Jeffrey Abbott, Avik Mukherjee, Wenxuan Wu, Tianyang Ye, Han Sae Jung, Kevin M. Cheung, Rona S. Gertner, Markus Basan, Donhee Ham, and Hongkun Park. 3/10/2022. “Multi-parametric functional imaging of cell cultures and tissues with a CMOS microelectrode array.” Lab on a Chip. Publisher's VersionAbstract
Electrode-based impedance and electrochemical measurements can provide cell-biology information that is difficult to obtain using optical-microscopy techniques. Such electrical methods are non-invasive, label-free, and continuous, eliminating the need for fluorescence reporters and overcoming optical imaging's throughput/temporal resolution limitations. Nonetheless, electrode-based techniques have not been heavily employed because devices typically contain few electrodes per well, resulting in noisy aggregate readouts. Complementary metal-oxide-semiconductor (CMOS) microelectrode arrays (MEAs) have sometimes been used for electrophysiological measurements with thousands of electrodes per well at sub-cellular pitches, but only basic impedance mappings of cell attachment have been performed outside of electrophysiology. Here, we report on new field-based impedance mapping and electrochemical mapping/patterning techniques to expand CMOS-MEA cell-biology applications. The methods enable accurate measurement of cell attachment, growth/wound healing, cell–cell adhesion, metabolic state, and redox properties with single-cell spatial resolution (20 μm electrode pitch). These measurements allow the quantification of adhesion and metabolic differences of cells expressing oncogenes versus wild-type controls. The multi-parametric, cell-population statistics captured by the chip-scale integrated device opens up new avenues for fully electronic high-throughput live-cell assays for phenotypic screening and drug discovery applications.
Severin Josef Schink, Dimitris Christodoulou, Avik Mukherjee, Edward Athaide, Viktoria Brunner, Tobias Fuhrer, Gary Andrew Bradshaw, Uwe Sauer, and Markus Basan. 1/2022. “Glycolysis/gluconeogenesis specialization in microbes is driven by biochemical constraints of flux sensing.” Molecular systems biology, 18, 1, Pp. e10704. Publisher's VersionAbstract
Central carbon metabolism is highly conserved across microbial species, but can catalyze very different pathways depending on the organism and their ecological niche. Here, we study the dynamic reorganization of central metabolism after switches between the two major opposing pathway configurations of central carbon metabolism, glycolysis, and gluconeogenesis in Escherichia coliPseudomonas aeruginosa, and Pseudomonas putida. We combined growth dynamics and dynamic changes in intracellular metabolite levels with a coarse-grained model that integrates fluxes, regulation, protein synthesis, and growth and uncovered fundamental limitations of the regulatory network: After nutrient shifts, metabolite concentrations collapse to their equilibrium, rendering the cell unable to sense which direction the flux is supposed to flow through the metabolic network. The cell can partially alleviate this by picking a preferred direction of regulation at the expense of increasing lag times in the opposite direction. Moreover, decreasing both lag times simultaneously comes at the cost of reduced growth rate or higher futile cycling between metabolic enzymes. These three trade-offs can explain why microorganisms specialize for either glycolytic or gluconeogenic substrates and can help elucidate the complex growth patterns exhibited by different microbial species.
Severin Schink, Mark Polk, Edward Athaide, Avik Mukherjee, Constantin Ammar, Xili Liu, Seungeun Oh, Yu-Fang Chang, and Markus Basan. 11/22/2021. “The energy requirements of ion homeostasis determine the lifespan of starving bacteria.” bioRxiv, Pp. 11/22/2021. 11.22.469587. Publisher's VersionAbstract
