@article{10812, abstract = {Several promising strategies based on combining or cycling different antibiotics have been proposed to increase efficacy and counteract resistance evolution, but we still lack a deep understanding of the physiological responses and genetic mechanisms that underlie antibiotic interactions and the clinical applicability of these strategies. In antibiotic-exposed bacteria, the combined effects of physiological stress responses and emerging resistance mutations (occurring at different time scales) generate complex and often unpredictable dynamics. In this Review, we present our current understanding of bacterial cell physiology and genetics of responses to antibiotics. We emphasize recently discovered mechanisms of synergistic and antagonistic drug interactions, hysteresis in temporal interactions between antibiotics that arise from microbial physiology and interactions between antibiotics and resistance mutations that can cause collateral sensitivity or cross-resistance. We discuss possible connections between the different phenomena and indicate relevant research directions. A better and more unified understanding of drug and genetic interactions is likely to advance antibiotic therapy.}, author = {Römhild, Roderich and Bollenbach, Mark Tobias and Andersson, Dan I.}, issn = {1740-1534}, journal = {Nature Reviews Microbiology}, keywords = {General Immunology and Microbiology, Microbiology, Infectious Diseases}, pages = {478--490}, publisher = {Springer Nature}, title = {{The physiology and genetics of bacterial responses to antibiotic combinations}}, doi = {10.1038/s41579-022-00700-5}, volume = {20}, year = {2022}, } @article{11341, abstract = {Intragenic regions that are removed during maturation of the RNA transcript—introns—are universally present in the nuclear genomes of eukaryotes1. The budding yeast, an otherwise intron-poor species, preserves two sets of ribosomal protein genes that differ primarily in their introns2,3. Although studies have shed light on the role of ribosomal protein introns under stress and starvation4,5,6, understanding the contribution of introns to ribosome regulation remains challenging. Here, by combining isogrowth profiling7 with single-cell protein measurements8, we show that introns can mediate inducible phenotypic heterogeneity that confers a clear fitness advantage. Osmotic stress leads to bimodal expression of the small ribosomal subunit protein Rps22B, which is mediated by an intron in the 5′ untranslated region of its transcript. The two resulting yeast subpopulations differ in their ability to cope with starvation. Low levels of Rps22B protein result in prolonged survival under sustained starvation, whereas high levels of Rps22B enable cells to grow faster after transient starvation. Furthermore, yeasts growing at high concentrations of sugar, similar to those in ripe grapes, exhibit bimodal expression of Rps22B when approaching the stationary phase. Differential intron-mediated regulation of ribosomal protein genes thus provides a way to diversify the population when starvation threatens in natural environments. Our findings reveal a role for introns in inducing phenotypic heterogeneity in changing environments, and suggest that duplicated ribosomal protein genes in yeast contribute to resolving the evolutionary conflict between precise expression control and environmental responsiveness9.}, author = {Lukacisin, Martin and Espinosa-Cantú, Adriana and Bollenbach, Mark Tobias}, issn = {1476-4687}, journal = {Nature}, pages = {113--118}, publisher = {Springer Nature}, title = {{Intron-mediated induction of phenotypic heterogeneity}}, doi = {10.1038/s41586-022-04633-0}, volume = {605}, year = {2022}, } @article{12261, abstract = {Dose–response relationships are a general concept for quantitatively describing biological systems across multiple scales, from the molecular to the whole-cell level. A clinically relevant example is the bacterial growth response to antibiotics, which is routinely characterized by dose–response curves. The shape of the dose–response curve varies drastically between antibiotics and plays a key role in treatment, drug interactions, and resistance evolution. However, the mechanisms shaping the dose–response curve remain largely unclear. Here, we show in Escherichia coli that the distinctively shallow dose–response curve of the antibiotic trimethoprim is caused by a negative growth-mediated feedback loop: Trimethoprim slows growth, which in turn weakens the effect of this antibiotic. At the molecular level, this feedback is caused by the upregulation of the drug target dihydrofolate reductase (FolA/DHFR). We show that this upregulation is not a specific response to trimethoprim but follows a universal trend line that depends primarily on the growth rate, irrespective of its cause. Rewiring the feedback loop alters the dose–response curve in a predictable manner, which we corroborate using a mathematical model of cellular resource allocation and growth. Our results indicate that growth-mediated feedback loops may shape drug responses more generally and could be exploited to design evolutionary traps that enable selection against drug resistance.}, author = {Angermayr, Andreas and Pang, Tin Yau and Chevereau, Guillaume and Mitosch, Karin and Lercher, Martin J and Bollenbach, Mark Tobias}, issn = {1744-4292}, journal = {Molecular Systems Biology}, keywords = {Applied Mathematics, Computational Theory and Mathematics, General Agricultural and Biological Sciences, General Immunology and Microbiology, General Biochemistry, Genetics and Molecular Biology, Information Systems}, number = {9}, publisher = {Embo Press}, title = {{Growth‐mediated negative feedback shapes quantitative antibiotic response}}, doi = {10.15252/msb.202110490}, volume = {18}, year = {2022}, } @article{10271, abstract = {Understanding interactions between antibiotics used in combination is an important theme in microbiology. Using the interactions between the antifolate drug trimethoprim and the ribosome-targeting antibiotic erythromycin in Escherichia coli as a model, we applied a transcriptomic approach for dissecting interactions between two antibiotics with different modes of action. When trimethoprim and erythromycin were combined, the transcriptional response of genes from the sulfate reduction pathway deviated from the dominant effect of trimethoprim on the transcriptome. We successfully altered the drug interaction from additivity to suppression by increasing the sulfate level in the growth environment and identified sulfate reduction as an important metabolic determinant that shapes the interaction between the two drugs. Our work highlights the potential of using prioritization of gene expression patterns as a tool for identifying key metabolic determinants that shape drug-drug interactions. We further demonstrated that the sigma factor-binding protein gene crl shapes the interactions between the two antibiotics, which provides a rare example of how naturally occurring variations between strains of the same bacterial species can sometimes generate very different drug interactions.