be modelled as a stochastic process evolving one genotype by means of mutation and selection between the resident and the mutated genotype. The probability of accepting the mutated genotype then depends on the change in fitness. We study this process, SSWM, from an algorithmic perspective, quantifying its expected optimisation time for various parameters and investigating differences to a similar evolutionary algorithm, the well-known (1+1) EA. We show that SSWM can have a moderate advantage over the (1+1) EA at crossing fitness valleys and study an example where SSWM outperforms the (1+1) EA by taking advantage of information on the fitness gradient.}, author = {Paixao, Tiago and Pérez Heredia, Jorge and Sudholt, Dirk and Trubenova, Barbora}, issn = {01784617}, journal = {Algorithmica}, number = {2}, pages = {681 -- 713}, publisher = {Springer}, title = {{Towards a runtime comparison of natural and artificial evolution}}, doi = {10.1007/s00453-016-0212-1}, volume = {78}, year = {2017}, } @article{1084, abstract = {BceRS and PsdRS are paralogous two-component systems in Bacillus subtilis controlling the response to antimicrobial peptides. In the presence of extracellular bacitracin and nisin, respectively, the two response regulators (RRs) bind their target promoters, PbceA or PpsdA, resulting in a strong up-regulation of target gene expression and ultimately antibiotic resistance. Despite high sequence similarity between the RRs BceR and PsdR and their known binding sites, no cross-regulation has been observed between them. We therefore investigated the specificity determinants of PbceA and PpsdA that ensure the insulation of these two paralogous pathways at the RR–promoter interface. In vivo and in vitro analyses demonstrate that the regulatory regions within these two promoters contain three important elements: in addition to the known (main) binding site, we identified a linker region and a secondary binding site that are crucial for functionality. Initial binding to the high-affinity, low-specificity main binding site is a prerequisite for the subsequent highly specific binding of a second RR dimer to the low-affinity secondary binding site. In addition to this hierarchical cooperative binding, discrimination requires a competition of the two RRs for their respective binding site mediated by only slight differences in binding affinities.}, author = {Fang, Chong and Nagy-Staron, Anna A and Grafe, Martin and Heermann, Ralf and Jung, Kirsten and Gebhard, Susanne and Mascher, Thorsten}, issn = { 0950382X}, journal = {Molecular Microbiology}, number = {1}, pages = {16 -- 31}, publisher = {Wiley-Blackwell}, title = {{Insulation and wiring specificity of BceR like response regulators and their target promoters in Bacillus subtilis}}, doi = {10.1111/mmi.13597}, volume = {104}, year = {2017}, } @article{954, abstract = {Understanding the relation between genotype and phenotype remains a major challenge. The difficulty of predicting individual mutation effects, and particularly the interactions between them, has prevented the development of a comprehensive theory that links genotypic changes to their phenotypic effects. We show that a general thermodynamic framework for gene regulation, based on a biophysical understanding of protein-DNA binding, accurately predicts the sign of epistasis in a canonical cis-regulatory element consisting of overlapping RNA polymerase and repressor binding sites. Sign and magnitude of individual mutation effects are sufficient to predict the sign of epistasis and its environmental dependence. Thus, the thermodynamic model offers the correct null prediction for epistasis between mutations across DNA-binding sites. Our results indicate that a predictive theory for the effects of cis-regulatory mutations is possible from first principles, as long as the essential molecular mechanisms and the constraints these impose on a biological system are accounted for.}, author = {Lagator, Mato and Paixao, Tiago and Barton, Nicholas H and Bollback, Jonathan P and Guet, Calin C}, issn = {2050084X}, journal = {eLife}, publisher = {eLife Sciences Publications}, title = {{On the mechanistic nature of epistasis in a canonical cis-regulatory element}}, doi = {10.7554/eLife.25192}, volume = {6}, year = {2017}, } @article{1007, abstract = {A nonlinear system possesses an invariance with respect to a set of transformations if its output dynamics remain invariant when transforming the input, and adjusting the initial condition accordingly. Most research has focused on invariances with respect to time-independent pointwise transformations like translational-invariance (u(t) -> u(t) + p, p in R) or scale-invariance (u(t) -> pu(t), p in R>0). In this article, we introduce the concept of s0-invariances with respect to continuous input transformations exponentially growing/decaying over time. We show that s0-invariant systems not only encompass linear time-invariant (LTI) systems with transfer functions having an irreducible zero at s0 in R, but also that the input/output relationship of nonlinear s0-invariant systems possesses properties well known from their linear counterparts. Furthermore, we extend the concept of s0-invariances to second- and higher-order s0-invariances, corresponding to invariances with respect to transformations of the time-derivatives of the input, and encompassing LTI systems with zeros of multiplicity two or higher. Finally, we show that nth-order 0-invariant systems realize – under mild conditions – nth-order nonlinear differential operators: when excited by an input of a characteristic functional form, the system’s output converges to a constant value only depending on the nth (nonlinear) derivative of the input.