As part of our series of multiple perspectives on gene editing, we asked Joe Zagorski from the Michigan State University Institute of Integrative Toxicology how CRISPR has impacted his research.
CRISPR is here to stay.
There are few topics that garnish such diametrically opposed stances as genetic modifications of organisms, whether it be for gene therapy or development of disease resistant crops. Over the last 4 years, I have been doing just that. In a couple of months, I will be graduating with a PhD from Michigan State University, in Cell and Molecular Biology. During my time at MSU, I researched how a stress activated protein called Nrf2 controls human T cell (a type of cell in your immune system) function, and creating genetic modifications was a major component of my graduate work. The tool I used to achieve my goal was, at the time, a ground-breaking technique called CRISPR. Although I utilized CRISPR for basic science, it has extremely broad applications and is currently being used to generate genetically modified food crops, including corn, rice, and wheat, as well as being used as a potential method for the treatment of some diseases, such as cancer. This technique is not going away, as it has revolutionized all aspects of gene manipulation. Since we will be hearing about and dealing with CRISPR for the foreseeable future, it is important to understand what CRISPR is, what it can do, and why scientists are so excited about it.
What is CRISPR and how did it get started?
CRISPR stands for Clustered Regularly Interspaced Short Palendromic Repeats and was originally discovered as a very crude immune system in some bacteria, and was first described in 1987 by a research group in Japan. Bacteria, just like you and me, are at risk of being infected, and potentially killed, by viruses. Viruses that infect bacteria are called bacteriophages, but you will frequently see this shortened to phage. Some types of bacteria have the ability to incorporate small pieces of the phage’s own DNA into their genome. Then, if the bacterium is infected by the same phage a second time, it has memory, of sorts, of the initial infection. The bacteria can then use that memory to specifically target the phage for destruction. This is frequently accomplished by a protein called CRISPR associated protein 9 (Cas9). This protein is guided to the targeted DNA sequence and cuts it – resulting in its disruption.
The ability to specifically cut DNA is what has garnished the attention of basic, translational, and industrial scientists. In 2012, CRISPR was demonstrated to have the ability to modify specific DNA sequences designed by scientists. At this point, the CRISPR revolution began. Less than one year later, two different groups, one from Harvard and one from MIT, would publish manuscripts showing the ability of this gene editing technique to disrupt genes in both humans and mice.
One thing to remember, however, is that we have been using molecular techniques to edit genes for well over a decade. So, what makes CRISPR so different and so exciting? I think there are really three main reasons CRISPR is so enticing: price, speed, and flexibility.
Overcoming the price tag of genetic research
The reality is, that whether it be industry or academia, price is often one of the biggest hurdles to overcome in research. When I first started looking at gene modification techniques in my dissertation research (in the fall of 2012), I considered using zinc-finger endonucleases and TAL Effector Nucleaes. Both techniques are similar to CRISPR in that they rely on very specific cuts in the cell’s DNA, resulting in the disruption (or knock-out) of a gene. The problem with these techniques is that they are much harder to develop, on top of being expensive. For my own research, these techniques were going to cost $2,000-$8,000 to accomplish. In the end, I would use CRISPR-Cas9 technology, and it cost my lab about $70.
Speeding up the research process
Although the raw materials are substantially cheaper using this technique, cost and speed are intimately intertwined, as one often affects the other. In my research, I use what is termed a cell line. A cell line is similar to cells in your own body, but they have been changed to survive longer outside of the host. Sometimes, researchers induce this change from a normal cell to a cell line, while in other instances, they simply use donated cancerous cells. In the case of the T cells I study (Jurkat E6-1 cells), they are a type of T cell leukemia, which allows them to be almost immortal. Currently, using Jurkat cells, I can take this process (i.e., from non-disrupted gene to disrupted gene) in about a month. This timeline becomes even more impressive when you look at the amount of time it takes to make different animals with desired mutations (termed transgenics). Using previous techniques, such as selective breeding, making a transgenic mouse would take about 18 months and cost, at times, in excess of $20,000. Now, using CRISPR, this can be accomplished in approximately one month and cost less than $5,000. This increase in speed allows for more scenarios to be studied, as the hurdles of both time and money have been lowered. This means that more potential drug targets can be studied, more potential disease resistant plants can be generated, and more gene therapies can be pursued. This diverse use, however, requires something be customizable and flexible and not just have the ability to ablate genes of interest.
Increasing the flexibility of research design
There is good reason that scientists are so excited about this technique. Since the beginning of the CRISPR revolution, scientists have gotten quite creative with finding more uses for CRISPR, which has made this powerful technique incredibly flexible. Now, its use isn’t limited to just gene disruption. For instance, CRISPR can be used to introduce new genes, to turn genes on, to turn genes down (but not off), and to make subtle changes (polymorphisms) within genes. These numerous variations to CRISPR technology have resulted in its application in varying fields, from treating diseases like HIV and cancer, to basic science, and even into the development of genetically modified foods and food products.
CRISPR had primitive beginnings in microorganisms, including archaea and bacteria. But because of the innovation of some scientists, it is no longer only a primitive immune system for bacteria. We now have the ability to quickly, easily, and inexpensively alter the function of genes in plants and animals alike, in a very specific manner, allowing potentially new solutions to many difficult challenges in science and health. As populations increase, climates change, and diseases arise, science needs to adapt to the present demands in order to solve these problems sustainably.
CRISPR is one way we can adapt to address these challenges.
Joe Zagorski is an immunotoxicologist and is working to better understand how exposure of to compounds in your environment (i.e. what you eat, drink, breathe in, etc.) change the way your immune system functions, specifically in regards to a stress activated protein called Nrf2. Since a young age, Joe wanted to be a scientist. You can read a personal description of his journey from Bill Nye admirer to scientist on the blog, Civilians of Science.
This article is the first in a series of multiple perspectives on the topic of gene editing. Once additional perspectives are published, we’ll post them here.