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    Home > Active Ingredient News > Study of Nervous System > Cell: The next generation of gene editing technology CRISPRoff opens the surface...

    Cell: The next generation of gene editing technology CRISPRoff opens the surface...

    • Last Update: 2021-05-10
    • Source: Internet
    • Author: User
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    Text | Li Yuan

    Genome editing tools represented by the CRISPR/Cas9 system need to change the DNA sequence when functioning, and often face safety issues caused by off-target.

    More and more studies have proved that the epigenome plays a central role in viral infections and cancer.
    Epigenetics affect gene expression through DNA methylation.
    When a methyl group tag is attached to DNA, it can cause gene silencing, and the epigenome is heritable like the genome.

    On April 9, in a paper published in Cell, researchers from the University of California, San Francisco and Massachusetts Institute of Technology developed a new type of epigenetic genome editing tool "CRISPRoff".
    Silencing almost all genes in human cells while changing the genetic code.
    This silencing will be maintained for hundreds of generations in the offspring of the cell and is reversible.
    The silenced genes can be reactivated after using another "CRISPRon" tool.

    Source: Cell

    Source: Cell

    The CRISPR system can become an effective gene editing tool due to two important functional components: one is DNA endonuclease that performs DNA cutting function.
    The other is the homing device, which can play a specific targeting function and can precisely control the editing position.

    In order to construct CRISPRoff that acts on the epigenome, the researchers retained the homing device of CRISPR, removed the function of the DNA-cutting enzyme, and then added an enzyme that acts on the epigenome.
    Of the two CRISPRoff tools, the silencing effect of CRISPRoff-V2 is more powerful.

    Source: Cell

    Source: Cell

    Researchers used this tool to target the silencing of endogenous CLTA genes, observe the persistence of epigenetic memory, and detect the expression of CLTA in single-cell clones.
    The results showed that this gene not only remained silent in the treated cells, but also the CLTA gene in 38 clones (39 in total) remained silent 15 months after transfection or after 450 generations of cell division.

    Source: Cell

    Source: Cell

    In addition to its excellent durability characteristics, CRISPRoff is also reversible.
    Using another CRISPRon tool can remove the methylation markers programmed by CRISPRoff, reactivate the silenced genes, and realize the complete reversibility of this process.

    CRISPRon reverses gene silencing (Source: Cell)

    CRISPRon reverses gene silencing (Source: Cell)

    The traditional view is that methylation can only silence cytosine in CpG islands (CGI), and nearly one-third of human genes lack CpG islands, so researchers suspect that about 30% of human genes do not respond to new tools.
    But the performance of CRISPRoff overturned this view.
    CRISPRoff's epigenetic editing is not limited to genes with typical CpG islands, but can also silence genes without CpG island structures.

    CRISPRoff silences genes without CpG island structure (Source: Cell)

    CRISPRoff silences genes without CpG island structure (Source: Cell)

    Finally, the researchers explored the therapeutic potential of CRISPRoff in induced pluripotent stem cells.
    Although the epigenome changes adaptively during the differentiation of stem cells into mature cells, researchers have observed that 90% of the cells differentiated by stem cells whose genes have been silenced by CRISPRoff remain encoded by CRISPRoff.
    Gene silencing.

    CRISPRoff gene silencing in iPSC and iPSC differentiated neurons (Source: Cell)

    CRISPRoff gene silencing in iPSC and iPSC differentiated neurons (Source: Cell)

    This shows that CRISPRoff can achieve long-lasting treatment with only one administration, which reflects the huge therapeutic potential of CRISPRoff.

    In general, this programmable epigenome editing is adjustable and reversible without cutting DNA, thereby effectively avoiding the cytotoxicity associated with gene editing.
    But the researchers also pointed out that the full therapeutic potential still needs further work to realize.

    Reference materials:

    Reference materials:

    [1] James Nuñez et al.
    Genome-wide programmable transcriptional memory by CRISPR-based epigenome editing.
    Cell (2021)

    [1] James Nuñez et al.
    Genome-wide programmable transcriptional memory by CRISPR-based epigenome editing.
    Cell (2021)

    [2] New CRISPR Technology Offers Unrivaled Control of Epigenetic Inheritance (Source: UCSF)

    [2] New CRISPR Technology Offers Unrivaled Control of Epigenetic Inheritance (Source: UCSF)

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