Dr. Hong Li, professor in Van Andel Institute’s Department of Structural Biology, leverages CRISPR, cryo-EM and other leading-edge technologies to explore the mechanisms underlying RNA-mediated processes. Her research has far-reaching implications for informing new diagnostic and treatment strategies for cancer and other disorders.
During a recent public lecture, Dr. Li provided an overview of gene editing methods and described how this technology is supporting new approaches to studying disease.
Watch the lecture below:
Video transcript
Dr. Mary Winn [0:00]
I’m truly excited to dive into the latest research today. Every year, we have countless new technologies and tools that have the potential to change health and humanity for the better. Many of them seem straight out of fantastical TV shows and movies giving us hope and fueling our dreams of the future. Today we’re going to talk about a topic science fiction fans may be familiar with: gene editing. Thanks to technological advances, gene editing has become a powerful real world tool for studying and potentially treating disease. Today we’ll be joined by VAI’s Dr. Hong Li, an expert in CRISPR-Cas gene editing technology for a crash course on gene editing and how it is changing the game in human research. I’ve had the pleasure of working with and supporting Dr. Li since she joined VAI in the summer of 2024, and I’m delighted for you to hear more about our work today. She leverages CRISPR cryo-electron microscopy and other leading edge technologies to explore the mechanisms underlying RNA mediated processes. She has built a multifaceted research program that has informed new strategies for gene editing, cancer diagnosis, and virus detection. We’ll have time for questions after Dr. Li’s presentation, and we encourage you to post your questions in the chat. In the meantime, please join me in welcoming Dr. Li.
Dr. Hong Li [1:31]
Thank you Mary, uh, for that introduction. Um, I welcome everyone, uh, to the public lecture on gene editing. Uh, so this, uh, talk, uh, will be divided into four parts. If I can advance the next slide, um, then I’ll pro provide an introduction, um, to the, to the talk and leading to the CRISPR technology. And I’ll describe what is the CRISPR technology. Um, and then I will provide some examples of what we study, uh, here in VAI and I finally provide topics and for discussion. And so the first, uh, I wanted to introduce myself a little bit. Uh, next slide that I moved to here, as Mary said, uh, last year, uh, to VAI. Um, most of our studies, uh, is discovering, uh, as well as characterizing and repurposing CRISPR technology as well as related technology for both fundamental biology studies as well as a potential therapeutic use.
So, um, prior to joining VAI was a professor of chemistry and biochemistry at Florida State University. Uh, and, uh, uh, when I was, uh, and, and as a graduate student actually in University of Rochester, studying at the interface of physics and biology, um, that, uh, inspired me to, uh, the research that I’m carrying out today. Uh, while reading the book of the Double Helix by James Watson and, uh, James Watson and Francis Crick, uh, used a technology called X-ray diffraction, shown here, uh, as, uh, taken by scientist Rosalind Franklin, uh, of a, of what we call a diffraction of this mysterious, uh, DNA molecule that led them to the elucidation of the three dimensional shape of this, uh, DNA molecule, um, that really started this field of molecular biology. And, uh, started an exciting journey of understanding the DNA and, uh, and led the foundation to today’s genome editing.
And I was fortunate enough at Florida State and, uh, become the colleague of Professor, uh, Don Casper, uh, who was a dear friend of Rosalind Franklin, uh, back, uh, in Cambridge, England. And my post-doctor training was in Caltech, uh, where under the guidance of Professor John Abelson, who introduced to me this, uh, uh, equally exciting molecule called RNA, um, that is also a foundation to the discovery and use of, uh, CRISPR technology. So if the mid 20th century was a DNA revolution and the discovery of CRISPR technology, uh, would be one of the RNA revolution. So, uh, next slide. Um, many of you might know that the DNA is actually the molecule of heredity, and that it gave rise to, uh, the living organism instructions as well as inheritance. But many of you may not know that the discovery was made by, uh, studying, uh, this simple system phage, uh, how it infect bacteria.
