Browsing by Author "Singh, Abhyudai"
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Item A mechanistic stochastic framework for regulating bacterial cell division(Nature Publishing Group, 2016-07-26) Ghusinga,Khem Raj; Vargas-Garcia,Cesar A.; Singh,Abhyudai; Khem Raj Ghusinga, Cesar A. Vargas-Garcia,Abhyudai Singh; Singh, AbhyudaiHow exponentially growing cells maintain size homeostasis is an important fundamental problem. Recent single-cell studies in prokaryotes have uncovered the adder principle, where cells add a fixed size (volume) from birth to division, irrespective of their size at birth. To mechanistically explain the adder principle, we consider a timekeeper protein that begins to get stochastically expressed after cell birth at a rate proportional to the volume. Cell-division time is formulated as the first-passage time for protein copy numbers to hit a fixed threshold. Consistent with data, the model predicts that the noise in division timing increases with size at birth. Intriguingly, our results show that the distribution of the volume added between successive cell-division events is independent of the newborn cell size. This was dramatically seen in experimental studies, where histograms of the added volume corresponding to different newborn sizes collapsed on top of each other. The model provides further insights consistent with experimental observations: the distribution of the added volume when scaled by its mean becomes invariant of the growth rate. In summary, our simple yet elegant model explains key experimental findings and suggests a mechanism for regulating both the mean and fluctuations in cell-division timing for controlling size.Item A comparative approach to stabilizing mechanisms between discrete- and continuous-time consumer-resource models(PLoS ONE, 2022-04-12) Singh, AbhyudaiThere is rich literature on using continuous-time and discrete-time models for studying population dynamics of consumer-resource interactions. A key focus of this contribution is to systematically compare between the two modeling formalisms the stabilizing/destabilizing impacts of diverse ecological processes that result in a density-dependent consumer attack rate. Inspired by the Nicholson-Bailey/Lotka-Volterra models in discrete-time/continuous-time, respectively, we consider host-parasitoid interactions with an arbitrary parasitoid attack rate that is a function of both the host/parasitoid population densities. Our analysis shows that a Type II functional response is stabilizing in both modeling frameworks only when combined with other mechanisms, such as mutual interference between parasitoids. A Type III functional response is by itself stabilizing, but the extent of attack-rate acceleration needed is much higher in the discrete-time framework, and its stability regime expands with increasing host reproduction. Finally, our results show that while mutual parasitoid interference can stabilize population dynamics, cooperation between parasitoids to handle hosts is destabilizing in both frameworks. In summary, our comparative analysis systematically characterizes diverse ecological processes driving stable population dynamics in discrete-time and continuous-time consumer-resource models.Item Fundamental limits of parasitoid-driven host population suppression: Implications for biological control(PLoS ONE, 2023-12-22) Singh, AbhyudaiParasitoid wasps are increasingly being used to control insect pest populations, where the pest is the host species parasitized by the wasp. Here we use the discrete-time formalism of the Nicholson-Bailey model to investigate a fundamental question—are there limits to parasitoid-driven suppression of the host population density while still ensuring a stable coexistence of both species? Our model formulation imposes an intrinsic self-limitation in the host’s growth resulting in a carrying capacity in the absence of the parasitoid. Different versions of the model are considered with parasitism occurring at a developmental stage that is before, during, or after the growth-limiting stage. For example, the host’s growth limitation may occur at its larval stage due to intraspecific competition, while the wasps attack either the host egg, larval or pupal stage. For slow-growing hosts, models with parasitism occurring at different life stages are identical in terms of their host suppression dynamics but have contrasting differences for fast-growing hosts. In the latter case, our analysis reveals that wasp parasitism occurring after host growth limitation yields the lowest pest population density conditioned on stable host-parasitoid coexistence. For ecologically relevant parameter regimes we estimate this host suppression to be roughly 10-20% of the parasitoid-free carrying capacity. We further expand the models to consider a fraction of hosts protected from parasitism (i.e., a host refuge). Our results show that for a given host reproduction rate there exists a critical value of protected host fraction beyond which, the system dynamics are stable even for high levels of parasitism that drive the host to arbitrary low population densities. In summary, our systematic analysis sheds key insights into the combined effects of density-dependence in host growth and parasitism refuge in stabilizing the host-parasitoid population dynamics with important implications for biological