A tunable dual-input system for on-demand dynamic gene expression regulation

Pedone, Elisa; Postiglione, Lorena; Aulicino, Francesco; Rocca, Dan L.; Montes-Olivas, Sandra; Khazim, Mahmoud; Bernardo, Diego di; Cosma, María Pía; Marucci, Lucia

doi:10.1038/s41467-019-12329-9

Cited by 42 publications

(35 citation statements)

References 69 publications

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“…Pelleted cells from a subconfluent 75-cm flask were resuspended in 500 l of complete medium and chip-loaded as in Ref. 30. Cells were cultured for 24 h in a tissue culture incubator (5% CO 2 , 37°C) under constant perfusion with standard or kenpaullone-containing medium.…”

Section: Microfluidics/microscopy-based Time Lapsesmentioning

confidence: 99%

A dual druggable genome-wide siRNA and compound library screening approach identifies modulators of parkin recruitment to mitochondria

Scott

Buckner

Fernández-Albert

et al. 2020

Journal of Biological Chemistry

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Genetic and biochemical evidence points to an association between mitochondrial dysfunction and Parkinson's disease (PD). PD-associated mutations in several genes have been identified and include those encoding PTEN-induced putative kinase 1 (PINK1) and parkin. To identify genes, pathways, and pharmacological targets that modulate the clearance of damaged or old mitochondria (mitophagy), here we developed a high-content imaging-based assay of parkin recruitment to mitochondria and screened both a druggable genome-wide siRNA library and a small neuroactive compound library. We used a multiparameter principal component analysis and an unbiased parameter-agnostic machine-learning approach to analyze the siRNA-based screening data. The hits identified in this analysis included specific genes of the ubiquitin proteasome system, and inhibition of ubiquitin-conjugating enzyme 2 N (UBE2N) with a specific antagonist, Bay 11-7082, indicated that UBE2N modulates parkin recruitment and downstream events in the mitophagy pathway. Screening of the compound library identified kenpaullone, an inhibitor of cyclin-dependent kinases and glycogen synthase kinase 3, as a modulator of parkin recruitment. Validation studies revealed that kenpaullone augments the mitochondrial network and protects against the complex I inhibitor MPP+. Finally, we used a microfluidics platform to assess the timing of parkin recruitment to depolarized mitochondria and its modulation by kenpaullone in real time and with single-cell resolution. We demonstrate that the high-content imaging-based assay presented here is suitable for both genetic and pharmacological screening approaches, and we also provide evidence that pharmacological compounds modulate PINK1-dependent parkin recruitment.

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Section: Microfluidics/microscopy-based Time Lapsesmentioning

confidence: 99%

A dual druggable genome-wide siRNA and compound library screening approach identifies modulators of parkin recruitment to mitochondria

Scott

Buckner

Fernández-Albert

et al. 2020

Journal of Biological Chemistry

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“…Some attempts have also been made to use control engineering principles to design cellular systems able to adapt to fluctuating environments (Hsiao et al, 2018). To date, these attempts have mostly focused on the basic genetic parts and regulatory motifs (e.g., negative feedback) needed to implement control algorithms (Ceroni et al, 2018;Aoki et al, 2019;Pedone et al, 2019;Bartoli et al, 2020). Moving forward, multi-agent models offer a means to make simulations of these systems more realistic by accurately capturing how individual cells and their complex environment change over time.…”

Section: Challenges In Scaling-up Biotechnologymentioning

confidence: 99%

Toward Engineering Biosystems With Emergent Collective Functions

Gorochowski

Hauert

Kreft

et al. 2020

Front. Bioeng. Biotechnol.

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Many complex behaviors in biological systems emerge from large populations of interacting molecules or cells, generating functions that go beyond the capabilities of the individual parts. Such collective phenomena are of great interest to bioengineers due to their robustness and scalability. However, engineering emergent collective functions is difficult because they arise as a consequence of complex multi-level feedback, which often spans many length-scales. Here, we present a perspective on how some of these challenges could be overcome by using multi-agent modeling as a design framework within synthetic biology. Using case studies covering the construction of synthetic ecologies to biological computation and synthetic cellularity, we show how multi-agent modeling can capture the core features of complex multi-scale systems and provide novel insights into the underlying mechanisms which guide emergent functionalities across scales. The ability to unravel design rules underpinning these behaviors offers a means to take synthetic biology beyond single molecules or cells and toward the creation of systems with functions that can only emerge from collectives at multiple scales.

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“…Designing synthetic gene networks by modeling and integrating them within WCM formalisms [as in Purcell et al (2013)] could be critical to investigate how gene expression correlates with codon usage, explore possible cell burden effects (Borkowski et al, 2016), and predict modularity of synthetic gene networks and tools to modulate gene expression across different chassis (Way et al, 2014;Pedone et al, 2019;Gomide et al, 2020).…”

Section: Model Granularity Of Gene Network (Re)designmentioning

confidence: 99%

Computer-Aided Whole-Cell Design: Taking a Holistic Approach by Integrating Synthetic With Systems Biology

Marucci

Barberis

Karr

et al. 2020

Front. Bioeng. Biotechnol.

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Computer-aided design (CAD) for synthetic biology promises to accelerate the rational and robust engineering of biological systems. It requires both detailed and quantitative mathematical and experimental models of the processes to (re)design biology, and software and tools for genetic engineering and DNA assembly. Ultimately, the increased precision in the design phase will have a dramatic impact on the production of designer cells and organisms with bespoke functions and increased modularity. CAD strategies require quantitative models of cells that can capture multiscale processes and link genotypes to phenotypes. Here, we present a perspective on how whole-cell, multiscale models could transform design-build-test-learn cycles in synthetic biology. We show how these models could significantly aid in the design and learn phases while reducing experimental testing by presenting case studies spanning from genome minimization to cell-free systems. We also discuss several challenges for the realization of our vision. The possibility to describe and build whole-cells in silico offers an opportunity to develop increasingly automatized, precise and accessible CAD tools and strategies.

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A tunable dual-input system for on-demand dynamic gene expression regulation

Cited by 42 publications

References 69 publications

A dual druggable genome-wide siRNA and compound library screening approach identifies modulators of parkin recruitment to mitochondria

A dual druggable genome-wide siRNA and compound library screening approach identifies modulators of parkin recruitment to mitochondria

Toward Engineering Biosystems With Emergent Collective Functions

Computer-Aided Whole-Cell Design: Taking a Holistic Approach by Integrating Synthetic With Systems Biology

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