A gene signal amplifier platform for monitoring the unfolded protein response

Marmolejo, Carlos A. Origel; Bachhav, Bhagyashree; Patibandla, Sahiti D; Yang, Alexander L; Segatori, Laura

doi:10.1038/s41589-020-0497-x

Cited by 12 publications

(12 citation statements)

References 50 publications

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“…Signal-to-noise ratio: a measure of the activated signal from a reporter compared with the basal activity or background. Single-molecule fluorescence in situ hybridization (smFISH): a technique where target RNAs of interest are hybridized with fluorescently labeled oligonucleotides to form subnuclear While most reporter assays directly connect the event of interest to transcriptional activation of the reporter gene, synthetic gene circuits can process signals from reporters to clarify the results of a screen [11,16]. For instance, synthetic circuits can increase the dynamic range of reporter assays, which may be limited by the promoter strength of a gene of interest.…”

Section: Microphysiological Systems (Mps)mentioning

confidence: 99%

“…For instance, synthetic circuits can increase the dynamic range of reporter assays, which may be limited by the promoter strength of a gene of interest. A recently developed signal amplifier expresses a synthetic transcriptional activator in response to activation of the unfolded protein response (UPR) [16]. The UPR is a stress-associated response triggered by the accumulation of unfolded proteins in the endoplasmic reticulum.…”

Section: Microphysiological Systems (Mps)mentioning

confidence: 99%

“…However, while aggregates provide markers of degeneration, their role in degeneration and impact on protein processing pathways remain less clear. By monitoring changes in protein processing through the proteolysis and UPR pathways, recently developed circuits can be used to identify drugs that abrogate UPRassociated neurodegeneration [16,81,83,84]. A circuit that reports on aggregate accumulation in yeast provides one example for how such circuits could be used to identify molecules that limit or accelerate aggregation (Figure 5A) [85].…”

Section: A Vision For Circuit-based Screening In Neurodegenerative Diseasesmentioning

confidence: 99%

“…(G) The dynamic range amplifier works via a synthetic transcription factor that activates reporter expression while also repressing negative regulators of the reporter. Range is expanded by inclusion of a transcriptional repressor (dark blue) that limits reporter expression in response to basal-level signaling while the post-translational repressor (orange) degrades the reporter, leading to enhanced resolution of signaling dynamics[16]. (H) Dynamic decoder circuits work by activating in response to a defined dynamic signaling pattern.…”

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confidence: 99%

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Synthetic gene circuits as tools for drug discovery

Beitz

Oakes

Galloway

2022

Trends in Biotechnology

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Within mammalian systems, there exists enormous opportunity to use synthetic gene circuits to enhance phenotype-based drug discovery, to map the molecular origins of disease, and to validate therapeutics in complex cellular systems. While drug discovery has relied on marker staining and high-content imaging in cell-based assays, synthetic gene circuits expand the potential for precision and speed. Here we present a vision of how circuits can improve the speed and accuracy of drug discovery by enhancing the efficiency of hit triage, capturing disease-relevant dynamics in cell-based assays, and simplifying validation and readouts from organoids and microphysiological systems (MPS). By tracking events and cellular states across multiple length and time scales, circuits will transform how we decipher the causal link between molecular events and phenotypes to improve the selectivity and sensitivity of cell-based assays. Enhancing phenotypic drug discovery with synthetic biologyDevelopment of new drugs requires identification and optimization of functional molecules that often proceeds sequentially through high-throughput discovery screening, hit validation, hit expansion and optimization, and preclinical disease modeling (see Glossary) (Figure 1, Key figure)[1]. Target-based drug discovery (TDD) programs aim to identify drug candidates that modulate a defined biological target that is hypothesized to play a causal role in initiating or sustaining a disease. TDD programs offer precision by directly screening for drug candidate efficacy using purified molecular targets [2]. By contrast, phenotypic drug discovery (PDD) programs aim to identify drug candidates that modulate a physiologically-relevant biological system or cellular signaling pathway and screen for drug efficacy using cell-based assays [2]. Cell-based assays are target-agnostic and provide the opportunity to identify therapeutics that resolve diseaseassociated phenotypic deficits even when the disease etiology remains obscure and targets are undefined. Thus, PDD has recently received renewed interest for its potential to identify first-in-class drugs that act via a novel mechanism of action (MoA) as well as identify drugs for disease with complex or unknown etiology [1][2][3][4]. In order to identify drug candidates that translate to effective therapeutics in the clinic, PDD programs require cell-based assays that faithfully recapitulate and report on disease-relevant processes. To efficiently and effectively screen in phenotypic models, assay development for initial discovery screens must balance throughput with the ability to capture these disease processes [1]. As PDD probes a larger biological space than TDD to identify potential therapeutics, the number of hits from an initial discovery screen in a PDD program may be very large [2]. Following initial discovery screens, hit validation and triage are required to produce a small number of high-confidence candidates to proceed to the resource-intensive stages of hit expansion, compound optimization, and hig...

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Section: Microphysiological Systems (Mps)mentioning

confidence: 99%

Section: Microphysiological Systems (Mps)mentioning

confidence: 99%

Section: A Vision For Circuit-based Screening In Neurodegenerative Diseasesmentioning

confidence: 99%

mentioning

confidence: 99%

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Synthetic gene circuits as tools for drug discovery

Beitz

Oakes

Galloway

2022

Trends in Biotechnology

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“…Although pioneered in single-cell organisms, synthetic biology’s recent expansion to mammalian systems provides a rich landscape of cell fates to examine and engineer [ 36 , 52 , 91 , 106 , 107 ]. As novel tools for studying signaling, chromatin structure, and gene regulation in mammalian systems continue to develop, synthetic biology will enable an unprecedented insight into the mechanism of reprogramming and expand our power to engineer reprogrammed cells.…”

Section: A Vision For the Future Of Synthetic Biology In Reprogrammingmentioning

confidence: 99%

Engineering cell fate: Applying synthetic biology to cellular reprogramming

Wang¹,

Beitz²,

Galloway³

2020

Current Opinion in Systems Biology

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Cell factory engineering: Challenges and opportunities for synthetic biology applications

Bachhav

Rossi

Llanos

et al. 2023

Biotech & Bioengineering

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The production of high‐quality recombinant proteins is critical to maintaining a continuous supply of biopharmaceuticals, such as therapeutic antibodies. Engineering mammalian cell factories presents a number of limitations typically associated with the proteotoxic stress induced upon aberrant accumulation of off‐pathway protein folding intermediates, which eventually culminate in the induction of apoptosis. In this review, we will discuss advances in cell engineering and their applications at different hierarchical levels of control of the expression of recombinant proteins, from transcription and translational to posttranslational modifications and subcellular trafficking. We also highlight challenges and unique opportunities to apply modern synthetic biology tools to the design of programmable cell factories for improved biomanufacturing of therapeutic proteins.

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A gene signal amplifier platform for monitoring the unfolded protein response

Cited by 12 publications

References 50 publications

Synthetic gene circuits as tools for drug discovery

Synthetic gene circuits as tools for drug discovery

Engineering cell fate: Applying synthetic biology to cellular reprogramming

Cell factory engineering: Challenges and opportunities for synthetic biology applications

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