}, author = {Qi, Qin and Angermayr, S. Andreas and Bollenbach, Mark Tobias}, issn = {1664-302X}, journal = {Frontiers in Microbiology}, keywords = {microbiology}, publisher = {Frontiers}, title = {{Uncovering Key Metabolic Determinants of the Drug Interactions Between Trimethoprim and Erythromycin in Escherichia coli}}, doi = {10.3389/fmicb.2021.760017}, volume = {12}, year = {2021}, } @article{8997, abstract = {Phenomenological relations such as Ohm’s or Fourier’s law have a venerable history in physics but are still scarce in biology. This situation restrains predictive theory. Here, we build on bacterial “growth laws,” which capture physiological feedback between translation and cell growth, to construct a minimal biophysical model for the combined action of ribosome-targeting antibiotics. Our model predicts drug interactions like antagonism or synergy solely from responses to individual drugs. We provide analytical results for limiting cases, which agree well with numerical results. We systematically refine the model by including direct physical interactions of different antibiotics on the ribosome. In a limiting case, our model provides a mechanistic underpinning for recent predictions of higher-order interactions that were derived using entropy maximization. We further refine the model to include the effects of antibiotics that mimic starvation and the presence of resistance genes. We describe the impact of a starvation-mimicking antibiotic on drug interactions analytically and verify it experimentally. Our extended model suggests a change in the type of drug interaction that depends on the strength of resistance, which challenges established rescaling paradigms. We experimentally show that the presence of unregulated resistance genes can lead to altered drug interaction, which agrees with the prediction of the model. While minimal, the model is readily adaptable and opens the door to predicting interactions of second and higher-order in a broad range of biological systems.}, author = {Kavcic, Bor and Tkačik, Gašper and Bollenbach, Tobias}, issn = {1553-7358}, journal = {PLOS Computational Biology}, keywords = {Modelling and Simulation, Genetics, Molecular Biology, Antibiotics, Drug interactions}, publisher = {Public Library of Science}, title = {{Minimal biophysical model of combined antibiotic action}}, doi = {10.1371/journal.pcbi.1008529}, volume = {17}, year = {2021}, } @article{8037, abstract = {Genetic perturbations that affect bacterial resistance to antibiotics have been characterized genome-wide, but how do such perturbations interact with subsequent evolutionary adaptation to the drug? Here, we show that strong epistasis between resistance mutations and systematically identified genes can be exploited to control spontaneous resistance evolution. We evolved hundreds of Escherichia coli K-12 mutant populations in parallel, using a robotic platform that tightly controls population size and selection pressure. We find a global diminishing-returns epistasis pattern: strains that are initially more sensitive generally undergo larger resistance gains. However, some gene deletion strains deviate from this general trend and curtail the evolvability of resistance, including deletions of genes for membrane transport, LPS biosynthesis, and chaperones. Deletions of efflux pump genes force evolution on inferior mutational paths, not explored in the wild type, and some of these essentially block resistance evolution. This effect is due to strong negative epistasis with resistance mutations. The identified genes and cellular functions provide potential targets for development of adjuvants that may block spontaneous resistance evolution when combined with antibiotics.}, author = {Lukacisinova, Marta and Fernando, Booshini and Bollenbach, Mark Tobias}, issn = {20411723}, journal = {Nature Communications}, publisher = {Springer Nature}, title = {{Highly parallel lab evolution reveals that epistasis can curb the evolution of antibiotic resistance}}, doi = {10.1038/s41467-020-16932-z}, volume = {11}, year = {2020}, } @article{8250, abstract = {Antibiotics that interfere with translation, when combined, interact in diverse and difficult-to-predict ways. Here, we explain these interactions by “translation bottlenecks”: points in the translation cycle where antibiotics block ribosomal progression. To elucidate the underlying mechanisms of drug interactions between translation inhibitors, we generate translation bottlenecks genetically using inducible control of translation factors that regulate well-defined translation cycle steps. These perturbations accurately mimic antibiotic action and drug interactions, supporting that the interplay of different translation bottlenecks causes these interactions. We further show that growth laws, combined with drug uptake and binding kinetics, enable the direct prediction of a large fraction of observed interactions, yet fail to predict suppression. However, varying two translation bottlenecks simultaneously supports that dense traffic of ribosomes and competition for translation factors account for the previously unexplained suppression. These results highlight the importance of “continuous epistasis” in bacterial physiology.}, author = {Kavcic, Bor and Tkačik, Gašper and Bollenbach, Tobias}, issn = {2041-1723}, journal = {Nature Communications}, publisher = {Springer Nature}, title = {{Mechanisms of drug interactions between translation-inhibiting antibiotics}}, doi = {10.1038/s41467-020-17734-z}, volume = {11}, year = {2020}, } @unpublished{7673, abstract = {Combining drugs can improve the efficacy of treatments. However, predicting the effect of drug combinations is still challenging. The combined potency of drugs determines the drug interaction, which is classified as synergistic, additive, antagonistic, or suppressive. While probabilistic, non-mechanistic models exist, there is currently no biophysical model that can predict antibiotic interactions. Here, we present a physiologically relevant model of the combined action of antibiotics that inhibit protein synthesis by targeting the ribosome. This model captures the kinetics of antibiotic binding and transport, and uses bacterial growth laws to predict growth in the presence of antibiotic combinations. We find that this biophysical model can produce all drug interaction types except suppression. We show analytically that antibiotics which cannot bind to the ribosome simultaneously generally act as substitutes for one another, leading to additive drug interactions. Previously proposed null expectations for higher-order drug interactions follow as a limiting case of our model. We further extend the model to include the effects of direct physical or allosteric interactions between individual drugs on the ribosome. Notably, such direct interactions profoundly change the combined drug effect, depending on the kinetic parameters of the drugs used. The model makes additional predictions for the effects of resistance genes on drug interactions and for interactions between ribosome-targeting antibiotics and antibiotics with other targets. These findings enhance our understanding of the interplay between drug action and cell physiology and are a key step toward a general framework for predicting drug interactions.