}, author = {Lang, Moritz and Sontag, Eduardo}, issn = {0005-1098}, journal = {Automatica}, pages = {46 -- 55}, publisher = {International Federation of Automatic Control}, title = {{Zeros of nonlinear systems with input invariances}}, doi = {10.1016/j.automatica.2017.03.030}, volume = {81C}, year = {2017}, } @misc{5564, abstract = {Compressed Fastq files with whole-genome sequencing data of IS-wt strain D and clones from four evolved populations (A11, C08, C10, D08). Information on this data collection is available in the Methods Section of the primary publication.}, author = {Steinrück, Magdalena and Guet, Calin C}, publisher = {Institute of Science and Technology Austria}, title = {{Fastq files for "Complex chromosomal neighborhood effects determine the adaptive potential of a gene under selection"}}, doi = {10.15479/AT:ISTA:65}, year = {2017}, } @misc{5560, abstract = {This repository contains the data collected for the manuscript "Biased partitioning of the multi-drug efflux pump AcrAB-TolC underlies long-lived phenotypic heterogeneity". The data is compressed into a single archive. Within the archive, different folders correspond to figures of the main text and the SI of the related publication. Data is saved as plain text, with each folder containing a separate readme file describing the format. Typically, the data is from fluorescence microscopy measurements of single cells growing in a microfluidic "mother machine" device, and consists of relevant values (primarily arbitrary unit or normalized fluorescence measurements, and division times / growth rates) after raw microscopy images have been processed, segmented, and their features extracted, as described in the methods section of the related publication.}, author = {Bergmiller, Tobias and Andersson, Anna M and Tomasek, Kathrin and Balleza, Enrique and Kiviet, Daniel and Hauschild, Robert and Tkacik, Gasper and Guet, Calin C}, keywords = {single cell microscopy, mother machine microfluidic device, AcrAB-TolC pump, multi-drug efflux, Escherichia coli}, publisher = {Institute of Science and Technology Austria}, title = {{Biased partitioning of the multi-drug efflux pump AcrAB-TolC underlies long-lived phenotypic heterogeneity}}, doi = {10.15479/AT:ISTA:53}, year = {2017}, } @article{665, abstract = {The molecular mechanisms underlying phenotypic variation in isogenic bacterial populations remain poorly understood.We report that AcrAB-TolC, the main multidrug efflux pump of Escherichia coli, exhibits a strong partitioning bias for old cell poles by a segregation mechanism that is mediated by ternary AcrAB-TolC complex formation. Mother cells inheriting old poles are phenotypically distinct and display increased drug efflux activity relative to daughters. Consequently, we find systematic and long-lived growth differences between mother and daughter cells in the presence of subinhibitory drug concentrations. A simple model for biased partitioning predicts a population structure of long-lived and highly heterogeneous phenotypes. This straightforward mechanism of generating sustained growth rate differences at subinhibitory antibiotic concentrations has implications for understanding the emergence of multidrug resistance in bacteria.}, author = {Bergmiller, Tobias and Andersson, Anna M and Tomasek, Kathrin and Balleza, Enrique and Kiviet, Daniel and Hauschild, Robert and Tkacik, Gasper and Guet, Calin C}, issn = {00368075}, journal = {Science}, number = {6335}, pages = {311 -- 315}, publisher = {American Association for the Advancement of Science}, title = {{Biased partitioning of the multidrug efflux pump AcrAB TolC underlies long lived phenotypic heterogeneity}}, doi = {10.1126/science.aaf4762}, volume = {356}, year = {2017}, } @article{1028, abstract = {Optogenetics and photopharmacology provide spatiotemporally precise control over protein interactions and protein function in cells and animals. Optogenetic methods that are sensitive to green light and can be used to break protein complexes are not broadly available but would enable multichromatic experiments with previously inaccessible biological targets. Herein, we repurposed cobalamin (vitamin B12) binding domains of bacterial CarH transcription factors for green-light-induced receptor dissociation. In cultured cells, we observed oligomerization-induced cell signaling for the fibroblast growth factor receptor 1 fused to cobalamin-binding domains in the dark that was rapidly eliminated upon illumination. In zebrafish embryos expressing fusion receptors, green light endowed control over aberrant fibroblast growth factor signaling during development. Green-light-induced domain dissociation and light-inactivated receptors will critically expand the optogenetic toolbox for control of biological processes.