One of the most significant recent scientific discoveries was that of CRISPR, a bacterial immune system mechanism. CRISPR stands for “clustered regularly interspaced short palindromic repeats.” It may have a complex name, but the mechanism itself only involves 2 components: an enzyme that cuts DNA and a “guide” RNA strand. The CRISPR sequences themselves are naturally contained in bacterial DNA, and they are copies of the genomes of viruses who have at one time infiltrated a specific bacterium. If one of the viruses, whose genome is already contained in a bacterium’s CRISPR sequences, happens to infect the bacterium once again, an RNA strand complementary to the CRISPR sequence that corresponds to the specific virus will be transcribed and then attached to an enzyme, such as Cas9 (Cpf1 is also sometimes used). Cas9 is an enzyme that can cut out DNA strands complementary to the guide RNA it is attached to. In the case of viral destruction, Cas9 will cut out a viral genomic sequence based on its guide RNA, which was transcribed from the specific CRISPR sequence that matches the virus’ genome. For a new virus, the viral genome will be cut into fragments by various enzymes, and some of the remnants will then be incorporated into the bacterial DNA (after it’s cut by other enzymes) as new CRISPR sequences so that that virus can be destroyed by specific CRISPR-complementary guide RNA and Cas9, if virus ever returns.
CRISPR has been harnessed as a new gene editing technology to help answer the 3 essential questions that biology has been consumed with for the past century:
- “What does each gene do?”,
- “How do we find the genetic mutations that make us sick?”, and
- “How can we overcome them?”
Cas9 and other similar enzymes can be outfitted with artificial guide RNA sequences that are complementary to DNA sequences in specific genes. Cas9 will then cut out that DNA from the gene. The DNA will immediately attempt to reconnect, and in doing so, it will incorporate any surrounding DNA fragments or nucleotides into the final, reconnected DNA. The newly incorporated DNA will also be copied through homology-directed repair into the homologous chromosome of the chromosome on which the CRISPR editing took place. So, geneticists can insert new genes into a cell that they want to incorporate into its genome, and after Cas9 cuts out a specific old gene, the new gene will be incorporated during the DNA’s healing process. This gene editing/replacement system has already been used successfully in mice and wheat. In mice, CRISPR technology has been able to successfully replace the alleles that cause sickle-cell anemia, muscular dystrophy, cystic fibrosis, cataracts, and more. Also, a specific gene in wheat has been successfully deleted through CRISPR, resulting in wheat that is resistant to mildew.
The gene-editing uses of CRISPR may also offer more effective treatments or even cures for various genetic diseases, specifically cancer. Cancer is different from other diseases, especially infectious diseases, mainly because each new case of cancer has had no experience with previous cancer treatments and thus cannot be resistant to any of them, as it has not learned from the experiences of past cancer cases. So, the same cancer treatments can be used over and over again without fear of subsequent resistant cancer cases. However, infectious diseases are caused by living organisms or viruses that can develop beneficial genetic mutations over time that eventually lead to completely resistant new generations. This means that they “learn” from the experiences of their ancestors; therefore, the treatments used to destroy them must be constantly updated to keep up with the rate at which the bacteria, viruses, etc. develop resistance. But, cancer is still extremely difficult to treat due to the uniqueness of the genetic mutations that cause each case of cancer. CRISPR could be used to treat some cancers and other genetic diseases if the exact mutated genes that cause them are found. Complementary guide RNA sequences to sections of these genes could be attached to Cas9 enzymes, which would then cut out the mutated genes and allow for replacement with healthy genes.
So, when and how was CRISPR first identified? In 1987, a group of Japanese scientists sequenced a gene from a E. coli bacteria. As part of their examination of the gene, they sequenced some of the surrounding DNA. These surrounding DNA sequences happened to be CRISPR sequences, which had not been observed until then. The scientists therefore had no knowledge of the sequences before them and did not know what to make of them. However, in 2005, a microbiologist from the University of Alicante made an astonishing comparison: the unknown E. coli sequences were identical to sections of viral genomes! He had compared the E. coli sequences to sequences from hundreds of other organisms but found the most similarity in viral genomes. It was then inferred that these sequences had originated from viruses that had at one time infected the bacteria.