}, author = {Kavcic, Bor and Tkačik, Gašper and Bollenbach, Tobias}, booktitle = {bioRxiv}, publisher = {Cold Spring Harbor Laboratory}, title = {{A minimal biophysical model of combined antibiotic action}}, doi = {10.1101/2020.04.18.047886}, year = {2020}, } @article{6046, abstract = {Sudden stress often triggers diverse, temporally structured gene expression responses in microbes, but it is largely unknown how variable in time such responses are and if genes respond in the same temporal order in every single cell. Here, we quantified timing variability of individual promoters responding to sublethal antibiotic stress using fluorescent reporters, microfluidics, and time‐lapse microscopy. We identified lower and upper bounds that put definite constraints on timing variability, which varies strongly among promoters and conditions. Timing variability can be interpreted using results from statistical kinetics, which enable us to estimate the number of rate‐limiting molecular steps underlying different responses. We found that just a few critical steps control some responses while others rely on dozens of steps. To probe connections between different stress responses, we then tracked the temporal order and response time correlations of promoter pairs in individual cells. Our results support that, when bacteria are exposed to the antibiotic nitrofurantoin, the ensuing oxidative stress and SOS responses are part of the same causal chain of molecular events. In contrast, under trimethoprim, the acid stress response and the SOS response are part of different chains of events running in parallel. Our approach reveals fundamental constraints on gene expression timing and provides new insights into the molecular events that underlie the timing of stress responses.}, author = {Mitosch, Karin and Rieckh, Georg and Bollenbach, Mark Tobias}, journal = {Molecular systems biology}, number = {2}, publisher = {Embo Press}, title = {{Temporal order and precision of complex stress responses in individual bacteria}}, doi = {10.15252/msb.20188470}, volume = {15}, year = {2019}, } @article{7026, abstract = {Effective design of combination therapies requires understanding the changes in cell physiology that result from drug interactions. Here, we show that the genome-wide transcriptional response to combinations of two drugs, measured at a rigorously controlled growth rate, can predict higher-order antagonism with a third drug in Saccharomyces cerevisiae. Using isogrowth profiling, over 90% of the variation in cellular response can be decomposed into three principal components (PCs) that have clear biological interpretations. We demonstrate that the third PC captures emergent transcriptional programs that are dependent on both drugs and can predict antagonism with a third drug targeting the emergent pathway. We further show that emergent gene expression patterns are most pronounced at a drug ratio where the drug interaction is strongest, providing a guideline for future measurements. Our results provide a readily applicable recipe for uncovering emergent responses in other systems and for higher-order drug combinations. A record of this paper’s transparent peer review process is included in the Supplemental Information.}, author = {Lukacisin, Martin and Bollenbach, Tobias}, issn = {2405-4712}, journal = {Cell Systems}, number = {5}, pages = {423--433.e1--e3}, publisher = {Cell Press}, title = {{Emergent gene expression responses to drug combinations predict higher-order drug interactions}}, doi = {10.1016/j.cels.2019.10.004}, volume = {9}, year = {2019}, } @article{674, abstract = {Navigation of cells along gradients of guidance cues is a determining step in many developmental and immunological processes. Gradients can either be soluble or immobilized to tissues as demonstrated for the haptotactic migration of dendritic cells (DCs) toward higher concentrations of immobilized chemokine CCL21. To elucidate how gradient characteristics govern cellular response patterns, we here introduce an in vitro system allowing to track migratory responses of DCs to precisely controlled immobilized gradients of CCL21. We find that haptotactic sensing depends on the absolute CCL21 concentration and local steepness of the gradient, consistent with a scenario where DC directionality is governed by the signal-to-noise ratio of CCL21 binding to the receptor CCR7. We find that the conditions for optimal DC guidance are perfectly provided by the CCL21 gradients we measure in vivo. Furthermore, we find that CCR7 signal termination by the G-protein-coupled receptor kinase 6 (GRK6) is crucial for haptotactic but dispensable for chemotactic CCL21 gradient sensing in vitro and confirm those observations in vivo. These findings suggest that stable, tissue-bound CCL21 gradients as sustainable “roads” ensure optimal guidance in vivo.}, author = {Schwarz, Jan and Bierbaum, Veronika and Vaahtomeri, Kari and Hauschild, Robert and Brown, Markus and De Vries, Ingrid and Leithner, Alexander F and Reversat, Anne and Merrin, Jack and Tarrant, Teresa and Bollenbach, Tobias and Sixt, Michael K}, issn = {09609822}, journal = {Current Biology}, number = {9}, pages = {1314 -- 1325}, publisher = {Cell Press}, title = {{Dendritic cells interpret haptotactic chemokine gradients in a manner governed by signal to noise ratio and dependent on GRK6}}, doi = {10.1016/j.cub.2017.04.004}, volume = {27}, year = {2017}, } @article{666, abstract = {Antibiotics elicit drastic changes in microbial gene expression, including the induction of stress response genes. While certain stress responses are known to “cross-protect” bacteria from other stressors, it is unclear whether cellular responses to antibiotics have a similar protective role. By measuring the genome-wide transcriptional response dynamics of Escherichia coli to four antibiotics, we found that trimethoprim induces a rapid acid stress response that protects bacteria from subsequent exposure to acid. Combining microfluidics with time-lapse imaging to monitor survival and acid stress response in single cells revealed that the noisy expression of the acid resistance operon gadBC correlates with single-cell survival. Cells with higher gadBC expression following trimethoprim maintain higher intracellular pH and survive the acid stress longer. The seemingly random single-cell survival under acid stress can therefore be predicted from gadBC expression and rationalized in terms of GadB/C molecular function. Overall, we provide a roadmap for identifying the molecular mechanisms of single-cell cross-protection between antibiotics and other stressors.