The majority of microbes on earth, whether they live in the ocean, the soil or in animals, are not growing, but instead struggling to survive starvation16. Some genes and environmental conditions affecting starvation survival have been identified713, but despite almost a century of study1416, we do not know which processes lead to irreversible loss of viability, which maintenance processes counteract them and how lifespan is determined from the balance of these opposing processes. Here, we used time-lapse microscopy to capture and characterize the cell death process of E. coli during carbon starvation for the first time. We found that a lack of nutrients results in the collapse of ion homeostasis, triggering a positive-feedback cascade of osmotic swelling and membrane permeabilization that ultimately results in lysis. Based on these findings, we hypothesized that ion transport is the major energetic requirement for starving cells and the primary determinant of the timing of lysis. We therefore developed a mathematical model that integrates ion homeostasis and ‘cannibalistic’ nutrient recycling from perished cells16,17 to predict lifespan changes under diverse conditions, such as changes of cell size, medium composition, and prior growth conditions. Guided by model predictions, we found that cell death during starvation could be dramatically slowed by replacing inorganic ions from the medium with a non-permeating osmoprotectant, removing the cost of ion homeostasis and preventing lysis. Our quantitative and predictive model explains how survival kinetics are determined in starvation and elucidates the mechanistic underpinnings of starvation survival.
Seungeun Oh, Changhee Lee, Wenlong Yang, Ang Li, Avik Mukherjee, Markus Basan, Chongzhao Ran, Wei Yin, Clifford J. Tabin, Dan Fu, X. Sunney Xie, and Marc W. Kirschner. 9/15/2021. “Protein and Lipid Mass Concentration Measurement in Tissues by Stimulated Raman Scattering Microscopy.” bioRxiv 629543; doi: Publisher's Version
Severin Schink, Zara Gough, Elena Biselli, Mariel Garcia Huiman, Yu-Fang Chang, Markus Basan, and Ulrich Gerland. 8/4/2021. “MetA is a ‘thermal fuse’ that arrests growth and protects Escherichia coli at elevated temperatures.” bioRxiv 8/4/2021. 06.14.448417; doi: 06.14.448417. Publisher's Version
Markus Basan, Tomoya Honda, Dimitris Christodoulou, Manuel Hörl, Yu-Fang Chang, Emanuele Leoncini, Avik Mukherjee, Hiroyuki Okano, Brian R. Taylor, Josh M. Silverman, Carlos Sanchez, James R. Williamson, Johan Paulsson, Terence Hwa, and Uwe Sauer. 7/15/2020. “A universal trade-off between growth and lag in fluctuating environments.” Nature, 584, 7821, Pp. 470-474. Publisher's VersionAbstract
The rate of cell growth is crucial for bacterial fitness and drives the allocation of bacterial resources, affecting, for example, the expression levels of proteins dedicated to metabolism and biosynthesis. It is unclear, however, what ultimately determines growth rates in different environmental conditions. Moreover, increasing evidence suggests that other objectives are also important, such as the rate of physiological adaptation to changing environments. A common challenge for cells is that these objectives cannot be independently optimized, and maximizing one often reduces another. Many such trade-offs have indeed been hypothesized on the basis of qualitative correlative studies. Here we report a trade-off between steady-state growth rate and physiological adaptability in Escherichia coli, observed when a growing culture is abruptly shifted from a preferred carbon source such as glucose to fermentation products such as acetate. These metabolic transitions, common for enteric bacteria, are often