In Vivo Validation of a Reversible Small Molecule-Based Switch for Synthetic Self-Amplifying mRNA Regulation

Cafferty, Séan Mc; Temmerman, Joyca De; Kitada, Tasuku; Becraft, Jacob; Weiss, Ron; Irvine, Darrell J.; Devreese, Mathias; Baere, Siegrid De; Combes, Francis; Sanders, Niek N.

doi:10.1016/j.ymthe.2020.11.010

Cited by 19 publications

(16 citation statements)

References 57 publications

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“…Thus, microbial RBPs are primarily used in mRNA-based mammalian synthetic biology. The representative microbial RBPs used to regulate synthetic mRNAs are coat proteins derived from the bacteriophages MS2 (MS2CP) [2][3][4][5][6][10][11][12][13][14][15][16][17][18][19][20][21] and PP7 (PP7CP) [6,14,21], the archaeal ribosomal protein L7Ae [2][3][4]6,7,11,12,[21][22][23][24][25], and the tetracycline-responsive repressor protein (TetR) from Escherichia coli [23,26]. Among these, TetR is unique in its ability to conditionally dissociate from the target RNA motif by doxycycline addition.…”

Section: Binding To Target Mrnasmentioning

confidence: 99%

“…Destabilizing domains (also called degrons) that induce rapid degradation of fused proteins are used to regulate the abundance of translational regulator proteins (Figure 2, the 4th row). For example, rapid degradation of a translational repressor protein results in an enhancement of its target mRNA translation [4,6,14,20,23,25]. Conversely, rapid degradation of a translational activator protein diminishes the translation of its target mRNA [20].…”

Section: Destabilizing Proteinsmentioning

confidence: 99%

“…Importantly, the destabilizing activity of the eDHFR-based degron can be inhibited by trimethoprim, an antibiotic that is used clinically. Translational regulator proteins fused with the eDHFRbased degron are rapidly degraded in the absence of trimethoprim, but can be stabilized by the addition of trimethoprim, allowing them to repress or activate the translation of their target mRNAs [4,6,20,23,25]. In addition, we recently showed that the combination of the eDHFR-based degron and photocaged trimethoprim, which can bind the degron only after light irradiation, enables photo-regulatable translation [20].…”

Section: Destabilizing Proteinsmentioning

confidence: 99%

“…One approach to producing an optimal amount of protein is to deliberately control it with an external cue such as small molecule administration or light irradiation. In this case, one uses protein-protein interaction modules [4][5][6]14,20], degrons [4,6,20,23,25], or RBPs [23,26] that can be controlled by small molecules or light. These types of translational regulation systems allow for a gradual decrease or increase in translation by adjusting the dosage of a small molecule or the duration of light irradiation [20].…”

Section: Deliberate Regulation By External Cuesmentioning

confidence: 99%

“…Small molecules such as doxycycline [23,26] or trimethoprim [4,6,20,23,25] are more suitable for whole body-translational regulation than light. Additionally, there are a number of small molecules that can be administered in a noninvasive manner like oral administration.…”

Section: Deliberate Regulation By External Cuesmentioning

confidence: 99%

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Protein-Based Systems for Translational Regulation of Synthetic mRNAs in Mammalian Cells

Nakanishi

2021

Life

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Synthetic mRNAs, which are produced by in vitro transcription, have been recently attracting attention because they can express any transgenes without the risk of insertional mutagenesis. Although current synthetic mRNA medicine is not designed for spatiotemporal or cell-selective regulation, many preclinical studies have developed the systems for the translational regulation of synthetic mRNAs. Such translational regulation systems will cope with high efficacy and low adverse effects by producing the appropriate amount of therapeutic proteins, depending on the context. Protein-based regulation is one of the most promising approaches for the translational regulation of synthetic mRNAs. As synthetic mRNAs can encode not only output proteins but also regulator proteins, all components of protein-based regulation systems can be delivered as synthetic mRNAs. In addition, in the protein-based regulation systems, the output protein can be utilized as the input for the subsequent regulation to construct multi-layered gene circuits, which enable complex and sophisticated regulation. In this review, I introduce what types of proteins have been used for translational regulation, how to combine them, and how to design effective gene circuits.

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Section: Binding To Target Mrnasmentioning

confidence: 99%

Section: Destabilizing Proteinsmentioning

confidence: 99%

Section: Destabilizing Proteinsmentioning

confidence: 99%

Section: Deliberate Regulation By External Cuesmentioning

confidence: 99%

Section: Deliberate Regulation By External Cuesmentioning

confidence: 99%

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Protein-Based Systems for Translational Regulation of Synthetic mRNAs in Mammalian Cells

Nakanishi

2021

Life

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Self-amplifying mRNA-Based Vaccine Technology and Its Mode of Action

Maruggi

Ulmer

Rappuoli³

et al. 2021

Current Topics in Microbiology and Immunology

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Self-amplifying mRNAs derived from the genomes of positive-strand RNA viruses have recently come into focus as a promising technology platform for vaccine development. Non-virally delivered self-amplifying mRNA vaccines have the potential to be highly versatile, potent, streamlined, scalable, and inexpensive. By amplifying their genome and the antigen encoding mRNA in the host cell, the self-amplifying mRNA mimics a viral infection, resulting in sustained levels of the target protein combined with self-adjuvanting innate immune responses, ultimately leading to potent and long-lasting antigen-specific humoral and cellular immune responses. Moreover, in principle, any eukaryotic sequence could be encoded by self-amplifying mRNA without the need to change the manufacturing process, thereby enabling a much faster and flexible research and development timeline than the current vaccines and hence a quicker response to emerging infectious diseases. This chapter highlights the rapid progress made in using non-virally delivered self-amplifying mRNA-based vaccines against infectious diseases in animal models. We provide an overview of the unique attributes of this vaccine approach, summarize the growing body of work defining its mechanism of action, discuss the current challenges and latest advances, and highlight perspectives about the future of this promising technology.

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