Researchers from University of California—Santa Barbara say they have developed a method of genome editing that gives the user complete spatiotemporal control and also treads lightly on DNA.
"Human cells don't like to take in stuff," says Norbert Reich, a professor in the department of chemistry and biochemistry. The human cell has evolved a "trash disposal" mechanism that isolates and breaks down foreign proteins and other unwanted biomolecules, pathogens, and even damaged cellular structures. So, for researchers working with CRISPR-Cas9 technology, results are only as good as their ability to efficiently bypass this defense mechanism and accurately introduce proteins into animal cells, Reich explains
In a paper published in Small recently, Reich and his team describe such a method. Their technique, which they estimate to be 100 to 1,000 times more efficient than current methods, gives users complete spatiotemporal control of the genome editing delivery, in effect allowing them to decide exactly when and where to release genome editing proteins.
"We can actually hit individual cells," Reich said. "We can even hit parts of a cell so we could release the protein into only a part of the cell. But the main point is that we have the control over where and when this protein that's going to cut the DNA is going to be released."
Key to the Reich group's light-triggered genome editing are hollow gold nanospheres onto which are coated DNA reporter strands and a protein fusion of Cre recombinase and cell-penetrating peptides. And near-infrared light. "So now we've got a homing device and a delivery agent," Reich says, explaining that the Cre recombinase and peptide fusion act as the targeting system, one that goes into play when the target cell does its cellular trash disposal.
Once taken into the cell, the nanoshell is enveloped in an endosome. "But the nanoshells don't do anything because they're entrapped," Reich said. Ultrafast pulsed near-infrared laser light, which is harmless to cells and is efficient at tissue penetration, is then aimed at the entrapped nanoshells and their protein coats.
"Near-infrared wavelengths cause a really interesting thing to happen," Reich notes. "It causes the gold nanoshell to get excited and it causes whatever we've attached to come off." At the same time, nanobubbles form, causing openings in the endosome and allowing its protein contents to escape. The proteins are now free to home in on the cell's nucleus, where its genetic material is stored, and gain entry with the cell-penetrating peptide. And Cre can get to work finding, cutting, and pasting its reporter strands into the helix.