accompanied by multi-hour lags before growth resumes. Metabolomic analysis reveals that long lags result from the depletion of key metabolites that follows the sudden reversal in the central carbon flux owing to the imposed nutrient shifts. A model of sequential flux limitation not only explains the observed trade-off between growth and adaptability, but also allows quantitative predictions regarding the universal occurrence of such tradeoffs, based on the opposing enzyme requirements of glycolysis versus gluconeogenesis. We validate these predictions experimentally for many different nutrient shifts in E. coli, as well as for other respiro-fermentative microorganisms, including Bacillus subtilis and Saccharomyces cerevisiae.
Markus Basan. 3/2018. “Resource allocation and metabolism: the search for governing principles.” Current Opinion in Microbiology, 45, Pp. 77-83. Publisher's VersionAbstract
Elucidating strategies of resource allocation and metabolism is crucial for a better understanding of microbial phenotypes. In particular, uncovering the governing principles underlying these processes would be a crucial step for achieving a central aim of systems microbiology, which is to quantitatively predict phenotypes of microbial cells or entire populations in diverse conditions. Here, some of the key concepts for understanding cellular resource allocation and metabolism that have been suggested over the past years are reviewed. In particular, recent experimental studies that have shown how phenotypic patterns from orthogonal genetic and environmental perturbations can help to differentiate between competing hypotheses and their respective predictions are discussed. Phenomenological models have proven to be a valuable addition to genome-scale models, capable of making quantitative predictions with only few parameters and having aided the identification of molecular mechanisms.
Markus Basan, Sheng Hui, and James R. Williamson. 9/19/2017. “ArcA overexpression induces fermentation and results in enhanced growth rates of E. coli.” Scientific Reports, 7, Pp. 11866. Publisher's VersionAbstract
Overflow metabolism in the presence of oxygen occurs at fast growth rates in a wide range of organisms including bacteria, yeast and cancer cells and plays an important role in biotechnology during production of proteins or metabolic compounds. As recently suggested, overflow metabolism can be understood in terms of proteome allocation, since fermentation has lower proteome cost for energy production than respiration. Here, we demonstrate that ArcA overexpression in aerobic conditions, results in downregulation of respiratory pathways and enhanced growth rates on glycolytic substrates of E. coli, coinciding with acetate excretion and increased carbon uptake rates. These results suggest that fermentation enables faster growth and demonstrate that fermentation on many glycolytic carbon sources is not limited by carbon uptake. Hence, these findings are difficult to reconcile with many alternative hypotheses that have been proposed for the origin of overflow metabolism and the growth rate dependence of fermentation and respiration, which are based on limited capacity of respiration or limitations in uptake rates and catabolic pathways. Instead, as suggested by increased lag phases of ArcA overexpression strains, respiratory energy metabolism may be related to a general preparatory response, observed for decreasing growth rates, but with limited advantages for maximizing steady-state growth rate.
Markus Basan, Sheng Hui, Hiroyuki Okano, Zhongge Zhang, Yang Shen, James R. Williamson, and Terence Hwa. 12/3/2015. “Overflow metabolism in Escherichia coli results from efficient proteome allocation.” Nature, 528, 7580, Pp. 99-104. Publisher's VersionAbstract