}, author = {Mitosch, Karin and Rieckh, Georg and Bollenbach, Tobias}, issn = {24054712}, journal = {Cell Systems}, number = {4}, pages = {393 -- 403}, publisher = {Cell Press}, title = {{Noisy response to antibiotic stress predicts subsequent single cell survival in an acidic environment}}, doi = {10.1016/j.cels.2017.03.001}, volume = {4}, year = {2017}, } @article{822, abstract = {Polymicrobial infections constitute small ecosystems that accommodate several bacterial species. Commonly, these bacteria are investigated in isolation. However, it is unknown to what extent the isolates interact and whether their interactions alter bacterial growth and ecosystem resilience in the presence and absence of antibiotics. We quantified the complete ecological interaction network for 72 bacterial isolates collected from 23 individuals diagnosed with polymicrobial urinary tract infections and found that most interactions cluster based on evolutionary relatedness. Statistical network analysis revealed that competitive and cooperative reciprocal interactions are enriched in the global network, while cooperative interactions are depleted in the individual host community networks. A population dynamics model parameterized by our measurements suggests that interactions restrict community stability, explaining the observed species diversity of these communities. We further show that the clinical isolates frequently protect each other from clinically relevant antibiotics. Together, these results highlight that ecological interactions are crucial for the growth and survival of bacteria in polymicrobial infection communities and affect their assembly and resilience. }, author = {De Vos, Marjon and Zagórski, Marcin P and Mcnally, Alan and Bollenbach, Mark Tobias}, issn = {00278424}, journal = {PNAS}, number = {40}, pages = {10666 -- 10671}, publisher = {National Academy of Sciences}, title = {{Interaction networks, ecological stability, and collective antibiotic tolerance in polymicrobial infections}}, doi = {10.1073/pnas.1713372114}, volume = {114}, year = {2017}, } @article{1027, abstract = {The rising prevalence of antibiotic resistant bacteria is an increasingly serious public health challenge. To address this problem, recent work ranging from clinical studies to theoretical modeling has provided valuable insights into the mechanisms of resistance, its emergence and spread, and ways to counteract it. A deeper understanding of the underlying dynamics of resistance evolution will require a combination of experimental and theoretical expertise from different disciplines and new technology for studying evolution in the laboratory. Here, we review recent advances in the quantitative understanding of the mechanisms and evolution of antibiotic resistance. We focus on key theoretical concepts and new technology that enables well-controlled experiments. We further highlight key challenges that can be met in the near future to ultimately develop effective strategies for combating resistance.}, author = {Lukacisinova, Marta and Bollenbach, Mark Tobias}, journal = {Current Opinion in Biotechnology}, pages = {90 -- 97}, publisher = {Elsevier}, title = {{Toward a quantitative understanding of antibiotic resistance evolution}}, doi = {10.1016/j.copbio.2017.02.013}, volume = {46}, year = {2017}, } @article{1154, abstract = {Cellular locomotion is a central hallmark of eukaryotic life. It is governed by cell-extrinsic molecular factors, which can either emerge in the soluble phase or as immobilized, often adhesive ligands. To encode for direction, every cue must be present as a spatial or temporal gradient. Here, we developed a microfluidic chamber that allows measurement of cell migration in combined response to surface immobilized and soluble molecular gradients. As a proof of principle we study the response of dendritic cells to their major guidance cues, chemokines. The majority of data on chemokine gradient sensing is based on in vitro studies employing soluble gradients. Despite evidence suggesting that in vivo chemokines are often immobilized to sugar residues, limited information is available how cells respond to immobilized chemokines. We tracked migration of dendritic cells towards immobilized gradients of the chemokine CCL21 and varying superimposed soluble gradients of CCL19. Differential migratory patterns illustrate the potential of our setup to quantitatively study the competitive response to both types of gradients. Beyond chemokines our approach is broadly applicable to alternative systems of chemo- and haptotaxis such as cells migrating along gradients of adhesion receptor ligands vs. any soluble cue. }, author = {Schwarz, Jan and Bierbaum, Veronika and Merrin, Jack and Frank, Tino and Hauschild, Robert and Bollenbach, Mark Tobias and Tay, Savaş and Sixt, Michael K and Mehling, Matthias}, journal = {Scientific Reports}, publisher = {Nature Publishing Group}, title = {{A microfluidic device for measuring cell migration towards substrate bound and soluble chemokine gradients}}, doi = {10.1038/srep36440}, volume = {6}, year = {2016}, } @article{1581, abstract = {In animal embryos, morphogen gradients determine tissue patterning and morphogenesis. Shyer et al. provide evidence that, during vertebrate gut formation, tissue folding generates graded activity of signals required for subsequent steps of gut growth and differentiation, thereby revealing an intriguing link between tissue morphogenesis and morphogen gradient formation.}, author = {Bollenbach, Mark Tobias and Heisenberg, Carl-Philipp J}, journal = {Cell}, number = {3}, pages = {431 -- 432}, publisher = {Cell Press}, title = {{Gradients are shaping up}}, doi = {10.1016/j.cell.2015.04.009}, volume = {161}, year = {2015}, } @article{1810, abstract = {Combining antibiotics is a promising strategy for increasing treatment efficacy and for controlling resistance evolution. When drugs are combined, their effects on cells may be amplified or weakened, that is the drugs may show synergistic or antagonistic interactions. Recent work revealed the underlying mechanisms of such drug interactions by elucidating the drugs'; joint effects on cell physiology. Moreover, new treatment strategies that use drug combinations to exploit evolutionary tradeoffs were shown to affect the rate of resistance evolution in predictable ways. High throughput studies have further identified drug candidates based on their interactions with established antibiotics and general principles that enable the prediction of drug interactions were suggested. Overall, the conceptual and technical foundation for the rational design of potent drug combinations is rapidly developing.}, author = {Bollenbach, Mark Tobias}, journal = {Current Opinion in Microbiology}, pages = {1 -- 9}, publisher = {Elsevier}, title = {{Antimicrobial interactions: Mechanisms and implications for drug discovery and resistance evolution}}, doi = {10.1016/j.mib.2015.05.008}, volume = {27}, year = {2015}, } @article{1823, abstract = {Abstract Drug combinations are increasingly important in disease treatments, for combating drug resistance, and for elucidating fundamental relationships in cell physiology. When drugs are combined, their individual effects on cells may be amplified or weakened. Such drug interactions are crucial for treatment efficacy, but their underlying mechanisms remain largely unknown. To uncover the causes of drug interactions, we developed a systematic approach based on precise quantification of the individual and joint effects of antibiotics on growth of genome-wide Escherichia coli gene deletion strains. We found that drug interactions between antibiotics representing the main modes of action are highly robust to genetic perturbation. This robustness is encapsulated in a general principle of bacterial growth, which enables the quantitative prediction of mutant growth rates under drug combinations. Rare