15 years ago, scientists finally concluded that CRISPR (Clustered Regularly Interspaced Short Palindromic Repeat), together with CRISPR-associated proteins (Cas) play a key role as a safeguard against bacteriophages. After this discovery it took only a couple of years for Emmanuelle Charpentier and Jennifer Doudnathe to develop precise genome-editing technology - CRISPR - Cas9. This gene-editing tool was one of the greatest breakthroughs of the 2010s, it allowed us to experiment with gene-edited mosquitos to reduce the spread of malaria, for engineering agriculture to withstand climate change, and in human clinical trials to treat a range of diseases, but how did it become so influential and what lead to its implementation?
The CRISPR sequences were initially discovered in the E. coli genome in 1987. Even though scientists hypothesized that prokaryotes used CRISPR as part of an adaptive immune system, it was not until 21. century that this function and its importance was confirmed. This system evolved to protect bacterial cells from its most common parasites - bacteriophages. When viral infection occurs this system cuts viral DNA into small fragments (around 20 nucleotides long). Then, it stores some of these viral fragments (called spacers) in a specific part of the bacterial genome - in between CRISPR sequences. Next time the same virus tries to attack - the CRISPR defense system will recognize the sequence that was previously stored in the spacers and immediately destroy viral DNA. This is possible due to two key components of this system. First is CRISPR RNA formed by transcription of spacers regions (transcription is a process of copying DNA sequence into RNA sequence). This RNA serves as a guide for molecular machinery to locate viral DNA in the cell. The second is a protein that is part of molecular machinery called Cas 9. This protein forms a complex with CRISPR RNA and then cuts viral DNA at a very precise spot that is identified by a sequence of CRISPR RNA.
First step in the adaptation of this technique is designing guide RNAs that will be specific for the place in genome that we want to edit. Then Cas protein will cut cell DNA at the desired spot. After that, we can use the cell's own DNA repair machinery to add or delete pieces of genetic material, or to make changes to the DNA by replacing an existing segment with a customized DNA sequence. Besides that, there are multiple upgrades of Cas proteins that allow us to activate or repress certain genes or even modify RNA molecules.
This was one of the first applications of this system because it is most similar to its original purpose - the only difference being the source of the spacers - in this case using artificial ones instead of real viruses.
Even though GMO in agriculture existed long before the development of CRISPR, this system enabled this process to become cheap and easy, thus significantly speeding up production.
There are more than 300000 research published that are somehow related to the usage of CRISPR technology. From reverse genetics, knock out and knock down mutants, epigenetics modifiers to the addition of fluorescent tags are all just a part of the techniques that were hard, expensive, or almost impossible before CRISPR.
CRISPR has a great potential for treating genetic diseases and cancer. From blood diseases such as sickle cell disease (SCD) and beta-thalassemiaa to neurodegenerative diseases, like transthyretin amyloidosis, CRISPR showed promising results in clinical trials. However, there is still ethical debate about certain usage of CRISPR on humans.
Genome engineers continue to work to develop a highly specific, programmable platform well-suited for various biological and translational technologies. It will also be important to find a way to deliver CRISPR therapies into the body before they can become widely used in medicine, with the caveat that off-target editing risk must be made very low or nonexistent. Scientists continue to wrestle with the possibility of postnatal and germline editing, but there is still a huge ethical debate in front of us before actual usage.
CRISPR is a technology that can be used to edit genes and, as such, will likely change the world. CRISPR is already widely used for scientific research, but it made its way in the industry and also has the potential to transform medicine, enabling us to not only treat but also prevent many diseases. This powerful tool speed up many technological innovations, thus brought us closer to the future. However, it opens other ethical questions about using it to change the genomes of our children.