Overflow metabolism refers to the seemingly wasteful strategy in which cells use fermentation instead of the more efficient respiration to generate energy, despite the availability of oxygen. Known as the Warburg effect in the context of cancer growth, this phenomenon occurs ubiquitously for fast-growing cells, including bacteria, fungi and mammalian cells, but its origin has remained unclear despite decades of research. Here we study metabolic overflow in Escherichia coli, and show that it is a global physiological response used to cope with changing proteomic demands of energy biogenesis and biomass synthesis under different growth conditions. A simple model of proteomic resource allocation can quantitatively account for all of the observed behaviours, and accurately predict responses to new perturbations. The key hypothesis of the model, that the proteome cost of energy biogenesis by respiration exceeds that by fermentation, is quantitatively confirmed by direct measurement of protein abundances via quantitative mass spectrometry.

Markus Basan, Manlu Zhu, Xiongfeng Dai, Mya Warren, Daniel Sévin, Yi‐Ping Wang, and Terence Hwa. 10/30/2015. “Inflating bacterial cells by increased protein synthesis.” Molecular Systems Biology, 11, 10. Publisher's VersionAbstract

Understanding how the homeostasis of cellular size and composition is accomplished by different organisms is an outstanding challenge in biology. For exponentially growing Escherichia coli cells, it is long known that the size of cells exhibits a strong positive relation with their growth rates in different nutrient conditions. Here, we characterized cell sizes in a set of orthogonal growth limitations. We report that cell size and mass exhibit positive or negative dependences with growth rate depending on the growth limitation applied. In particular, synthesizing large amounts of “useless” proteins led to an inversion of the canonical, positive relation, with slow growing cells enlarged 7‐ to 8‐fold compared to cells growing at similar rates under nutrient limitation. Strikingly, this increase in cell size was accompanied by a 3‐ to 4‐fold increase in cellular DNA content at slow growth, reaching up to an amount equivalent to ~8 chromosomes per cell. Despite drastic changes in cell mass and macromolecular composition, cellular dry mass density remained constant. Our findings reveal an important role of protein synthesis in cell division control.

Sheng Hui, Josh M Silverman, Stephen S Chen, David W Erickson, Markus Basan, Jilong Wang, Terence Hwa, and James R Williamson. 2015. “Quantitative proteomic analysis reveals a simple strategy of global resource allocation in bacteria.” Molecular Systems Biology, 11, 2. Publisher's VersionAbstract

A central aim of cell biology was to understand the strategy of gene expression in response to the environment. Here, we study gene expression response to metabolic challenges in exponentially growing Escherichia coli using mass spectrometry. Despite enormous complexity in the details of the underlying regulatory network, we find that the proteome partitions into several coarse‐grained sectors, with each sector's total mass abundance exhibiting positive or negative linear relations with the growth rate. The growth rate‐dependent components of the proteome fractions comprise about half of the proteome by mass, and their mutual dependencies can be characterized by a simple flux model involving only two effective parameters. The success and apparent generality of this model arises from tight coordination between proteome partition and metabolism, suggesting a principle for resource allocation in proteome economy of the cell. This strategy of global gene regulation should serve as a basis for future studies on gene expression and constructing synthetic biological circuits. Coarse graining may be an effective approach to derive predictive phenomenological models for other ‘omics’ studies.

Juliane Zimmermann, Ryan L. Hayes, Markus Basan, José N. Onuchic, Wouter-Jan Rappel, and Herbert Levine. 8/5/2014. “Intercellular stress reconstitution from traction force data.” Biophysical Journal, 107, 3, Pp. 548–554. Publisher's VersionAbstract

Cells migrate collectively during development, wound healing, and cancer metastasis. Recently, a method has been developed to recover intercellular stress in monolayers from measured traction forces upon the substrate. To calculate stress maps in two dimensions, the cell sheet was assumed to behave like an elastic material, and it remains unclear to what extent this assumption is valid. In this study, we simulate our recently developed model for collective cell migration, and compute intercellular stress maps using the method employed in the experiments. We also compute these maps using a method that does not depend on the traction forces or material properties. The two independently obtained stress patterns agree well for the parameters we have probed and provide a verification of the validity of the experimental method.

J. Zimmermann, M. Basan, and H. Levine. 6/2014. “An instability at the edge of a tissue of collectively migrating cells can lead to finger formation during wound healing.” The European Physical Journal Special Topics, 223, 7, Pp. 1259–1264. Publisher's VersionAbstract

In wound healing assays, a monolayer of epithelial cells is allowed to migrate onto empty surface area. When the motile cells close the artificial wound, the edge of the tissue does usually not move uniformly but characteristic fingerlike protrusions are observed. We model the collectively moving cells as a system of self-propelled particles using the Toner-Tu equations for an active fluid. A linear stability analysis of perturbations at the tissue edge reveals an instability in the disordered nonmoving state. The instability is purely due to spontaneous motility and velocity alignment between cells. It can account for finger formation in wound healing experiments.

