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CRISPR

CRISPR

Genome editing technologies allow scientists to modify an organism’s DNA via the addition, removal, or rearrangement of genetic material at specific genomic locations. These types of techniques could potentially be used to cure genetic disorders such as hemophilia and sickle cell anemia. One popular and widely used DNA-editing research tool that could lead to safe and effective cures for genetic disorders is the CRISPR-Cas9 system. CRISPR-Cas9 stands for Clustered Regularly Interspaced...

CRISPR and crRNAs

CRISPR and crRNAs

Bacteria and archaea are susceptible to viral infections just like eukaryotes; therefore, they have developed a unique adaptive immune system to protect themselves. Clustered regularly interspaced short palindromic repeats and CRISPR-associated proteins (CRISPR-Cas) are present in more than 45% of known bacteria and 90% of known archaea.
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CRISPR stands for Clustered Regularly Interspaced Short Palindromic Repeats is a adaptive immune system found in bacteria and archaea that protects against viral infections. This system enables prokaryotic cells to identify, remember, and neutralize foreign genetic elements, primarily bacteriophages, by storing fragments of the invader’s DNA as a genetic memory.The CRISPR immune response begins during an initial infection. Cas (CRISPR-associated) proteins play a central role in this...

CRISPR/Cas9 Genome Editing

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The basic reaction of homologous recombination (HR) involves two chromatids that contain DNA sequences sharing a significant stretch of identity. One of these sequences uses a strand from another as a template to synthesize DNA in an enzyme-catalyzed reaction. The final product is a novel amalgamation of the two substrates. To ensure an accurate recombination of sequences, HR is restricted to the S and G2 phases of the cell cycle. At these stages, the DNA has been replicated already and the...

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CRISPR/Cas9-mediated Targeted Integration In Vivo Using a Homology-mediated End Joining-based Strategy

CRISPR/Cas9-mediated Targeted Integration In Vivo Using a Homology-mediated End Joining-based Strategy

Published on: March 12, 2018

Structures of the CRISPR genome integration complex.

Addison V Wright¹, Jun-Jie Liu^1,2, Gavin J Knott¹

¹Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720, USA.

Science (New York, N.Y.)

|July 22, 2017

Summary

This summary is machine-generated.

The Cas1-Cas2 integrase uses DNA structure, not just sequence, to integrate foreign DNA into CRISPR loci. This mechanism, aided by IHF, ensures accurate bacterial adaptive immunity against viruses.

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Gene Digital Circuits Based on CRISPR-Cas Systems and Anti-CRISPR Proteins

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Published on: October 18, 2022

Area of Science:

Molecular Biology
Microbiology
Structural Biology

Background:

CRISPR-Cas systems provide adaptive immunity in bacteria against phages.
The Cas1-Cas2 integrase is essential for acquiring new spacers and integrating foreign DNA into the CRISPR locus.

Purpose of the Study:

To elucidate the structural mechanisms of DNA integration by the Cas1-Cas2 integrase.
To understand how integration site selection is achieved during CRISPR array expansion.

Main Methods:

X-ray crystallography to determine structures of Cas1-Cas2-DNA complexes.
Cryo-electron microscopy for the full CRISPR locus integration complex with IHF.

Main Results:

Crystal structures reveal Cas1-Cas2 bound to donor and target DNA in intermediate and product states.
Cryo-EM structure shows the integration complex including integration host factor (IHF).
Integration site selection is dictated by indirect DNA sequence recognition and sequence-dependent DNA deformation.

Conclusions:

Cas1-Cas2 utilizes DNA structural features, favoring repeat and flanking sequence deformation, for site-specific integration.
IHF binding induces sharp DNA bending, enhancing specificity and efficiency of integration by bringing an upstream motif into contact with Cas1.
These findings explain the mechanism of sequence-dependent DNA structure recognition by Cas1-Cas2 for site-selective CRISPR array expansion in bacterial adaptive immunity.