violations of this principle exposed recurring cellular functions controlling drug interactions. In particular, we found that polysaccharide and ATP synthesis control multiple drug interactions with previously unexplained mechanisms, and small molecule adjuvants targeting these functions synthetically reshape drug interactions in predictable ways. These results provide a new conceptual framework for the design of multidrug combinations and suggest that there are universal mechanisms at the heart of most drug interactions. Synopsis A general principle of bacterial growth enables the prediction of mutant growth rates under drug combinations. Rare violations of this principle expose cellular functions that control drug interactions and can be targeted by small molecules to alter drug interactions in predictable ways. Drug interactions between antibiotics are highly robust to genetic perturbations. A general principle of bacterial growth enables the prediction of mutant growth rates under drug combinations. Rare violations of this principle expose cellular functions that control drug interactions. Diverse drug interactions are controlled by recurring cellular functions, including LPS synthesis and ATP synthesis. A general principle of bacterial growth enables the prediction of mutant growth rates under drug combinations. Rare violations of this principle expose cellular functions that control drug interactions and can be targeted by small molecules to alter drug interactions in predictable ways.}, author = {Chevereau, Guillaume and Bollenbach, Mark Tobias}, journal = {Molecular Systems Biology}, number = {4}, publisher = {Nature Publishing Group}, title = {{Systematic discovery of drug interaction mechanisms}}, doi = {10.15252/msb.20156098}, volume = {11}, year = {2015}, } @misc{9711, author = {Chevereau, Guillaume and Lukacisinova, Marta and Batur, Tugce and Guvenek, Aysegul and Ayhan, Dilay Hazal and Toprak, Erdal and Bollenbach, Mark Tobias}, publisher = {Public Library of Science}, title = {{Excel file containing the raw data for all figures}}, doi = {10.1371/journal.pbio.1002299.s001}, year = {2015}, } @misc{9765, author = {Chevereau, Guillaume and Lukacisinova, Marta and Batur, Tugce and Guvenek, Aysegul and Ayhan, Dilay Hazal and Toprak, Erdal and Bollenbach, Mark Tobias}, publisher = {Public Library of Science}, title = {{Gene ontology enrichment analysis for the most sensitive gene deletion strains for all drugs}}, doi = {10.1371/journal.pbio.1002299.s008}, year = {2015}, } @article{1509, abstract = {The Auxin Binding Protein1 (ABP1) has been identified based on its ability to bind auxin with high affinity and studied for a long time as a prime candidate for the extracellular auxin receptor responsible for mediating in particular the fast non-transcriptional auxin responses. However, the contradiction between the embryo-lethal phenotypes of the originally described Arabidopsis T-DNA insertional knock-out alleles (abp1-1 and abp1-1s) and the wild type-like phenotypes of other recently described loss-of-function alleles (abp1-c1 and abp1-TD1) questions the biological importance of ABP1 and relevance of the previous genetic studies. Here we show that there is no hidden copy of the ABP1 gene in the Arabidopsis genome but the embryo-lethal phenotypes of abp1-1 and abp1-1s alleles are very similar to the knock-out phenotypes of the neighboring gene, BELAYA SMERT (BSM). Furthermore, the allelic complementation test between bsm and abp1 alleles shows that the embryo-lethality in the abp1-1 and abp1-1s alleles is caused by the off-target disruption of the BSM locus by the T-DNA insertions. This clarifies the controversy of different phenotypes among published abp1 knock-out alleles and asks for reflections on the developmental role of ABP1.}, author = {Michalko, Jaroslav and Dravecka, Marta and Bollenbach, Tobias and Friml, Jirí}, journal = {F1000 Research }, publisher = {F1000 Research}, title = {{Embryo-lethal phenotypes in early abp1 mutants are due to disruption of the neighboring BSM gene}}, doi = {10.12688/f1000research.7143.1}, volume = {4}, year = {2015}, } @article{1619, abstract = {The emergence of drug resistant pathogens is a serious public health problem. It is a long-standing goal to predict rates of resistance evolution and design optimal treatment strategies accordingly. To this end, it is crucial to reveal the underlying causes of drug-specific differences in the evolutionary dynamics leading to resistance. However, it remains largely unknown why the rates of resistance evolution via spontaneous mutations and the diversity of mutational paths vary substantially between drugs. Here we comprehensively quantify the distribution of fitness effects (DFE) of mutations, a key determinant of evolutionary dynamics, in the presence of eight antibiotics representing the main modes of action. Using precise high-throughput fitness measurements for genome-wide Escherichia coli gene deletion strains, we find that the width of the DFE varies dramatically between antibiotics and, contrary to conventional wisdom, for some drugs the DFE width is lower than in the absence of stress. We show that this previously underappreciated divergence in DFE width among antibiotics is largely caused by their distinct drug-specific dose-response characteristics. Unlike the DFE, the magnitude of the changes in tolerated drug concentration resulting from genome-wide mutations is similar for most drugs but exceptionally small for the antibiotic nitrofurantoin, i.e., mutations generally have considerably smaller resistance effects for nitrofurantoin than for other drugs. A population genetics model predicts that resistance evolution for drugs with this property is severely limited and confined to reproducible mutational paths. We tested this prediction in laboratory evolution experiments using the “morbidostat”, a device for evolving bacteria in well-controlled drug environments. Nitrofurantoin resistance indeed evolved extremely slowly via reproducible mutations—an almost paradoxical behavior since this drug causes DNA damage and increases the mutation rate. Overall, we identified novel quantitative characteristics of the evolutionary landscape that provide the conceptual foundation for predicting the dynamics of drug resistance evolution.