Thomas Risler and Markus Basan. 6/20/2013. “Morphological instabilities of stratified epithelia: a mechanical instability in tumour formation.” New Journal of Physics, 15. Publisher's VersionAbstract

Interfaces between stratified epithelia and their supporting stromas commonly exhibit irregular shapes. Undulations are particularly pronounced in dysplastic tissues and typically evolve into long, finger-like protrusions in carcinomas. In previous work (Basan et al 2011 Phys. Rev. Lett. 106 158101), we demonstrated that an instability arising from viscous shear stresses caused by the constant flow due to cell turnover in the epithelium could drive this phenomenon. While interfacial tension between the two tissues as well as mechanical resistance of the stroma tend to maintain a flat interface, an instability occurs for sufficiently large viscosity, cell-division rate and thickness of the dividing region in the epithelium. Here, extensions of this work are presented, where cell division in the epithelium is coupled to the local concentration of nutrients or growth factors diffusing from the stroma. This enhances the instability by a mechanism similar to that of the Mullins–Sekerka instability in single-diffusion processes of crystal growth. We furthermore present the instability for the generalized case of a viscoelastic stroma.

Markus Basan, Jens Elgeti, Edouard Hannezo, Wouter-Jan Rappel, and Herbert Levine. 2/12/2013. “Alignment of cellular motility forces with tissue flow as a mechanism for efficient wound healing.” Proceedings of the National Academy of Sciences USA, 110, 7, Pp. 2452–2459. Publisher's VersionAbstract

Recent experiments have shown that spreading epithelial sheets exhibit a long-range coordination of motility forces that leads to a buildup of tension in the tissue, which may enhance cell division and the speed of wound healing. Furthermore, the edges of these epithelial sheets commonly show finger-like protrusions whereas the bulk often displays spontaneous swirls of motile cells. To explain these experimental observations, we propose a simple flocking-type mechanism, in which cells tend to align their motility forces with their velocity. Implementing this idea in a mechanical tissue simulation, the proposed model gives rise to efficient spreading and can explain the experimentally observed long-range alignment of motility forces in highly disordered patterns, as well as the buildup of tensile stress throughout the tissue. Our model also qualitatively reproduces the dependence of swirl size and swirl velocity on cell density reported in experiments and exhibits an undulation instability at the edge of the spreading tissue commonly observed in vivo. Finally, we study the dependence of colony spreading speed on important physical and biological parameters and derive simple scaling relations that show that coordination of motility forces leads to an improvement of the wound healing process for realistic tissue parameters.

Fabien Montel, Morgan Delarue, Jens Elgeti, Laurent Malaquin, Markus Basan, Thomas Risler, Bernard Cabane, Danijela Vignjevic, Jacques Prost, Giovanni Cappello, and Jean-François Joanny. 10/28/2011. “Stress Clamp Experiments on Multicellular Tumor Spheroids.” Physical Review Letters , 107, 18. Publisher's VersionAbstract

The precise role of the microenvironment on tumor growth is poorly understood. Whereas the tumor is in constant competition with the surrounding tissue, little is known about the mechanics of this interaction. Using a novel experimental procedure, we study quantitatively the effect of an applied mechanical stress on the long-term growth of a spheroid cell aggregate. We observe that a stress of 10 kPa is sufficient to drastically reduce growth by inhibition of cell proliferation mainly in the core of the spheroid. We compare the results to a simple numerical model developed to describe the role of mechanics in cancer progression.

Markus Basan, Jean-François Joanny, Jacques Prost, and Thomas Risler. 4/11/2011. “Undulation instability of epithelial tissues.” Physical Review Letters , 106, 15. Publisher's VersionAbstract

Treating the epithelium as an incompressible fluid adjacent to a viscoelastic stroma, we find a novel hydrodynamic instability that leads to the formation of protrusions of the epithelium into the stroma. This instability is a candidate for epithelial fingering observed in vivo. It occurs for sufficiently large viscosity, cell-division rate and thickness of the dividing region in the epithelium. Our work provides physical insight into a potential mechanism by which interfaces between epithelia and stromas undulate and potentially by which tissue dysplasia leads to cancerous invasion.