And by, uh, uh, mostly three scientists, Max Delbrück, um, Alfred Hershey and Martha, Martha Chase. And then they use this simple manipulable, um, viruses phage, uh, to really, uh, firm the discovery that the DNA is the inherit, inherit, uh, heredity molecule. And phages are the viruses that infect the bacteria, much like the flu viruses that infect the humans. And so, uh, and this discovery set the stage of the, you know, decades of research on what the DNA, how does it give instruction to the cell, and how does it, when it’s, uh, uh, going wrong, uh, cause diseases. Um, so the, the, um, next slides please.
So, uh, one of the, uh, first, uh, study was to what is actually DNA molecules made of, and then they are made of, of what we call the building blocks nucleotides that’s sticking together by, uh, strain, uh, by this, uh, what’s called a phosphosugar backbone. And these nucleotides, it has complementarity to the opposing strand of the DNA much like the zippers when they’re trying to be zipping together. So they always come in pairs. Um, and so, uh, next slide please. And in simple organisms, uh, such as bacteria, uh, the DNA molecule, uh, the genome, or we call the genome that comprise the DNA molecule are pretty simple. They’re usually circular, uh, double stranded circular DNA molecules avoid of free of all these higher order structures. But in animal and plants, uh, the DNA molecules are double stranded, but wrapping around, um, protein molecules that they call the histones and to form these higher ordered structures. Uh, so in addition, the genome has a lot of diversity. And, for instance, in higher eukaryotes such as animal and plants, uh, they are actually have additional chemical modifications on the DNA as well as the histones. And we call these epigenetic modifications in addition to the, the, the sequences of these nucleotides and giving instructions to the cell, these epigenetic modifications, as well as DNA modification that further alters the signal of the genome. And that can cause different biology as well as when it goes wrong, cause diseases.
Um, so to direct the transfer the information, genetic information that the DNA usually have to make, uh, another molecule, according to the nucleotide sequences called RNA, uh, that they, some of these RNA molecules actually give the instructions to the cell to make the molecules called proteins. And this principle is called a central dogma. Uh, that’s true for living organisms that the protein molecules, obviously what we call the workers of the cell, they make up our tissues, they make up skins, bones, and, uh, just the structure as well as acting as enzyme, for instance, for digesting food. So therefore, the instruction given by the DNA can, uh, direct what is the, uh, property of these proteins. And so therefore, um, if there is a change in the DNA sequences, you could change the RNA and the protein, which give a rise to different traits of the organism.
For instance, a different co, color of a plant. So much like, uh, one changes the letters in the sentence that will give rise to a different meaning. And so, given the importance of the, what we call the, the, the information that’s encoded in the DNA and scientists always wanted to, to say that we wish there is a way we could rewrite the code of life, meaning we can change the code of the DNA, um, that will enable us to, first of all, to study the fundamental biology. If you can alter the DNA sequences, that’s, uh, the blueprint of the organism. And you could also use the change the DNA to correct what we call the, the mutations in the patient genome. And you could also, in fact, to remove, uh, certain genes, certain stretch of sequences, or even add certain stretch of sequences that can change the desire, the traits of the organism.
Uh, you can also, uh, maybe influence the in genetic information without even changing the sequence and such as these epigenetic, uh, modifications of, of the genome. So, despite that, this has been desired for many years prior to, to the discovery of this new technology called a CRISPR. Uh, there was just no good, uh, safe and convenient simple methods for doing so. So, uh, we lack the such, um, technology. So, uh, excitingly, uh, it is in around 2012, uh, this, uh, enzyme called a CRISPR uh, technology was born. Uh, I’m showing this phage and bacteria system again. And so the study of the phage and the bacteria, uh, the, you know, virus infecting the bacteria led to the discovery of the CRISPR. So, in fact, the sort of first landmark study was made, uh, by study in the yogurt, uh, the, the bacteria that may in our yogurt, uh, culture, our yogurt, that, uh, the, the company who studied this, uh, bacteria, uh, wanted to make sure the yogurt is not infected by, by the viruses.
