Dataset Information

Anti-CRISPR-mediated control of gene editing and synthetic circuits in eukaryotic cells.

ABSTRACT: Repurposed CRISPR-Cas molecules provide a useful tool set for broad applications of genomic editing and regulation of gene expression in prokaryotes and eukaryotes. Recent discovery of phage-derived proteins, anti-CRISPRs, which serve to abrogate natural CRISPR anti-phage activity, potentially expands the ability to build synthetic CRISPR-mediated circuits. Here, we characterize a panel of anti-CRISPR molecules for expanded applications to counteract CRISPR-mediated gene activation and repression of reporter and endogenous genes in various cell types. We demonstrate that cells pre-engineered with anti-CRISPR molecules become resistant to gene editing, thus providing a means to generate "write-protected" cells that prevent future gene editing. We further show that anti-CRISPRs can be used to control CRISPR-based gene regulation circuits, including implementation of a pulse generator circuit in mammalian cells. Our work suggests that anti-CRISPR proteins should serve as widely applicable tools for synthetic systems regulating the behavior of eukaryotic cells.

SUBMITTER: Nakamura M

PROVIDER: S-EPMC6331597 | biostudies-literature | 2019 Jan

REPOSITORIES: biostudies-literature

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Anti-CRISPR-mediated control of gene editing and synthetic circuits in eukaryotic cells.

Nakamura Muneaki M Srinivasan Prashanth P Chavez Michael M Carter Matthew A MA Dominguez Antonia A AA La Russa Marie M Lau Matthew B MB Abbott Timothy R TR Xu Xiaoshu X Zhao Dehua D Gao Yuchen Y Kipniss Nathan H NH Smolke Christina D CD Bondy-Denomy Joseph J Qi Lei S LS

Nature communications 20190114 1

Repurposed CRISPR-Cas molecules provide a useful tool set for broad applications of genomic editing and regulation of gene expression in prokaryotes and eukaryotes. Recent discovery of phage-derived proteins, anti-CRISPRs, which serve to abrogate natural CRISPR anti-phage activity, potentially expands the ability to build synthetic CRISPR-mediated circuits. Here, we characterize a panel of anti-CRISPR molecules for expanded applications to counteract CRISPR-mediated gene activation and repressio ...[more]

PMID: 30643127

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Project description:BackgroundThe modular design of synthetic gene circuits via composable parts (DNA segments) and pools of signal carriers (molecules such as RNA polymerases and ribosomes) has been successfully applied to bacterial systems. However, eukaryotic cells are becoming a preferential host for new synthetic biology applications. Therefore, an accurate description of the intricate network of reactions that take place inside eukaryotic parts and pools is necessary. Rule-based modeling approaches are increasingly used to obtain compact representations of reaction networks in biological systems. However, this approach is intrinsically non-modular and not suitable per se for the description of composable genetic modules. In contrast, the Model Description Language (MDL) adopted by the modeling tool ProMoT is highly modular and it enables a faithful representation of biological parts and pools.ResultsWe developed a computational framework for the design of complex (eukaryotic) gene circuits by generating dynamic models of parts and pools via the joint usage of the BioNetGen rule-based modeling approach and MDL. The framework converts the specification of a part (or pool) structure into rules that serve as inputs for BioNetGen to calculate the part's species and reactions. The BioNetGen output is translated into an MDL file that gives a complete description of all the reactions that take place inside the part (or pool) together with a proper interface to connect it to other modules in the circuit. In proof-of-principle applications to eukaryotic Boolean circuits with more than ten genes and more than one thousand reactions, our framework yielded proper representations of the circuits' truth tables.ConclusionsFor the model-based design of increasingly complex gene circuits, it is critical to achieve exact and systematic representations of the biological processes with minimal effort. Our computational framework provides such a detailed and intuitive way to design new and complex synthetic gene circuits.

Dataset Information

Anti-CRISPR-mediated control of gene editing and synthetic circuits in eukaryotic cells.

Publications

Anti-CRISPR-mediated control of gene editing and synthetic circuits in eukaryotic cells.

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