A synthetic circuit for buffering gene dosage variation between individual mammalian cells

Yang, Jin; Lee, Jihwan; Land, Michelle A.; Lai, Syeling; Igoshin, Oleg A.; St-Pierre, François

doi:10.1038/s41467-021-23889-0

Cited by 15 publications

(18 citation statements)

References 68 publications

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“…Plasmid buffering in such systems allows for copy number adjustment in response to different growth environments. A recent study by Yang et al created a synthetic circuit for eliminating gene dosage variation in individual mammalian cells 24 . Their method buffers plasmid CN variability in mammalian cells to reduce heterogeneity while we use plasmid CN flexibility to buffer circuit burden.…”

Section: Discussionmentioning

confidence: 99%

Exploiting heterogeneity in coupled, two plasmid systems for dynamic population adaptation

Kumar

Lezia

Hasty

2023

Preprint

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In synthetic multi-plasmid systems, it is standard to use only plasmids with orthogonal replication mechanisms to avoid phenotypic heterogeneity and ensure plasmid stability. In nature, however, microbial populations actively exploit heterogeneity to survive in fluctuating environments. Here we show that the intentional use of distinct plasmids with identical origins of replications (oris) can help an engineered bacterial population adapt to its environment. We find that copy number coupling between distinct plasmids in such systems allows for copy number buffering of an essential, but high-burden construct through the action of a stably maintained, nonessential plasmid. Plasmid coupling also generates population state memory without additional layers of regulatory control. This work reimagines how we design synthetic populations to survive and adapt by strategically giving control back to the cells.

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Section: Discussionmentioning

confidence: 99%

Exploiting heterogeneity in coupled, two plasmid systems for dynamic population adaptation

Kumar

Lezia

Hasty

2023

Preprint

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“…As an example of the latter, a recent study demonstrated a five-fold noise reduction by incorporating negative feedback regulation gene circuits (i.e., TetR repressor fused with a Tet-inhibiting peptide; Guinn and Balázsi, 2019 ). Such synthetic circuits can also be multiplexed orthogonally to probe multigene expressions in contribution of cell phenotypes (Szenk et al, 2020 ) and buffering gene dosage variation across cell populations for a more homogeneous gene expression control (Yang et al, 2021 ).…”

Section: Optogenetics In Genome Modification and Regulationmentioning

confidence: 99%

Optogenetic Application to Investigating Cell Behavior and Neurological Disease

Zhu

Johnson

Schaffer

2022

Front. Cell. Neurosci.

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Cells reside in a dynamic microenvironment that presents them with regulatory signals that vary in time, space, and amplitude. The cell, in turn, interprets these signals and accordingly initiates downstream processes including cell proliferation, differentiation, migration, and self-organization. Conventional approaches to perturb and investigate signaling pathways (e.g., agonist/antagonist addition, overexpression, silencing, knockouts) are often binary perturbations that do not offer precise control over signaling levels, and/or provide limited spatial or temporal control. In contrast, optogenetics leverages light-sensitive proteins to control cellular signaling dynamics and target gene expression and, by virtue of precise hardware control over illumination, offers the capacity to interrogate how spatiotemporally varying signals modulate gene regulatory networks and cellular behaviors. Recent studies have employed various optogenetic systems in stem cell, embryonic, and somatic cell patterning studies, which have addressed fundamental questions of how cell-cell communication, subcellular protein localization, and signal integration affect cell fate. Other efforts have explored how alteration of signaling dynamics may contribute to neurological diseases and have in the process created physiologically relevant models that could inform new therapeutic strategies. In this review, we focus on emerging applications within the expanding field of optogenetics to study gene regulation, cell signaling, neurodevelopment, and neurological disorders, and we comment on current limitations and future directions for the growth of the field.

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“…Studies in the field of biosynthetic gene cluster expression 19 and microbial community engineering 20 have shown that similarities in phylogeny and genotypic profiles can accurately predict metabolic phenotype, which suggests genome relatedness could be a potential predictor of genetic circuit performance. On the other hand, the functional phenotype of expression plasmids and genetic devices has been shown to be coupled to physiological metrics such as growth rate 21, 22 , gene copy number 23, 24 , codon usage bias 25, 26 and growth burden 27, 28 in a number of studies. These studies have however, only considered a single or select combination of physiology metrics as explanatory variables of phenotype within a single model chassis.…”

Section: Introductionmentioning

confidence: 99%

Revealing the chassis-effect on a broad-host-range genetic switch and its concordance with interspecies bacterial physiologies

Chan

Baldwin

Bernstein

2023

Preprint

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Broad-host-range synthetic biology is an emerging frontier that aims to expand our current engineerable domain of microbial hosts for biodesign applications. As more novel species are brought to model status, synthetic biologists are discovering that identically engineered genetic circuits can exhibit different performances depending on the organism it operates within, an observation referred to as the chassis-effect. It remains a major challenge to uncover which genome encoded and physiological biological determinants will underpin chassis effects that govern the performance of engineered genetic devices. In this study, we compared model and novel bacterial hosts to ask whether phylogenomic relatedness or similarity in host physiology is a better predictor of toggle switch performance. This was accomplished using comparative framework based on multivariate statistical approaches to systematically demonstrate the chassis-effect and characterize the performance dynamics of a genetic toggle switch operating within six Gammaproteobacteria. Our results solidify the notion that genetic devices are significantly impacted by host-context. Furthermore, we formally determined that hosts exhibiting more similar metrics of growth and molecular physiology also exhibit more similar toggle switch performance, indicating that specific bacterial physiology underpins measurable chassis effects. The result of this study contributes to the field of broad-host-range synthetic biology by lending increased predictive power to the implementation of genetic devices in less-established microbial hosts.

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A synthetic circuit for buffering gene dosage variation between individual mammalian cells

Cited by 15 publications

References 68 publications

Exploiting heterogeneity in coupled, two plasmid systems for dynamic population adaptation

Exploiting heterogeneity in coupled, two plasmid systems for dynamic population adaptation

Optogenetic Application to Investigating Cell Behavior and Neurological Disease

Revealing the chassis-effect on a broad-host-range genetic switch and its concordance with interspecies bacterial physiologies

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