}, author = {Chevereau, Guillaume and Dravecka, Marta and Batur, Tugce and Guvenek, Aysegul and Ayhan, Dilay and Toprak, Erdal and Bollenbach, Mark Tobias}, journal = {PLoS Biology}, number = {11}, publisher = {Public Library of Science}, title = {{Quantifying the determinants of evolutionary dynamics leading to drug resistance}}, doi = {10.1371/journal.pbio.1002299}, volume = {13}, year = {2015}, } @article{2040, abstract = {Development requires tissue growth as well as cell diversification. To address how these processes are coordinated, we analyzed the development of molecularly distinct domains of neural progenitors in the mouse and chick neural tube. We show that during development, these domains undergo changes in size that do not scale with changes in overall tissue size. Our data show that domain proportions are first established by opposing morphogen gradients and subsequently controlled by domain-specific regulation of differentiation rate but not differences in proliferation rate. Regulation of differentiation rate is key to maintaining domain proportions while accommodating both intra- and interspecies variations in size. Thus, the sequential control of progenitor specification and differentiation elaborates pattern without requiring that signaling gradients grow as tissues expand. }, author = {Kicheva, Anna and Bollenbach, Mark Tobias and Ribeiro, Ana and Pérez Valle, Helena and Lovell Badge, Robin and Episkopou, Vasso and Briscoe, James}, journal = {Science}, number = {6204}, publisher = {American Association for the Advancement of Science}, title = {{Coordination of progenitor specification and growth in mouse and chick spinal cord}}, doi = {10.1126/science.1254927}, volume = {345}, year = {2014}, } @article{2220, abstract = {In this issue of Chemistry & Biology, Cokol and colleagues report a systematic study of drug interactions between antifungal compounds. Suppressive drug interactions occur more frequently than previously realized and come in different flavors with interesting implications.}, author = {De Vos, Marjon and Bollenbach, Mark Tobias}, issn = {10745521}, journal = {Chemistry and Biology}, number = {4}, pages = {439 -- 440}, publisher = {Cell Press}, title = {{Suppressive drug interactions between antifungals}}, doi = {10.1016/j.chembiol.2014.04.004}, volume = {21}, year = {2014}, } @article{2001, abstract = {Antibiotics affect bacterial cell physiology at many levels. Rather than just compensating for the direct cellular defects caused by the drug, bacteria respond to antibiotics by changing their morphology, macromolecular composition, metabolism, gene expression and possibly even their mutation rate. Inevitably, these processes affect each other, resulting in a complex response with changes in the expression of numerous genes. Genome‐wide approaches can thus help in gaining a comprehensive understanding of bacterial responses to antibiotics. In addition, a combination of experimental and theoretical approaches is needed for identifying general principles that underlie these responses. Here, we review recent progress in our understanding of bacterial responses to antibiotics and their combinations, focusing on effects at the levels of growth rate and gene expression. We concentrate on studies performed in controlled laboratory conditions, which combine promising experimental techniques with quantitative data analysis and mathematical modeling. While these basic research approaches are not immediately applicable in the clinic, uncovering the principles and mechanisms underlying bacterial responses to antibiotics may, in the long term, contribute to the development of new treatment strategies to cope with and prevent the rise of resistant pathogenic bacteria.}, author = {Mitosch, Karin and Bollenbach, Tobias}, journal = {Environmental Microbiology Reports}, number = {6}, pages = {545 -- 557}, publisher = {Wiley}, title = {{Bacterial responses to antibiotics and their combinations}}, doi = {10.1111/1758-2229.12190}, volume = {6}, year = {2014}, } @article{2839, abstract = {Directional guidance of cells via gradients of chemokines is considered crucial for embryonic development, cancer dissemination, and immune responses. Nevertheless, the concept still lacks direct experimental confirmation in vivo. Here, we identify endogenous gradients of the chemokine CCL21 within mouse skin and show that they guide dendritic cells toward lymphatic vessels. Quantitative imaging reveals depots of CCL21 within lymphatic endothelial cells and steeply decaying gradients within the perilymphatic interstitium. These gradients match the migratory patterns of the dendritic cells, which directionally approach vessels from a distance of up to 90-micrometers. Interstitial CCL21 is immobilized to heparan sulfates, and its experimental delocalization or swamping the endogenous gradients abolishes directed migration. These findings functionally establish the concept of haptotaxis, directed migration along immobilized gradients, in tissues.}, author = {Weber, Michele and Hauschild, Robert and Schwarz, Jan and Moussion, Christine and De Vries, Ingrid and Legler, Daniel and Luther, Sanjiv and Bollenbach, Mark Tobias and Sixt, Michael K}, journal = {Science}, number = {6117}, pages = {328 -- 332}, publisher = {American Association for the Advancement of Science}, title = {{Interstitial dendritic cell guidance by haptotactic chemokine gradients}}, doi = {10.1126/science.1228456}, volume = {339}, year = {2013}, } @article{2970, abstract = {Morphogen gradients regulate the patterning and growth of many tissues, hence a key question is how they are established and maintained during development. Theoretical descriptions have helped to explain how gradient shape is controlled by the rates of morphogen production, spreading and degradation. These effective rates have been measured using fluorescence recovery after photobleaching (FRAP) and photoactivation. To unravel which molecular events determine the effective rates, such tissue-level assays have been combined with genetic analysis, high-resolution assays, and models that take into account interactions with receptors, extracellular components and trafficking. Nevertheless, because of the natural and experimental data variability, and the underlying assumptions of transport models, it remains challenging to conclusively distinguish between cellular mechanisms.}, author = {Kicheva, Anna and Bollenbach, Mark Tobias and Wartlick, Ortrud and Julicher, Frank and Gonzalez Gaitan, Marcos}, journal = {Current Opinion in Genetics & Development}, number = {6}, pages = {527 -- 532}, publisher = {Elsevier}, title = {{Investigating the principles of morphogen gradient formation: from tissues to cells}}, doi = {10.1016/j.gde.2012.08.004}, volume = {22}, year = {2012}, } @article{3160, abstract = {There is a long-running controversy about how early cell fate decisions are made in the developing mammalian embryo. 1,2 In particular, it is controversial when the first events that can predict the establishment of the pluripotent and extra-embryonic lineages in the blastocyst of the pre-implantation embryo occur. It has long been proposed that the position and polarity of cells at the 16- to 32-cell stage embryo influence their decision to either give rise to the pluripotent cell lineage that eventually contributes to the inner cell mass (ICM), comprising the primitive endoderm (PE) and the epiblast (EPI), or the extra-embryonic trophectoderm (TE) surrounding the blastocoel. The positioning of cells in the embryo at this developmental stage could largely be the result of random events, making this a stochastic model of cell lineage allocation. Contrary to such a stochastic model, some studies have detected putative differences in the lineage potential of individual blastomeres before compaction, indicating that the first cell fate decisions may occur as early as at the 4-cell stage. Using a non-invasive, quantitative in vivo imaging assay to study the kinetic behavior of Oct4 (also known as POU5F1), a key transcription factor (TF) controlling pre-implantation development in the mouse embryo, 3-5 a recent study identifies Oct4 kinetics as a predictive measure of cell lineage patterning in the early mouse embryo. 