This post is cross-posted with the PLOS Student Blog
If you’ve recently taken a glimpse at the front page of any major science news outlet, it is likely you are no stranger to an emerging genome editing technology known as CRISPR/Cas9. With the help of RNA, Cas9 (a bacterial enzyme) can be programmed to target specific locations within the human genome, enabling scientists to delete, modify, or insert sequences that may treat, or even cure patients with genetic diseases. Although the CRISPR/Cas9 field is still in its early stages, major breakthroughs have been made recently, paving the road for a new line of gene therapy.
Just a couple weeks ago, researchers at Nanjing Medical University and Yunnan Key Laboratory reported successful usage of Cas9 in monkeys, thereby progressing toward the exciting possibility of editing human genomes. They injected Cas9 into monkey embryos and impregnated female monkeys with the resulting eggs; quite
remarkably, the newly born monkeys had deletions in the targeted gene of interest. In the world of science, where expected results sometimes never come to fruition, this marks an important stepping-stone in Cas9 technology. Targeting genes with this high degree of specificity could potentially lead to therapies that will prevent individuals from developing genetic diseases.
In the same week, a paper published in Nature revealed the mechanism by which Cas9 finds target DNA sequences tens of base-pairs in size within a genome that contains three billion base pairs. Interestingly, the researchers showed that Cas9 searches for a specific sequence known as the PAM – if the target doesn’t carry this short DNA tag, the sequence is neither recognized nor cut. Samuel Sternberg, lead author on the paper, explains that the presence of PAM sequences “accelerates the rate at which the target can be located, and minimizes the time spent interrogating non-target DNA sites.”
One of the most recent discoveries came out last week in Science magazine: two structural biologists at UC Berkeley, Jennifer Doudna and Eva Nogales, published the structure of Cas9 from two different organisms, providing key insights into the mode of DNA recognition and cleavage by the RNA-guided enzyme. Fuguo Jiang, one of the lead authors on the paper, said, “although the two Cas9s are from different organisms, and overall they look very different, when you superimpose the two structures, their functional domains are very similar. This suggests all the Cas9s have a similar mechanism to make cuts in DNA.” Additionally, the paper revealed an important loop within Cas9 is responsible for recognizing the PAM. As for what this means for the future of CRISPR/Cas9 technology, Jiang elaborated, “The original Cas9 recognizes one particular PAM, but if you can engineer it to recognize a different PAM sequence through mutagenesis, that would be great. The genome sequence is usually fixed. But what you could do is change the PAM sequences that Cas9 recognizes and tailor it to target more genomic loci. In this way, we can expand our Cas9-based genome-editing toolbox.” This discovery, along with the physical mechanism by which Cas9 locates target sequences, may help improve the efficiency of targeted gene editing.
And that’s not all. Editas Medicine, a new company co-founded by some of the leading scientists studying Cas9 aim to translate its genome engineering technology into a novel class of human therapeutics. These therapies are destined to make significant medical advances for people with genetic diseases including, but not limited to, Huntington’s disease, cystic fibrosis, and Alzheimer’s. Since modern sequencing technology has produced a massive amount of human genome sequences, mapping diseases to certain genomic coordinates is becoming faster and easier. With this valuable sequence information, the CRISPR/Cas9 system can simply be engineered to make positive changes in specific diseased DNA sequences and restore normal function.
Genome engineering earned researchers a Nobel Prize in 2007, but with Cas9 speeding ahead, I wouldn’t be surprised if one is awarded to a Cas9-er in the near future.
**While writing this article, a paper was published in Cell that reveals another structure of Cas9, but now bound to its target DNA. This structure provides more information about the molecular mechanisms by which Cas9 cuts its targets and will further aid researchers in improving genome-editing tools**
If you want to learn more about how Cas9 functions, check out this video produced by a student in Eric Greene’s lab at Columbia University: