Section 1 – Summary
Study:
Scientists at The Scripps Research Institute (TSRI), in Florida, discovered a way to further optimize gene-editing technology. The scientists at TSRI were able to use CRISPR-Cpf1, and two types of RNA, to better target, cut and paste genes within human and animal cells (Zhong et al., 2017). With the help of “multiplexing” guide RNA’s, the CRISPR molecules would be led to a desired target, whether it be multiple genes or multiple locations within the same gene, to do its job (Zhong et al., 2017). Ultimately, the purpose of this advancement is not only to broaden the applications of the CRISPR-Cpf1 editing system, but instead it is also intended to be useful treating diseases. The uncertainty behind the Cpf1 molecule working on mammalian cells led to it first being tested on fireflies by editing their luminescence gene into the cell’s chromosome with the end result being successful (Zhong et al., 2017).
Section 2 – Key Concepts
The first genetic concept addressed in this study is the use of CRISPR, which stands for Clustered Regularly Interspace Short Palindromic Repeats. CRISPR is specifically chosen due to its function present in bacteria and archaea (Jinek et al., 2012). CRISPR has adaptive immunity against viruses. It has the ability to recognize viruses that it has encountered. Once recognized, the virus genetic information is cut up and disposed of (Zhong et al., 2017). CRISPR is typically attached to Cas9, however, in this study it was replaced by the Cpf1 molecule.
Guide RNA’s are also addressed in this study. The purpose of guide RNA’s is to map out where the CRISPR molecule should go in order to cut and edit genes or parts of a gene (Mefferd et al., 2017). This study uses RNA guided endonucleases Cpf1 and Cas9 in the beginning, but then focuses on Cpf1 due to its compatibility with mammalian cells.
Figure 1: How guide RNA’s works
Lastly, the guide RNA Cpf1 is specifically chosen due to its “multiplexing” capability (Zhong et al., 2017). Although Cpf1 and Cas9 are very similar, even in structure and function, Cpf1 contains the RuvC domain and putative novel nuclease domain (Yamano et al., 2016). Both domains enable Cpf1 to cleave both target and non-target strands, and it also generates a staggered DNA breaks (Yamano et al., 2016).
Figure 2: How the Cpf1 guide RNA works (Yamano et al., 2016)
Furthermore, Cpf1 is ideal because it can process its own CRISPR RNA (crRNA), unlike its counterpart Cas9 which is limited by its need to generate large constructs (Zetsche et al., 2016). Cpf1’s ability to process its own crRNA is what enables it to edit multiple parts of a gene or multiple gens at the same time.
Section 3 – Novelty
The novelty behind this study is connected to the behavior of CRISPR and Cpf1 combination. CRISPR gene editing on its own adopts a defense process present in bacteria and archaea. Microbes are able to fight of viral infection by retaining a piece of the virus’ foreign genetic material within its own DNA to serve as a template (Zhong et al, 2017). Since the virus’ genetic material is now within the microbe, it is able to recognize when the virus is present again, and it then is cut and disposed of with the help Cpf1. CRISPR and Cpf1 combined takes it one step further. Due to Cpf1’s ability to generate its own crRNA, this means that multiple genetic targets can be hit simultaneously; this improves the efficiency of editing multiple genes or multiple gene sites at the same time (Zhong et al., 2017). The prospect of this study is important when it comes to diseases that may be affected by multiple genes or multiple locations within the same gene. CRISPR-Cpf1 has revolutionized microbiology because it makes it more plausible for genetic engineering to become a treatment to combat diseases (Zhong et al., 2017).
Section 4 – Personal Reflection
I personally chose to write over this study because I was impressed by the versatility of the Cpf1 molecule, and how multiple molecules are able to simultaneously edit different areas of a gene or genes. This advancement further appealed to me due to my interest in the health profession and how it could heavily impact the health industry with continuous research. The scientist that participated in this study made claims that this advancement could combat diseases. As an example, they used the treatment of hepatitis B. If treated early enough using the studied method, it is possible for the viral DNA to be digested well before it causes irreversible damage to the liver and body (Zhong et al., 2017). I think it is exciting at the possibility of eradicating most, if not all, kinds of disease. Even if we’re not able to completely eliminate diseases, we would at least be able to use this studied method as a treatment to further minimize the effects of some diseases that have no ‘cure’. Aside from my excitement, I question whether the human body will eventually evolve/mutate and become immune to this method, and how long it would take for it to do so.
Section 5 – References
- Source of news: https://www.sciencedaily.com/releases/2017/06/170619120827.htm
- Jinek, M., Chylinski, K., Fonfara, I., Hauer, M., Doudna, J. A., & Charpentier, E. (2012, August 17). A Programmable Dual-RNA–Guided DNA Endonuclease in Adaptive Bacterial Immunity. Retrieved April 01, 2019, from http://science.sciencemag.org/content/337/6096/816
- Mefferd, A. L., Kornepati, A. V., Bogerd, H. P., & Kennedy, E. M. (2015, July 17). Expression of CRISPR/Cas single guide RNAs using small tRNA promoters. Retrieved April 01, 2019, from https://rnajournal.cshlp.org/content/21/9/1683.full
- Yamano, T., Nishimasu, H., Zetsche, B., Hirano, H., Slaymaker, I. M., Li, Y., . . . Nureki, O. (2016, April 21). Crystal Structure of Cpf1 in Complex with Guide RNA and Target DNA. Retrieved April 01, 2019, from https://www.sciencedirect.com/science/article/pii/S0092867416303944
- Zetsche, B., Heidenreich, M., Mohanraju, P., Fedorova, I., Kneppers, J., DeGennaro, E. M., . . . Zhang, F. (2016, December 05). Multiplex gene editing by CRISPR–Cpf1 using a single crRNA array. Retrieved April 01, 2019, from https://www.nature.com/articles/nbt.3737
- Zhong, G., Wang, H., Li, Y., Tran, M. H., & Farzan, M. (2017, June 19). Cpf1 proteins excise CRISPR RNAs from mRNA transcripts in mammalian cells. Retrieved April 1, 2019, from https://www.nature.com/articles/nchembio.2410
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