CRISPR/Cas9: The geneticist’s word processor

I’m old enough to remember the days of scribbling text on Big Chief Tablets. I typed final drafts on heavy-keyed typewriters that required strong fingers and a bottle of Wite-Out® nearby to rectify mistakes and edit copy. Then came word processors. Sentences glided past my eyes like fence posts along the Interstate. Editing became a relatively simple matter of cutting, pasting and deleting.

In 2012, geneticists acquired a similar tool, an RNA/protein combination, abbreviated CRISPR/Cas9, that seeks out targeted stretches of DNA to be edited and modified in a matter of days rather than the months that old techniques required. This wonder tool offers great promise, along with some potentially serious concerns.
A potential wonder tool

Geneticists’ eyes widen, contemplating what they can do with this tool:
• Fix genetic diseases
• Alter organs from pigs and other species so they can be used for human transplants
• Genetically engineer microbes as needed or desired for creating biofuels, eating waste materials, adapting to harsh environments, etc.
• Short circuit cancer growth and development
• Zap pesky viruses by slicing up their infective DNA or RNA
• Transform pluripotent stem cells into other kinds of cells like bone, muscle and liver
• Regulate normal gene function in new ways by creating the equivalent of stimulants, muzzles, dimmer switches and trackers for a host of genes and the metabolic pathways they control.

Colorado State University plant sciences Prof. Jan Leach is starting to use the CRISPR/Cas9 technology “to ask questions about how genes that help plants defend themselves against pathogens are regulated (turned on, turned up, or turned down) when the plant pathogens infect the plants.” Her plant of choice is Asian rice (Oryza sativa), the most widely consumed staple food crop for a large fraction of the world’s population.

Leach is also interested in the entire phytobiome surrounding crop plants: the complex networks of microbes that interact (in mostly positive ways) with plant root systems to maximize benefits for all soil inhabitants. Invasion by disease organisms usually occurs when these networks are disrupted or imbalanced in some way.
The fears and concerns

Powerful tools can create or destroy as well as have unintended consequences. CRISPR/Cas9 could potentially be used to edit germ cells or embryos, thus changing an organism’s entire genetic heritage. Some researchers are looking to impose regulations or some sort of moratorium so that the technology wouldn’t be used to create “designer babies.” Last April, Chinese researchers reported doing modifications to non-viable human embryos that stirred geneticists in other countries to call for a slowdown in this kind of work.

Others worry about an application of the technology called gene drives — genes engineered to break normal rules of inheritance so that changes get passed across generations. Gene drives have the potential of modifying mosquitoes, for example, in such a way that diseases like malaria could be eliminated entirely, but could rogue gene drives have unexpected consequences? Could bacteria or viruses, for example, pick up gene drives, and pass them around the biosphere in ways we can’t anticipate? How much do scientists need to worry about containing their creations to the lab?

In the case of Leach’s work at CSU, she notes, “We always work in contained (BL2P+) conditions. Our plants are contained in greenhouses or growth chambers, and we autoclave the plants when we are through with the experiments. Also, no rice is grown in Colorado, or for about 1,000 miles in any direction, so we are well isolated.”

Serendipity in science

In 2011, French geneticist Emmanuelle Charpentier approached Jennifer Doudna, an award-winning scientist from California, at a microbiology conference in Puerto Rico. Charpentier, a bubbly junior researcher, had become fascinated with a gene called CRISPR (short for “clustered regularly interspaced short palindromic repeats”) isolated from a flesh-eating bacterium. They talked shop about the then novel idea that bacteria might actually have pretty sophisticated immune systems. Charpentier had found that bacteria copy the DNA from ancient viral encounters and store it in their own genomes in an inactive form. When attacked by a viral repeat offender, the bacteria recognize the genetic sequence and use CRISPR in conjunction with the enzyme (organic catalyst) Cas9 to slice and dice its viral parasites into oblivion.

At the time, neither scientist thought about CRISPR/Cas9 as a gene-editing tool or even had any idea that the system could be lifted from bacteria and work successfully in other organisms. But the more they talked, the more excited they got.

Doudna said to Jennifer Kahn, a New York Times interviewer, regarding the CRISPR/Cas9 system, “I had this feeling. You know when you pick up a suspense novel, and read the first chapter, and you get a little chill, and you know, ‘Oh, this is going to be good’? It was like that.”

We all love good suspense novels and have faith that authors will lead us to happy endings. Scientists are now urging their colleagues to edit and police their studies carefully that make use of CRISPR/Cas9 technology so that the stories they ultimately write will have happy endings as well.

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