6 Here, we discuss the implications of such molecular heterogeneities in early development and offer potential avenues toward a mechanistic understanding of these observations, contributing to the resolution of the controversy of developmental cell lineage allocation.}, author = {Pantazis, Periklis and Bollenbach, Tobias}, journal = {Cell Cycle}, number = {11}, pages = {2055 -- 2058}, publisher = {Taylor and Francis}, title = {{Transcription factor kinetics and the emerging asymmetry in the early mammalian embryo}}, doi = {10.4161/cc.20118}, volume = {11}, year = {2012}, } @article{3429, abstract = {Transcription factors are central to sustaining pluripotency, yet little is known about transcription factor dynamics in defining pluripotency in the early mammalian embryo. Here, we establish a fluorescence decay after photoactivation (FDAP) assay to quantitatively study the kinetic behaviour of Oct4, a key transcription factor controlling pre-implantation development in the mouse embryo. FDAP measurements reveal that each cell in a developing embryo shows one of two distinct Oct4 kinetics, before there are any morphologically distinguishable differences or outward signs of lineage patterning. The differences revealed by FDAP are due to differences in the accessibility of Oct4 to its DNA binding sites in the nucleus. Lineage tracing of the cells in the two distinct sub-populations demonstrates that the Oct4 kinetics predict lineages of the early embryo. Cells with slower Oct4 kinetics are more likely to give rise to the pluripotent cell lineage that contributes to the inner cell mass. Those with faster Oct4 kinetics contribute mostly to the extra-embryonic lineage. Our findings identify Oct4 kinetics, rather than differences in total transcription factor expression levels, as a predictive measure of developmental cell lineage patterning in the early mouse embryo.}, author = {Plachta, Nicolas and Bollenbach, Mark Tobias and Pease, Shirley and Fraser, Scott and Pantazis, Periklis}, journal = {Nature Cell Biology}, number = {2}, pages = {117 -- 123}, publisher = {Nature Publishing Group}, title = {{Oct4 kinetics predict cell lineage patterning in the early mammalian embryo}}, doi = {10.1038/ncb2154}, volume = {13}, year = {2011}, } @article{3376, abstract = {Regulatory conflicts occur when two signals that individually trigger opposite cellular responses are present simultaneously. Here, we investigate regulatory conflicts in the bacterial response to antibiotic combinations. We use an Escherichia coli promoter-GFP library to study the transcriptional response of many promoters to either additive or antagonistic drug pairs at fine two-dimensional (2D) resolution of drug concentration. Surprisingly, we find that this data set can be characterized as a linear sum of only two principal components. Component one, accounting for over 70% of the response, represents the response to growth inhibition by the drugs. Component two describes how regulatory conflicts are resolved. For the additive drug pair, conflicts are resolved by linearly interpolating the single drug responses, while for the antagonistic drug pair, the growth-limiting drug dominates the response. Importantly, for a given drug pair, the same conflict resolution strategy applies to almost all genes. These results provide a recipe for predicting gene expression responses to antibiotic combinations.}, author = {Bollenbach, Mark Tobias and Kishony, Roy}, journal = {Molecular Cell}, number = {4}, pages = {413 -- 425}, publisher = {Cell Press}, title = {{Resolution of gene regulatory conflicts caused by combinations of antibiotics}}, doi = {10.1016/j.molcel.2011.04.016}, volume = {42}, year = {2011}, } @article{3428, abstract = {In this issue of Molecular Cell, Davies et al. (2009) work out a sequence of active cellular events that lead to the death of Escherichia coli in the presence of the drug hydroxyurea.}, author = {Bollenbach, Mark Tobias and Kishony, Roy}, journal = {Molecular Cell}, number = {5}, pages = {728 -- 729}, publisher = {Cell Press}, title = {{Hydroxyurea triggers cellular responses that actively cause bacterial cell death}}, doi = {10.1016/j.molcel.2009.11.027}, volume = {36}, year = {2009}, } @article{4223, abstract = {Both Gram-positive and Gram-negative bacteria contain bactoprenol-dependent biosynthetic pathways expressing non-essential cell surface polysaccharides that function as virulence factors. Although these polymers are not required for bacterial viability in vitro, genes in many of the biosynthetic pathways are conditionally essential: they cannot be deleted except in strains incapable of initiating polymer synthesis. We report a cell-based, pathway-specific strategy to screen for small molecule inhibitors of conditionally essential enzymes. The screen identifies molecules that prevent the growth of a wildtype bacterial strain but do not affect the growth of a mutant strain incapable of initiating polymer synthesis. We have applied this approach to discover inhibitors of wall teichoic acid (WTA) biosynthesis in Staphylococcus aureus. WTAs are anionic cell surface polysaccharides required for host colonization that have been suggested as targets for new antimicrobials. We have identified a small molecule, 7-chloro-N,N-diethyl-3-(phenylsulfonyl)-[1,2,3]triazolo[1,5-a]quinolin-5-amine (1835F03), that inhibits the growth of a panel of S. aureus strains (MIC = 1−3 μg mL−1), including clinical methicillin-resistant S. aureus (MRSA) isolates. Using a combination of biochemistry and genetics, we have identified the molecular target as TarG, the transmembrane component of the ABC transporter that exports WTAs to the cell surface. We also show that preventing the completion of WTA biosynthesis once it has been initiated triggers growth arrest. The discovery of 1835F03 validates our chemical genetics strategy for identifying inhibitors of conditionally essential enzymes, and the strategy should be applicable to many other bactoprenol-dependent biosynthetic pathways in the pursuit of novel antibacterials and probes of bacterial stress responses.}, author = {Swoboda, Jonathan and Meredith, Timothy and Campbell, Jennifer and Brown, Stephanie and Suzuki, Takashi and Bollenbach, Mark Tobias and Malhowski, Amy and Kishony, Roy and Gilmore, Michael and Walker, Suzanne}, journal = {ACS Chemical Biology}, number = {10}, pages = {875 -- 883}, publisher = {American Chemical Society}, title = {{Discovery of a Small Molecule that Blocks Wall Teichoic Acid Biosynthesis in Staphylococcus aureus}}, doi = {10.1021/cb900151k}, volume = {4}, year = {2009}, } @article{3398, abstract = {Why is a particular architecture for a pathway chosen over seemingly equivalent alternatives? Çağatay et al. (2009) use a synthetic biology approach to show that fluctuations—or noise—in protein levels may play a key role in determining which network design is selected during evolution.