A few other potential uses for CRISPR include the eradication of malaria, AIDS, and Lyme disease. To eradicate these diseases, gene drives must be utilized along with CRISPR technology. Gene drives increase the chance of a specific parental allele being inherited by offspring, guaranteeing that all offspring will carry that allele and that eventually, nearly the entire population will carry that allele. Mendelian genetics state that offspring have an equal chance of inheriting each allele of a particular gene. Thus, gene drives conflict with Mendelian genetics by greatly increasing a certain allele’s chances of being inherited. Gene drives work hand in hand with CRISPR; as mentioned earlier, a gene inserted using CRISPR will be copied through homology directed repair into the homologous chromosome of the chromosome in which the new gene has been inserted. This means that that organism will have 2 copies of that new gene and a 100% chance of passing the gene down to all offspring. Therefore, the gene will quickly accumulate in the population until every member of the population carries the gene. Most of the time, the genes that are used in gene drives are “selfish genes”: genes that exterminate or change species that threaten humans.
For example, a potential gene drive has been proposed that involves a so-called “selfish gene” and could eradicate malaria. The gene would prevent female A. gambiae mosquitoes (which carry the parasite that causes malaria) from producing fertile eggs, eventually leading to the disappearance of the species. This gene is considered “selfish”, as it will eradicate a species that is dangerous to humans. However, completely eliminating mosquitoes from the Earth might have some ecological consequences. Many animals use mosquitoes as an important food source, so if they were to be exterminated, these animals might starve and die off as well. Essentially, it would disrupt the food chain.
Another possible gene drive would guarantee the inheritance of the Delta32 mutation of the CCR5 gene by all human offspring. This would prevent all humans from contracting HIV, since HIV enters human blood cells through CCR5 receptors, and the Delta32 mutation produces faulty CCR5 receptors through which HIV cannot travel. However, functional CCR5 receptors are needed to transport leukocytes, which fight against viruses, into cells infected with the West Nile virus. Without them, humans would be much more susceptible to the West Nile virus, and a higher percentage of those infected would not survive. The debate over this gene drive comes down to this question: “Which is more important: protection against HIV or protection against the West Nile virus?”
Lastly, another gene drive is being developed with the goal of eradicating Lyme disease. This gene drive was developed by a researcher named Kevin Esvelt, and he has proposed using CRISPR to introduce a gene into white-footed mice that produces an antibody which protects against and destroys Lyme bacteria. White-footed mice are the main source of Lyme disease (they then transmit the Lyme bacteria to ticks, who transmit it to humans), so if the mice could no longer carry the Lyme bacteria, neither could any other organism. Esvelt’s goals with this gene drive have been described as those of an “effective altruist.” This means he is completely devoted to helping others (in this case, those infected with Lyme disease), and he is actually able to effectively aid them.
CRISPR could also be used as a tool to guide evolution, as it could introduce beneficial genes or alleles to a population that are then selected for by natural selection. This would cause changes in gene or allele frequencies in the population, and these changes are the definition of evolution. Thus, CRISPR could guide and influence evolution.
As a final note about this fascinating technology, CRISPR can only reveal the function of one gene at a time. If 2 or more genes are added/removed at once, their combined effects could be different from the the list of effects created from adding/removing each individual gene, since the whole is different from the sum of the parts. The same concept applies to species ecology, or the interactions between members of a species and their environment. Individual organisms interact with their environment in unique ways by themselves, but when they congregate, they exhibit different, more uniform behaviors. Therefore, the behaviors of an entire, assembled species are different from the list of each of the members’ individual behaviors.
2019-2-20-1550639184