control.Item Genome-wide screening reveals metabolic regulation of stop-codon readthrough by cyclic AMP(Nucleic Acids Research, 2023-09-06) Lyu, Zhihui; Villanueva, Patricia; O’Malley, Liam; Murphy, Parker; Augenstreich, Jacques; Briken, Volker; Singh, Abhyudai; Ling, JiqiangTranslational fidelity is critical for microbial fitness, survival and stress responses. Much remains unknown about the genetic and environmental control of translational fidelity and its single-cell heterogeneity. In this study, we used a high-throughput fluorescence-based assay to screen a knock-out library of Escherichia coli and identified over 20 genes critical for stop-codon readthrough. Most of these identified genes were not previously known to affect translational fidelity. Intriguingly, we show that several genes controlling metabolism, including cyaA and crp, enhance stop-codon readthrough. CyaA catalyzes the synthesis of cyclic adenosine monophosphate (cAMP). Combining RNA sequencing, metabolomics and biochemical analyses, we show that deleting cyaA impairs amino acid catabolism and production of ATP, thus repressing the transcription of rRNAs and tRNAs to decrease readthrough. Single-cell analyses further show that cAMP is a major driver of heterogeneity in stop-codon readthrough and rRNA expression. Our results highlight that carbon metabolism is tightly coupled with stop-codon readthrough. Graphical Abstract available at: https://doi.org/10.1093/nar/gkad725Item Intercellular Variability in Protein Levels from Stochastic Expression and Noisy Cell Cycle Processes(Public Library Science, 2016-08-18) Soltani,Mohammad; Vargas-Garcia,Cesar A.; Antunes,Duarte; Singh,Abhyudai; Mohammad Soltani, Cesar A. Vargas-Garcia, Duarte Antunes, Abhyudai Singh; Singh, AbhyudaiInside individual cells, expression of genes is inherently stochastic and manifests as cell-to-cell variability or noise in protein copy numbers. Since proteins half-lives can be comparable to the cell-cycle length, randomness in cell-division times generates additional intercellular variability in protein levels. Moreover, as many mRNA/protein species are expressed at low-copy numbers, errors incurred in partitioning of molecules between two daughter cells are significant. We derive analytical formulas for the total noise in protein levels when the cell-cycle duration follows a general class of probability distributions. Using a novel hybrid approach the total noise is decomposed into components arising from i) stochastic expression; ii) partitioning errors at the time of cell division and iii) random cell-division events. These formulas reveal that random cell-division times not only generate additional extrinsic noise, but also critically affect the mean protein copy numbers and intrinsic noise components. Counter intuitively, in some parameter regimes, noise in protein levels can decrease as cell-division times become more stochastic. Computations are extended to consider genome duplication, where transcription rate is increased at a random point in the cell cycle. We systematically investigate how the timing of genome duplication influences different protein noise components. Intriguingly, results show that noise contribution from stochastic expression is minimized at an optimal genome-duplication time. Our theoretical results motivate new experimental methods for decomposing protein noise levels from synchronized and asynchronized single-cell expression data. Characterizing the contributions of individual noise mechanisms will lead to precise estimates of gene expression parameters and techniques for altering stochasticity to change phenotype of individual cells.Item Modeling protein concentrations in cycling cells using stochastic hybrid systems(IFAC-PapersOnLine, 2021-07-16) Vahdat, Zahra; Xu, Zikai; Singh, AbhyudaiWe analyze a class of time-triggered stochastic hybrid systems where the state-space evolves as per a linear time-invariant dynamical system. This continuous-time evolution is interspersed with two kinds of stochastic resets. The first reset occurs based on an internal timer that measures the time elapsed since it last occurred. Whenever the first reset occurs, the states-space undergoes a random jump, and the timer is reset to zero. The second reset occurs based on an arbitrary timer-depended rate, and whenever this reset fires, the state-space is changed based on a given random map. We provide exact conditions for this class of systems that lead to finite statistical moments and the corresponding exact analytical expressions for the first two moments. This framework is applied to study random fluctuations in the concentration of a protein in a growing cell. In the context of this example, the timer denotes the time elapsed since the cell was born, and the cell division event (first reset) is triggered based on a timer-dependent rate. The second reset corresponds to the protein synthesis in stochastic bursts, and finally, during cell growth, protein concentration continuously decreases due to dilution. Our analysis provides closed-form formulas for the noise in the protein concentration and leads to a striking result - for a constant (timer-independent) protein synthesis rate, the noise in the protein concentration is invariant of the noise in the cell-cycle time. Finally, we provide a rigorous framework for investigating protein noise levels for