}, author = {Kainrath, Stephanie and Stadler, Manuela and Gschaider-Reichhart, Eva and Distel, Martin and Janovjak, Harald L}, issn = {14337851}, journal = {Angewandte Chemie - International Edition}, number = {16}, pages = {4608--4611}, publisher = {Wiley-Blackwell}, title = {{Green-light-induced inactivation of receptor signaling using cobalamin-binding domains}}, doi = {10.1002/anie.201611998}, volume = {56}, year = {2017}, } @article{704, abstract = {How the organization of genes on a chromosome shapes adaptation is essential for understanding evolutionary paths. Here, we investigate how adaptation to rapidly increasing levels of antibiotic depends on the chromosomal neighborhood of a drug-resistance gene inserted at different positions of the Escherichia coli chromosome. Using a dual-fluorescence reporter that allows us to distinguish gene amplifications from other up-mutations, we track in real-time adaptive changes in expression of the drug-resistance gene. We find that the relative contribution of several mutation types differs systematically between loci due to properties of neighboring genes: essentiality, expression, orientation, termination, and presence of duplicates. These properties determine rate and fitness effects of gene amplification, deletions, and mutations compromising transcriptional termination. Thus, the adaptive potential of a gene under selection is a system-property with a complex genetic basis that is specific for each chromosomal locus, and it can be inferred from detailed functional and genomic data.}, author = {Steinrück, Magdalena and Guet, Calin C}, issn = {2050084X}, journal = {eLife}, publisher = {eLife Sciences Publications}, title = {{Complex chromosomal neighborhood effects determine the adaptive potential of a gene under selection}}, doi = {10.7554/eLife.25100}, volume = {6}, year = {2017}, } @article{696, abstract = {Mutator strains are expected to evolve when the availability and effect of beneficial mutations are high enough to counteract the disadvantage from deleterious mutations that will inevitably accumulate. As the population becomes more adapted to its environment, both availability and effect of beneficial mutations necessarily decrease and mutation rates are predicted to decrease. It has been shown that certain molecular mechanisms can lead to increased mutation rates when the organism finds itself in a stressful environment. While this may be a correlated response to other functions, it could also be an adaptive mechanism, raising mutation rates only when it is most advantageous. Here, we use a mathematical model to investigate the plausibility of the adaptive hypothesis. We show that such a mechanism can be mantained if the population is subjected to diverse stresses. By simulating various antibiotic treatment schemes, we find that combination treatments can reduce the effectiveness of second-order selection on stress-induced mutagenesis. We discuss the implications of our results to strategies of antibiotic therapy.}, author = {Lukacisinova, Marta and Novak, Sebastian and Paixao, Tiago}, issn = {1553734X}, journal = {PLoS Computational Biology}, number = {7}, publisher = {Public Library of Science}, title = {{Stress induced mutagenesis: Stress diversity facilitates the persistence of mutator genes}}, doi = {10.1371/journal.pcbi.1005609}, volume = {13}, year = {2017}, } @article{735, abstract = {Cell-cell contact formation constitutes an essential step in evolution, leading to the differentiation of specialized cell types. However, remarkably little is known about whether and how the interplay between contact formation and fate specification affects development. Here, we identify a positive feedback loop between cell-cell contact duration, morphogen signaling, and mesendoderm cell-fate specification during zebrafish gastrulation. We show that long-lasting cell-cell contacts enhance the competence of prechordal plate (ppl) progenitor cells to respond to Nodal signaling, required for ppl cell-fate specification. We further show that Nodal signaling promotes ppl cell-cell contact duration, generating a positive feedback loop between ppl cell-cell contact duration and cell-fate specification. Finally, by combining mathematical modeling and experimentation, we show that this feedback determines whether anterior axial mesendoderm cells become ppl or, instead, turn into endoderm. Thus, the interdependent activities of cell-cell signaling and contact formation control fate diversification within the developing embryo.