Markus Basan, Jacques Prost, Jean-François Joanny, and Jens Elgeti. 4/1/2011. “Dissipative particle dynamics simulations for biological tissues: rheology and competition.” Physical Biology, 8, 2. Publisher's VersionAbstract

In this work, we model biological tissues using a simple, mechanistic simulation based on dissipative particle dynamics. We investigate the continuum behavior of the simulated tissue and determine its dependence on the properties of the individual cell. Cells in our simulation adhere to each other, expand in volume, divide after reaching a specific size checkpoint and undergo apoptosis at a constant rate, leading to a steady-state homeostatic pressure in the tissue. We measure the dependence of the homeostatic state on the microscopic parameters of our model and show that homeostatic pressure, rather than the unconfined rate of cell division, determines the outcome of tissue competitions. Simulated cell aggregates are cohesive and round up due to the effect of tissue surface tension, which we measure for different tissues. Furthermore, mixtures of different cells unmix according to their adhesive properties. Using a variety of shear and creep simulations, we study tissue rheology by measuring yield stresses, shear viscosities, complex viscosities as well as the loss tangents as a function of model parameters. We find that cell division and apoptosis lead to a vanishing yield stress and fluid-like tissues. The effects of different adhesion strengths and levels of noise on the rheology of the tissue are also measured. In addition, we find that the level of cell division and apoptosis drives the diffusion of cells in the tissue. Finally, we present a method for measuring the compressibility of the tissue and its response to external stress via cell division and apoptosis.

Jonas Ranft, Markus Basan, Jens Elgeti, Jean-François Joanny, Jacques Prost, and Frank Jülicher. 12/10/2010. “Fluidization of tissues by cell division and apoptosis.” Proceedings of the National Academy of Sciences USA, 107, 49, Pp. 20863–20868. Publisher's VersionAbstract

During the formation of tissues, cells organize collectively by cell division and apoptosis. The multicellular dynamics of such systems is influenced by mechanical conditions and can give rise to cell rearrangements and movements. We develop a continuum description of tissue dynamics, which describes the stress distribution and the cell flow field on large scales. In the absence of division and apoptosis, we consider the tissue to behave as an elastic solid. Cell division and apoptosis introduce stress sources that, in general, are anisotropic. By combining cell number balance with dynamic equations for the stress source, we show that the tissue effectively behaves as a viscoelastic fluid with a relaxation time set by the rates of division and apoptosis. If the system is confined in a fixed volume, it reaches a homeostatic state in which division and apoptosis balance. In this state, cells undergo a diffusive random motion driven by the stochasticity of division and apoptosis. We calculate the expression for the effective diffusion coefficient as a function of the tissue parameters and compare our results concerning both diffusion and viscosity to simulations of multicellular systems using dissipative particle dynamics.

Markus Basan, Timon Idema, Martin Lenz, Jean-François Joanny, and Thomas Risler. 6/16/2010. “A reaction-diffusion model of the cadherin-catenin system: a possible mechanism for contact inhibition and implications for tumorigenesis.” Biophysical Journal, 98, 12, Pp. 2770–2779. Publisher's VersionAbstract

Contact inhibition is the process by which cells switch from a motile growing state to a passive and stabilized state upon touching their neighbors. When two cells touch, an adhesion link is created between them by means of transmembrane E-cadherin proteins. Simultaneously, their actin filaments stop polymerizing in the direction perpendicular to the membrane and reorganize to create an apical belt that colocalizes with the adhesion links. Here, we propose a detailed quantitative model of the role of cytoplasmic β-catenin and α-catenin proteins in this process, treated as a reaction-diffusion system. Upon cell-cell contact the concentration in α-catenin dimers increases, inhibiting actin branching and thereby reducing cellular motility and expansion pressure. This model provides a mechanism for contact inhibition that could explain previously unrelated experimental findings on the role played by E-cadherin, β-catenin, and α-catenin in the cellular phenotype and in tumorigenesis. In particular, we address the effect of a knockout of the adenomatous polyposis coli tumor suppressor gene. Potential direct tests of our model are discussed.