And then they discovered that there’s existing, um, a mechanism in these bacterias that they can fight the in infecting phages. And this mechanism is called a CRISPR. And then the CRISPR is a stretch of a DNA sequences in many, many bacterias that they encode, uh, a series of CRISPR enzymes. So these are, uh, the enzymes that could upon the phage infection to the bacteria, they can, um, restrict, they can, uh, kill the, these viruses so that they can survive, much like the human, uh, antibodies that we develop that allow us to fight the viruses, you know, flu viruses or, or CoV-2 viruses. So it is really, the CRISPR enzymes is native to bacteria. They really, um, are used these enzymes to, to fight the infection of their own infection. Um, and this is the, uh, enzyme that the scientists studied to the detail and the discovery that this enzyme could be taken out of the bacteria and use it for, uh, genome editing.
And so, uh, why is this, uh, enzyme, uh, are able to do that? And so there are many CRISPR enzymes among which, uh, there are two that’s are mostly used in genome editing. And so we gave them names of a cas CRISPR associated proteins, Cas9, Cas1, Cas2, all the way to, you know, Cas, uh, 14 Now. So, uh, these two enzymes, Cas9 and Cas12 are enzymes the bacteria uses to cut virus DNA. And then, so that comes back to why we can use the CRISPR to alter the DNA sequences in, in genome, especially the Cas9, which is the most, um, widely used, uh, genome editing enzyme. Uh, so these blocks really shows the shape, uh, in a really cartoon, uh, representation of how the, uh, enzymes are made up, so they are made of several parts. Uh, one of the, uh, parts are the ones that’s holding the DNA as you will see later.
Uh, but these, uh, green and and blue, uh, parts are representing what we called the, uh, active sides of the enzyme. So these are the, uh, sites where they actually can break, uh, DNA phosphosugar backbone. So, as I introduced earlier this molecule called RNA molecule, that many of these RNA molecules that’s copied, made by the DNA, they do not, uh, encode the proteins. And these are called non-coding RNAs. So, CRISPR enzymes actually widely use these non-coding RNAs that these, they, they stopped being as the RNA as a functional molecule themself. So they work with the protein molecules, uh, in this case, they, they could work with Cas9 molecules. So they would, um, uh, partner, uh, with the Cas9. So obviously, the significance of the discovery, this Cas9 was the 2020, uh, uh, Nobel Prize by, uh, in chemistry, by, uh, giving to Emmanuelle Charpentier and Jennifer A. Doudna, um, for its potential in, in many, many research as well as therapeutics.
So, uh, RNA molecules that’s made also by the CRISPR, um, locus in bacteria. Uh, it has this remarkable ability, uh, property. First is part of this RNA that’s gonna partner with these Cas9 Cas12 protein enzymes came actually from virus themself. So they remember that inflammation of the unique code that in the phage molecule that previously infected them. By bearing this information of this DNA, uh, of, of the sequences that in the, in the phage, uh, it could allow, uh, this RNA to recognize the phage in a subsequent infection. Uh, so that’s, that’s the first remarkable, um, property. And so in phage, if the, if the be, uh, Cas9 molecule is were to cut the DNA from the phage, it would, uh, unzip the phage DNA, which are double stranded, but instead, replace one of the DNA strands, uh, of the phage by the RNA that it remembered, and that now become part of the Cas9 enzyme.
And that allows this Cas9 to be properly sitting on the phage DNA and make, um, break the chemical bond of the phage DNA, allow it to render it, uh, nonfunctional. And what’s remarkable about this, RNA, we also call them a guide, RNA, because they sort of take the Cas9 enzyme to where the phage DNA is. And so it knows not to cut its own DNA that will be devastating. And then, so we call them a guide RNA. What’s remarkable of the guide RNA of the Cas9, as well as Cas12, is you can change it, uh, when you wanna use for a different purpose. That’s, uh, made this Cas9 technology, what we call programmable. Meaning, if you’re, uh, synthetically making this guide RNA for Cas9 enzyme, but dial the sequence of the guide region of the guide, RNA to a sequence, let’s say, match, uh, part of the, uh, uh, gene of a in the human genome.
