Bulletin of the American Physical Society
APS March Meeting 2022
Volume 67, Number 3
Monday–Friday, March 14–18, 2022; Chicago
Session A04: Synthetic Biology IFocus Recordings Available
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Sponsoring Units: DBIO GSNP Chair: Gábor Balázsi, Stony Brook University Room: McCormick Place W-176C |
Monday, March 14, 2022 8:00AM - 8:12AM |
A04.00001: Molecular Implementations of Positive and Negative Feedback Inform Robustness in Biological Oscillator Motifs Chaitra Agrahar, Michael J Rust Biological oscillators like the cell cycle, the circadian sleep-wake cycle, etc. are vital to an organism's survival. Biochemical oscillator circuits are typically classified based on the net logic of the regulatory connections between interacting molecules. However, we show that the robustness response of biochemical oscillator motifs vary substantially based on the regulatory implementation of the logic of interactions encoded in the circuit topology. Nullcline analyses and linear stability arguments predict the robustness response of different mechanisms of a topology. Robust regulatory implementations not only enhance the probability of obtaining stable limit cycles over larger ranges of parameter variations, but also exhibit an increased resilience of oscillations to stochasticity. We further show that there are preferred regulatory implementations for particular biological functions, and that the most robust regulatory implementations of a topology are realized in naturally occurring oscillator systems where high phase coherence is desired. |
Monday, March 14, 2022 8:12AM - 8:24AM |
A04.00002: Metabolic Pathway Design Using Active Subspaces Andre Archer, Taylor Nichols, Niall Mangan, Danielle Tullman-Ercek Salmonella can utilize a biodiesel waste product, glycerol, to produce 1,3-PDO, a common commercial solvent. Experimental collaborators modify Salmonella to sequester the two enzymes needed for this pathway in microcompartments (MCPs), protein-bound shells that spatially segregate reactions. I formulated a differential equation model of this system to compare five candidate strains with different enzyme concentrations in the MCP. |
Monday, March 14, 2022 8:24AM - 9:00AM |
A04.00003: Building synthetic circuits to achieve cascading cell fate transitions Invited Speaker: Xiaojun Tian Previously, we proposed a cascading bistable switches (CBS) mechanism for epithelial to mesenchymal transition (EMT). Here we built a synthetic CBS (Syn-CBS) circuit in a single strain with two coupled self-activation modules to achieve two successive cell fate transitions. Interestingly, we find that the in vivo transition path was redirected as the activation of one switch always prevails against the other, contrary to the theoretically expected coactivation. This qualitatively different type of resource competition between the two modules follows a 'winner-takes-all' rule, where the winner is determined by the relative connection strength between the modules. To decouple the resource competition, we construct a two-strain circuit, which achieves successive activation and stable coactivation of the two switches. These results illustrate that a highly nonlinear hidden interaction between the circuit modules due to resource competition may cause counterintuitive consequences on circuit functions, which can be controlled with a division of labor strategy. |
Monday, March 14, 2022 9:00AM - 9:12AM |
A04.00004: Identifying Mechanisms of Gene Circuit Evolution that Elicit Mammalian Drug Resistance Joseph Cohen, Quanhua Mu, Ph.D, Yiming Wan, Kevin Farquar, Ph.D, Gabor Balazsi Stochasticity in gene expression is a prime determinant of drug resistance in mammalian cell populations, but the biological mechanisms that elicit such behaviors are poorly understood. Recently we used synthetic gene circuits harboring positive feedback (PF) and negative feedback (NF) regulation to tune mammalian drug resistance transgene (DRT) expression noise. Cell populations harboring PF circuitry exhibit high DRT expression noise, which favors the evolution of drug resistance in high stress environments. Cell populations harboring NF circuitry exhibit minimal DRT expression noise, which favors the evolution of drug resistance in low stress environments. Biological mechanisms that drive these observations are unknown. Here, we investigate transcriptional profiles of experimentally evolved NF and PF mammalian cell populations that became drug resistant. Identifying how mammalian populations combine and evolve drug-specific and pleiotropic drug resistance will provide insight into drug resistance mechanisms and may guide future applications of synthetic gene circuits in cell research and medicine. |
Monday, March 14, 2022 9:12AM - 9:24AM |
A04.00005: The Role of Energy Dissipation in Shaping Eukaryotic Transcriptional Regulation Zheng Diao, Caleb J Bashor, Oleg A Igoshin A mechanistic understanding of gene regulation is essential for quantitative control over gene expression. In prokaryotic systems, engineering rules were established via synergy between theory and experiments using model synthetic systems. However, a lack of interrelatable theoretical and experimental approaches have slowed progress in eukaryotes, where gene expression involves complex regulation driven by highly cooperative energy dissipative processes. To address this deficit, we developed a framework for modelling non-equilibrium regulation and used it to define underlying parameter regimes that set limits for cooperativity in various regulatory schemes, thereby discovering a trade-off between energy dissipation and cooperativity. To test our model, we developed an experimental system that enables sequence-level definition of a single-copy-integrated model fluorescent reporter locus in human cells. By expressing fluorescently labeled synthetic transcription factors—individually or in combination—that regulate the locus, we can precisely measure its gene regulatory functions. We envision using these theoretical and experimental approaches in tandem to establish quantitative rules for engineering complex gene networks in eukaryotes that incorporate non-equilibrium regulation. |
Monday, March 14, 2022 9:24AM - 10:00AM |