}, author = {Barone, Vanessa and Lang, Moritz and Krens, Gabriel and Pradhan, Saurabh and Shamipour, Shayan and Sako, Keisuke and Sikora, Mateusz K and Guet, Calin C and Heisenberg, Carl-Philipp J}, issn = {15345807}, journal = {Developmental Cell}, number = {2}, pages = {198 -- 211}, publisher = {Cell Press}, title = {{An effective feedback loop between cell-cell contact duration and morphogen signaling determines cell fate}}, doi = {10.1016/j.devcel.2017.09.014}, volume = {43}, year = {2017}, } @article{1008, abstract = {Feedback loops in biological networks, among others, enable differentiation and cell cycle progression, and increase robustness in signal transduction. In natural networks, feedback loops are often complex and intertwined, making it challenging to identify which loops are mainly responsible for an observed behavior. However, minimal synthetic replicas could allow for such identification. Here, we engineered a synthetic permease-inducer-repressor system in Saccharomyces cerevisiae to analyze if a transport-mediated positive feedback loop could be a core mechanism for the switch-like behavior in the regulation of metabolic gene networks such as the S. cerevisiae GAL system or the Escherichia coli lac operon. We characterized the synthetic circuit using deterministic and stochastic mathematical models. Similar to its natural counterparts, our synthetic system shows bistable and hysteretic behavior, and the inducer concentration range for bistability as well as the switching rates between the two stable states depend on the repressor concentration. Our results indicate that a generic permease–inducer–repressor circuit with a single feedback loop is sufficient to explain the experimentally observed bistable behavior of the natural systems. We anticipate that the approach of reimplementing natural systems with orthogonal parts to identify crucial network components is applicable to other natural systems such as signaling pathways.}, author = {Gnügge, Robert and Dharmarajan, Lekshmi and Lang, Moritz and Stelling, Jörg}, journal = {ACS Synthetic Biology}, number = {10}, pages = {1098 -- 1107}, publisher = {American Chemical Society}, title = {{An orthogonal permease–inducer–repressor feedback loop shows bistability}}, doi = {10.1021/acssynbio.6b00013}, volume = {5}, year = {2016}, } @article{1170, abstract = {The increasing complexity of dynamic models in systems and synthetic biology poses computational challenges especially for the identification of model parameters. While modularization of the corresponding optimization problems could help reduce the “curse of dimensionality,” abundant feedback and crosstalk mechanisms prohibit a simple decomposition of most biomolecular networks into subnetworks, or modules. Drawing on ideas from network modularization and multiple-shooting optimization, we present here a modular parameter identification approach that explicitly allows for such interdependencies. Interfaces between our modules are given by the experimentally measured molecular species. This definition allows deriving good (initial) estimates for the inter-module communication directly from the experimental data. Given these estimates, the states and parameter sensitivities of different modules can be integrated independently. To achieve consistency between modules, we iteratively adjust the estimates for inter-module communication while optimizing the parameters. After convergence to an optimal parameter set---but not during earlier iterations---the intermodule communication as well as the individual modules\' state dynamics agree with the dynamics of the nonmodularized network. Our modular parameter identification approach allows for easy parallelization; it can reduce the computational complexity for larger networks and decrease the probability to converge to suboptimal local minima. We demonstrate the algorithm\'s performance in parameter estimation for two biomolecular networks, a synthetic genetic oscillator and a mammalian signaling pathway.}, author = {Lang, Moritz and Stelling, Jörg}, journal = {SIAM Journal on Scientific Computing}, number = {6}, pages = {B988 -- B1008}, publisher = {Society for Industrial and Applied Mathematics }, title = {{Modular parameter identification of biomolecular networks}}, doi = {10.1137/15M103306X}, volume = {38}, year = {2016}, } @inproceedings{1220, abstract = {Theoretical and numerical aspects of aerodynamic efficiency of propulsion systems coupled to the boundary layer of a fuselage are studied. We discuss the effects of local flow fields, which are affected both by conservative flow acceleration as well as total pressure losses, on the efficiency of boundary layer immersed propulsion devices. We introduce the concept of a boundary layer retardation turbine that helps reduce skin friction over the fuselage. We numerically investigate efficiency gains offered by boundary layer and wake interacting devices. We discuss the results in terms of a total energy consumption framework and show that efficiency gains of any device depend on all the other elements of the propulsion system.}, author = {Mikić, Gregor and Stoll, Alex and Bevirt, Joe and Grah, Rok and Moore, Mark}, location = {Washington, D.C., USA}, pages = {1 -- 19}, publisher = {AIAA}, title = {{Fuselage boundary layer ingestion propulsion applied to a thin haul commuter aircraft for optimal efficiency}}, doi = {10.2514/6.2016-3764}, year = {2016}, }