different forms of timer-dependent synthesis rates, as is the case for cell-cycle regulated genes inside the cell.Item Regulating gene expression to achieve temporal precision(IFAC-PapersOnLine, 2021-07-16) Ghusinga, Khem Raj; Singh, AbhyudaiCellular response to a stimulus is often triggered by accumulation of an appropriate gene product up to a critical threshold. How cells regulate gene expression to achieve precision in timing of such responses is a question of interest. Earlier work has shown that for a stable gene product, a constant rate of accumulation provides minimum noise in the time of response, provided that initial gene product distribution is degenerate. Here, we show that this strategy is no longer optimal if the initial gene product level is drawn from a non-degenerate distribution. We also discuss biological relevance of this finding.Item Tradeoff between lag time and growth rate drives the plasmid acquisition cost(Nature Communications, 2023-04-24) Ahmad, Mehrose; Prensky, Hannah; Balestrieri, Jacqueline; ElNaggar, Shahd; Gomez-Simmonds, Angela; Uhlemann, Anne-Catrin; Traxler, Beth; Singh, Abhyudai; Lopatkin, Allison J.Conjugative plasmids drive genetic diversity and evolution in microbial populations. Despite their prevalence, plasmids can impose long-term fitness costs on their hosts, altering population structure, growth dynamics, and evolutionary outcomes. In addition to long-term fitness costs, acquiring a new plasmid introduces an immediate, short-term perturbation to the cell. However, due to the transient nature of this plasmid acquisition cost, a quantitative understanding of its physiological manifestations, overall magnitudes, and population-level implications, remains unclear. To address this, here we track growth of single colonies immediately following plasmid acquisition. We find that plasmid acquisition costs are primarily driven by changes in lag time, rather than growth rate, for nearly 60 conditions covering diverse plasmids, selection environments, and clinical strains/species. Surprisingly, for a costly plasmid, clones exhibiting longer lag times also achieve faster recovery growth rates, suggesting an evolutionary tradeoff. Modeling and experiments demonstrate that this tradeoff leads to counterintuitive ecological dynamics, whereby intermediate-cost plasmids outcompete both their low and high-cost counterparts. These results suggest that, unlike fitness costs, plasmid acquisition dynamics are not uniformly driven by minimizing growth disadvantages. Moreover, a lag/growth tradeoff has clear implications in predicting the ecological outcomes and intervention strategies of bacteria undergoing conjugation.Item Transcriptional Bursting Explains the Noise-Versus-Mean Relationship in mRNA and Protein Levels(Public Library Science, 2016-07-28) Dar,Roy D.; Shaffer,Sydney M.; Singh,Abhyudai; Razooky,Brandon S.; Simpson,Michael L.; Raj,Arjun; Weinberger,Leor S.; Roy D. Dar, Sydney M. Shaffer, Abhyudai Singh, Brandon S. Razooky, Michael L. Simpson, Arjun Raj, Leor S. Weinberger; Singh, AbhyudaiRecent analysis demonstrates that the HIV-1 Long Terminal Repeat (HIV LTR) promoter exhibits a range of possible transcriptional burst sizes and frequencies for any mean-expression level. However, these results have also been interpreted as demonstrating that cell-tocell expression variability (noise) and mean are uncorrelated, a significant Developmentiation from previous results. Here, we re-examine the available mRNA and protein abundance data for the HIV LTR and find that noise in mRNA and protein expression scales inversely with the mean along analytically predicted transcriptional burst-sizemanifolds. We then experimentally perturb transcriptional activity to test a prediction of the multiple burst-size model: that increasing burst frequency will cause mRNA noise to decrease along given burst-size lines as mRNA levels increase. The data show that mRNA and protein noise decrease as mean expression increases, supporting the canonical inverse correlation between noise and mean.Item Transcriptional regulation of Sis1 promotes fitness but not feedback in the heat shock response(eLife, 2023-05-17) Grade, Rania; Singh, Abhyudai; Ali, Asif; Pincus, DavidThe heat shock response (HSR) controls expression of molecular chaperones to maintain protein homeostasis. Previously, we proposed a feedback loop model of the HSR in which heat-denatured proteins sequester the chaperone Hsp70 to activate the HSR, and subsequent induction of Hsp70 deactivates the HSR (Krakowiak et al., 2018; Zheng et al., 2016). However, recent work has implicated newly synthesized proteins (NSPs) – rather than unfolded mature proteins – and the Hsp70 co-chaperone Sis1 in HSR regulation, yet their contributions to HSR dynamics have not been determined. Here, we generate a new mathematical model that incorporates NSPs and Sis1 into the HSR activation mechanism, and we perform genetic decoupling and pulse-labeling experiments to demonstrate that Sis1 induction is dispensable for HSR deactivation. Rather than providing negative feedback to the HSR, transcriptional regulation of Sis1 by Hsf1 promotes fitness by coordinating stress granules and carbon metabolism. These results support an overall model in which NSPs signal the HSR by sequestering Sis1 and Hsp70, while induction of Hsp70 – but not Sis1 – attenuates the response.