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Biological Chemistry

Easier Route To Genomic Changes

Genetic Engineering: Zinc finger nucleases induce duplications, inversions of DNA sequences

by Stu Borman
January 2, 2012 | A version of this story appeared in Volume 90, Issue 1

Zinc finger nucleases (scissors) make breaks in genomic double-stranded DNA that cells then repair to create duplications (left) and inversions (right).
This image shows how zinc fingers can change DNA sequencing.
Zinc finger nucleases (scissors) make breaks in genomic double-stranded DNA that cells then repair to create duplications (left) and inversions (right).

A research group in South Korea has shown for the first time that zinc finger nucleases can trigger sequence duplications and inversions at specific sites in living cells’ chromosomes. The researchers also engineered variations that corrected disease-associated genetic defects.

The technique could be used in gene therapy applications and in studies of how genomic variations affect the function of live cells or organisms.

Genomic variations such as deleted segments, duplicate sequences, and inversions (sequence reversals) often have significant effects on the health of cells and organisms. For example, DNA inversions cause almost half of severe hemophilia A cases.

But in many instances the variations’ consequences are poorly understood, in part because researchers lack simple, effective techniques to engineer them into specific genomic sites. Existing methods either introduce changes only at random sites or require laborious premanipulation of genomic sequences in cells, which also leaves behind undesired foreign sequence elements in the genome.

Now, associate professor of chemistry Jin-Soo Kim and coworkers at Seoul National University have used zinc finger nucleases—synthetic DNA-cleaving enzymes made from zinc-coordinating protein motifs—to create such variations more easily (Genome Res., DOI: 10.1101/gr.129635.111).

The researchers designed zinc finger nucleases that target specific genomic sequences in human embryonic kidney cells. The nucleases create DNA double-strand breaks that then induce cellular DNA-repair mechanisms to create duplications or inversions at the sites. Sequence translocations have been achieved this way before, but “we are the first to demonstrate that zinc finger nucleases can be used to induce duplications and inversions,” Kim says.

He and his coworkers detected, isolated, and characterized those variations. They also demonstrated that their nucleases could reverse the abnormal genomic inversion associated with hemophilia A.

They believe the technique could eventually be used to correct genomic aberrations in pluripotent stem cells of patients. Once corrected, the cells could then be differentiated into appropriate types of somatic cells and reintroduced into patients.

“Perhaps the most intriguing aspect of this study is the ability to generate individual structural variations at will so their biological consequences can be studied,” says biochemistry and molecular biology professor John H. Wilson of Baylor College of Medicine, who specializes in targeted genome modification. The new approach “should provide a useful gateway for such studies.”

The technique also “potentially allows for the reversal of some [sequence] rearrangements in patients,” although “application to patients will require surmounting other substantial obstacles,” Wilson notes.



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