}, author = {Bollenbach, Tobias and Kishony, Roy}, journal = {Cell}, number = {3}, pages = {460 -- 461}, publisher = {Cell Press}, title = {{Quiet gene circuit more fragile than its noisy peer}}, doi = {10.1016/j.cell.2009.10.005}, volume = {139}, year = {2009}, } @article{3427, abstract = {We present a general theoretical framework to discuss mechanisms of morphogen transport and gradient formation in a cell layer. Trafficking events on the cellular scale lead to transport on larger scales. We discuss in particular the case of transcytosis where morphogens undergo repeated rounds of internalization into cells and recycling. Based on a description on the cellular scale, we derive effective nonlinear transport equations in one and two dimensions which are valid on larger scales. We derive analytic expressions for the concentration dependence of the effective diffusion coefficient and the effective degradation rate. We discuss the effects of a directional bias on morphogen transport and those of the coupling of the morphogen and receptor kinetics. Furthermore, we discuss general properties of cellular transport processes such as the robustness of gradients and relate our results to recent experiments on the morphogen Decapentaplegic (Dpp) that acts in the wing disk of the fruit fly Drosophila. © 2007 The American Physical Society}, author = {Bollenbach, Mark Tobias and Kruse, Karsten and Pantazis, Periklis and Gonzalez Gaitan, Marcos and Julicher, Frank}, journal = {Physical Review E Statistical Nonlinear and Soft Matter Physics}, number = {1}, publisher = {American Institute of Physics}, title = {{Morphogen transport in epithelia}}, doi = {10.1103/PhysRevE.75.011901}, volume = {75}, year = {2007}, } @article{4225, abstract = {The discovery of the genetic code was one of the most important advances of modern biology. But there is more to a DNA code than protein sequence; DNA carries signals for splicing, localization, folding, and regulation that are often embedded within the protein-coding sequence. In this issue, Itzkovitz and Alon show that the specific 64-to-20 mapping found in the genetic code may have been optimized for permitting protein-coding regions to carry this extra information and suggest that this property may have evolved as a side benefit of selection to minimize the negative effects of frameshift errors.}, author = {Bollenbach, Mark Tobias and Vetsigian, Kalin and Kishony, Roy}, journal = {Genome Research}, number = {4}, pages = {401 -- 404}, publisher = {Cold Spring Harbor Laboratory Press}, title = {{Evolution and multilevel optimization of the genetic code}}, doi = {10.1101/gr.6144007}, volume = {17}, year = {2007}, } @article{3426, abstract = {We discuss the formation of graded morphogen profiles in a cell layer by nonlinear transport phenomena, important for patterning developing organisms. We focus on a process termed transcytosis, where morphogen transport results from the binding of ligands to receptors on the cell surface, incorporation into the cell, and subsequent externalization. Starting from a microscopic model, we derive effective transport equations. We show that, in contrast to morphogen transport by extracellular diffusion, transcytosis leads to robust ligand profiles which are insensitive to the rate of ligand production.}, author = {Bollenbach, Mark Tobias and Kruse, Karsten and Pantazis, Periklis and González Gaitán, Marcos and Jülicher, Frank}, journal = {Physical Review Letters}, number = {1}, publisher = {American Physical Society}, title = {{Robust formation of morphogen gradients}}, doi = {10.1103/PhysRevLett.94.018103}, volume = {94}, year = {2005}, } @article{4224, abstract = {Developing cells acquire positional information by reading the graded distribution of morphogens. In Drosophila, the Dpp morphogen forms a long-range concentration gradient by spreading from a restricted source in the developing wing. It has been assumed that Dpp spreads by extracellular diffusion. Under this assumption, the main role of endocytosis in gradient formation is to downregulate receptors at the cell surface. These surface receptors bind to the ligand and thereby interfere with its long-range movement. Recent experiments indicate that Dpp spreading is mediated by Dynamin-dependent endocytosis in the target tissue, suggesting that extracellular diffusion alone cannot account for Dpp dispersal. Here, we perform a theoretical study of a model for morphogen spreading based on extracellular diffusion, which takes into account receptor binding and trafficking. We compare profiles of ligand and surface receptors obtained in this model with experimental data. To this end, we monitored directly the pool of surface receptors and extracellular Dpp with specific antibodies. We conclude that current models considering pure extracellular diffusion cannot explain the observed role of endocytosis during Dpp long-range movement.}, author = {Kruse, Karsten and Pantazis, Periklis and Bollenbach, Mark Tobias and Julicher, Frank and Gonzalez Gaitan, Marcos}, journal = {Development}, number = {19}, pages = {4843 -- 4856}, publisher = {Company of Biologists}, title = {{Dpp gradient formation by dynamin-dependent endocytosis: receptor trafficking and the diffusion model}}, doi = {10.1242/dev.01335}, volume = {131}, year = {2004}, } @inproceedings{3425, author = {Bollenbach, Mark Tobias and Strother, T. and Bauer, Wolfgang}, pages = {277 -- 288}, publisher = {Springer}, title = {{3D supernova collapse calculations}}, doi = {10.1007/978-1-4020-2705-5_21}, volume = {166}, year = {2003}, } @article{3797, author = {Bauer, Wolfgang and Kleine Berkenbusch, Marco and Bollenbach, Mark Tobias}, journal = {Revista Mexicana De Fisica}, number = {4}, pages = {1 -- 6}, publisher = {Sociedad Mexicana de Física}, title = {{Breaking atomic nuclei into little pieces: evidence for a phase transition}}, volume = {49}, year = {2003}, } @inproceedings{3424, abstract = {We give a brief overview of the current understanding of the explosion mechanism of core collapse supernovae. Our main focus is the impact of rotation on the explosion. Recent observations of the polarization of the light emitted by supernova explosions indicate that there are large deviations from spherical symmetry in the very heart of the explosion the origin of which is unknown. We use the new approach of a three dimensional test particle based simulation to simulate the infall phase of a supernova event. The underlying microphysics is simplified to make this computationally possible. A systematic study of the influence of rotation mainly during the infall phase of the collapse of a typical iron core is performed. Indications for significant deviations from spherical symmetry are found in our very rapidly rotating models. © 2002 American Institute of Physics }, author = {Bollenbach, Mark Tobias and Bauer, Wolfgang}, isbn = {9781510832008}, location = {Catania, Italy}, pages = {219 -- 232}, publisher = {American Institute of Physics}, title = {{3d supernovae collapse calculations}}, doi = {10.1063/1.1523196 }, volume = {644}, year = {2002}, } @inproceedings{3423, author = {Bauer, Wolfgang and Bollenbach, Mark Tobias and Kleine Berkenbusch, Marko and Harreis, Holger}, booktitle = {Proceedings of the 18th Winter Workshop on Nuclear Dynamics}, location = {Nassau, Bahamas}, pages = {111 -- 118}, publisher = {EP Systema}, title = {{The percolation interpretation of the nuclear fragmentation phase transition}}, year = {2002}, }