Structures, mechanisms and applications of RNA-centric CRISPR–Cas13

Yang, Hui; Patel, Dinshaw J.

doi:10.1038/s41589-024-01593-6

Cited by 2 publications

References 147 publications

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Visualizing DNA/RNA, Proteins, and Small Molecule Metabolites within Live Cells

Jia,

Cui,

Ding

2024

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Live cell imaging is essential for obtaining spatial and temporal insights into dynamic molecular events within heterogeneous individual cells, in situ intracellular networks, and in vivo organisms. Molecular tracking in live cells is also a critical and general requirement for studying dynamic physiological processes in cell biology, cancer, developmental biology, and neuroscience. Alongside this context, this review provides a comprehensive overview of recent research progress in live‐cell imaging of RNAs, DNAs, proteins, and small‐molecule metabolites, as well as their applications in molecular diagnosis, immunodiagnosis, and biochemical diagnosis. A series of advanced live‐cell imaging techniques have been introduced and summarized, including high‐precision live‐cell imaging, high‐resolution imaging, low‐abundance imaging, multidimensional imaging, multipath imaging, rapid imaging, and computationally driven live‐cell imaging methods, all of which offer valuable insights for disease prevention, diagnosis, and treatment. This review article also addresses the current challenges, potential solutions, and future development prospects in this field.

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Visualizing DNA/RNA, Proteins, and Small Molecule Metabolites within Live Cells

Jia,

Cui,

Ding

2024

Small

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CRISPR-RNA binding drives structural ordering that primes Cas7-11 for target cleavage

Lin,

Li,

Brogan

et al. 2024

Preprint

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Type III-E CRISPR-Cas effectors, of which Cas7-11 is the first, are single proteins that cleave target RNAs without nonspecific collateral cleavage, opening new possibilities for RNA editing. Biochemical experiments combined with amide hydrogen-deuterium exchange (HDX-MS) experiments provide a first glimpse of the conformational dynamics of apo Cas7-11. HDX-MS revealed the backbone comprised of the four Cas7 zinc-binding RRM folds are well-folded but insertion sequences are highly dynamic and fold upon binding crRNA. The crRNA causes folding of disordered catalytic loops and β-hairpins, stronger interactions at domain-domain interfaces, and folding of the Cas7.1 processing site. Target RNA binding causes only minor ordering around the catalytic loops of Cas7.2 and Cas7.3. We show that Cas7-11 cannot fully process the CRISPR array and that binding of partially processed crRNA induces multiple states in Cas7-11 and reduces target RNA cleavage. The insertion domain shows the most ordering upon binding of mature crRNA. Finally, we show a crRNA-induced conformational change in one of the TPR-CHAT binding sites providing an explanation for why crRNA binding facilitates TPR-CHAT binding. The results provide the first glimpse of the apo state of Cas7-11 and reveal how its structure and function are regulated by crRNA binding.

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Structures, mechanisms and applications of RNA-centric CRISPR–Cas13

Cited by 2 publications

References 147 publications

Visualizing DNA/RNA, Proteins, and Small Molecule Metabolites within Live Cells

Visualizing DNA/RNA, Proteins, and Small Molecule Metabolites within Live Cells

CRISPR-RNA binding drives structural ordering that primes Cas7-11 for target cleavage

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