A04.00006: Advancing the frontiers of design-driven medicine: engineering programmable cell-based therapies and diagnostics Invited Speaker: Joshua N Leonard Engineered cell-based therapies are a transformative medical frontier. Employing living cells to perform sophisticated and complex tasks within the human body has already revolutionized the treatment of some cancers, and the prospect of extending these capabilities to promote health in myriad ways is now within reach. However, realizing the full potential of this approach will necessitate the development of improved engineering tools, ranging from improved biological technologies to computational and conceptual frameworks to guide their deployment. I will present recent advances made toward achieving the vision of design-driven engineering of novel mammalian cellular functions. This presentation will include recently developed "parts" for composing gene expression functions in mammalian cells, computational approaches for design-driven engineering of new parts such as biosensors, and computational tools and design principles that enable the efficient generation of novel functional programs that perform in a manner that is customizable, predictable, and robust. Each of these will be presented in the context of our primary motivation application—engineered cell-based devices for improving cancer treatment. |
Monday, March 14, 2022 10:00AM - 10:12AM |
A04.00007: Dynamics of DNA methylation and persistence of epigenetic memory Sagnik Ghosh, Arnab Datta, Colin Kunze, Jane Kondev, Ahmad S Khalil DNA methylation is a process by which cells can regulate gene expression. Recent experiments have used synthetic read-write molecules that bind to methylated sites and then methylate nearby unmethylated ones to create a positive feedback [1]. Additionally, methylated sites turn over to unmethylated sites at a constant rate. The feedback allows the methylation to persist, despite the turnover, giving rise to ‘memory’. We examine a theoretical model of this process by considering a one-dimensional lattice where each site can either be methylated (1) or unmethylated (0). Unlike the canonical Ising model, this model has asymmetric interactions since 1s can convert a 0 to a 1 but not vice versa. We compute the steady state number of methylated sites for interactions ranging from nearest neighbour to infinite range, for which we obtain an analytic solution. As the feedback is increased, while the turnover rate is kept constant, we find that the average steady state methylation jumps from zero to a non zero value, reminiscent of a phase transition. We also compute the relaxation time to the steady state and analyze how it depends on the strength of the feedback and the range of interactions. |
Monday, March 14, 2022 10:12AM - 10:24AM |
A04.00008: Deciphering the influence of genome architecture in a minimized bacterial genome Troy A Brier, Pratap Venepally, John I Glass, Zaida (Zan) Luthey-Schulten Probabilistic interactions of RNA polymerase with DNA dictate transcription, the process which evolves the bacterial transcriptome composed of rRNA, tRNA, mRNA, and sRNA. The boundaries of the transcriptional events are outlined by the genome architecture defined as the local arrangement of genetic features identified via sequence motifs (promoters, gene coding regions, and transcription termination sites, etc.) along the genome. The local arrangement of these sequence motifs form operon-like regions, transcriptional units, directly controlling the expression of encoded RNA species. Using bioinformatic analysis of these genetic features, their locations were predicted computationally, and integrated together to define transcriptional units in the minimized genomes of the minimal cell, JCVI-syn3A, and its previous iteration, JCVI-syn1.0. The computational predictions correlate well or strongly agree with an equivalent experimental genome architecture identification approach using Oxford Nanopore Technologies which provide long read native RNA sequencing. Comparison between the two genome architectures has resolved factors influencing the evolution of the transcriptome and potential downstream impacts on cellular processes. |
Monday, March 14, 2022 10:24AM - 10:36AM |
A04.00009: A novel riboswitch design for integrative, functional RNA detection in mammalian cells. Rafał Krzysztoń, Yiming Wan, Julia Petreczky, Gabor Balazsi Aberrant RNA expression profiles are characteristic to many pathologies including neurodegenerative diseases, viral infection or cancer and serve as sensitive biomarkers. Currently, RNA detection in mammalian cells involves cell lysis (RT qPCR), cell fixation (RNA-FISH) or laborious, costly protocols (RNAseq). Despite their diagnostic power such methods do not allow for in vivo RNA detection and cannot be used to trigger corrective effects in affected living human cells for therapeutic purposes. Here we propose a strategy for detection of mRNA or short RNAs by trigger-specific control of mammalian translation. We designed a novel class of riboswitches taking into account the mechanisms of translation initiation in eukaryotes. We demonstrate the efficiency of designed riboswitches in detecting trigger RNA inside living cells. Finally, combining experiment and computational modeling, we identify and characterize the design rules for their optimal performance. |
Monday, March 14, 2022 10:36AM - 10:48AM |
A04.00010: The state of water in lipidic mesophase Yang Yao, Sara Catalini, Bence Kutus, Johannes Hunger, Tao Zhou, Paolo Foggi, George Floudas, Raffaele Mezzenga Water is the most ubiquitous and essential liquid on earth, and it is fundamental to the existence of life. Many biological processes take place in crowded aqueous surroundings and water in living cells can be considered as confined water. However, water behaves differently under confinement with respect to the crystallization and the molecular dynamics compared to those in the bulk. Especially in biological systems, the role of confined water is of crucial importance though still far from fully understood. Here I will present our recent studies on water under soft confining media provided by lipidic mesophase (LMP). Formerly in our group, an unfrozen lipid was synthesized which opens the gate for the study of water in LMPs at subzero temperatures. Yet, the complex synthetic procedures with low yields narrows the large-scale application. We recently discoveried a commercially available lipid, phytantiol, which gives the access to the liquid water in LMPs at subzero temperatures. I will first discuss the crystallization and dynamics of water confined in an unfrozen LMP based on phytantriol at subzero temperatures. Then I will discuss the state of water in lipidic mesophase during the phase transition from bicontinuous cubic phase to reverse hexagonal phase. |
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