And you would be able to take the Cas9 or Cas12 enzyme to the human, uh, gene, uh, in the genome. And that’s what makes this, uh, technology powerful, uh, and that it’s such a simple, uh, technology that allows one, uh, to, to realize. All right? And so, um, the, obviously the realizing that this is the, uh, the, the potential for the Cas9 and Cas12 to be used in, in biotechnology, uh, and that set the stage for the why, the spread application of, of this enzyme. All right. So there are, uh, different ways one can use this. RNA guided double stranded, DNA break enzyme Cas9, as well as Cas12. So the very first mode of the use of the CRISPR technology, uh, would it be to direct the Cas9 enzyme to, uh, a part of the genome, uh, using a guide RNA, uh, that you dial it to, and you can make a double stranded break.
So the double stranded break, uh, we call it cutting that triggers in the human cell and called DNA repair. So the human cell has the machineries that the minute your DNA is broken, it’ll try to stitch them together. And so that’s called double stranded, uh, uh, break repair. So the way the cells repair our genome, there are two ways to repair it. One way is just to stitch them together as best that it can. But the way when it is stitched together, uh, usually will change the alter, the original sequence of the DNA, therefore, um, you know, eliminating the original message that’s encoded in the DNA. So in that case, uh, one would be able to destroy that message. So, um, they are also another way of repair in the human, uh, genome, which is to supply a new DNA and that we called a template DNA in conjunction with double stranded break, with a new DNA template, one can actually repair it according to what the new sequence encoded in a template will be, and that will allow us to alter, for instance, the disease mutations, uh, and unwanted changes that’s happening in the genome.
So this is the very first rudimentary, uh, direct application of the CRISPR Cas9. However, uh, because of the programmability of the CRISPR-Cas9 many, uh, uh, derived technology has been discovered. Uh, these are the technology that does not involve double stranded break. Some of them just breaks one, we call it “nick” on the DNA. So these CRISPR-Cas9 mediated, uh, technology, they are usually a fusion of the enzyme. So it’s like a, taking a CRISPR-Cas9 attaching to another tool to, to the Cas9. So, so that you can apply additional, uh, changes in the DNA. For instance, they are technology called base editors. So that means not to make a cut in the DNA at the location that guide RNA led you to, but rather to change, uh, the nucleotide to another nucleotide to swap the nucleotide sequence.
This is also powerful, but much safer way to do that. Uh, one can also use it to attach to other enzymes to make the CRISPR, such as what we call a prime editing. And that can actually insert a stretch of A DNA, um, although short, uh, piece of DNA at the site when targeted too. Um, you can also, not cutting anything on A DNA, but take the good CRISPR-Cas9 to the genome of the location that you can turn what we call silencing certain genes, or you can activate certain genes, um, in other words, interfere with the RNA production and or the, you know, subsequent protein production. So, uh, realize the goal of, uh, you know, uh, program, the changing of the, of, of the downstream product, of the gene product. And one, uh, technology has also made it possible, for instance, to guide the CRISPR-Cas9 to the genome of the interest to add these chemical modifications or to erase these chemical modifications, um, which in turn, uh, can turn gene on and off.
So many, uh, CRISPR, uh, derived technology that are equally exciting and advancing their applications. Uh, interestingly, uh, so far, all the CRISPR Cas9 or Cas12, uh, are not, they’re, they are not able to sense the chemical modifications on the genome. And so, despite the fact, we know the fact, which is one of the focus, the study area of Van Andel Institute, that the DNA methylation are incredibly important in fundamental biology as well as in the development of diseases. But none of these CRISPR current tech CRISPR technology, uh, that is not able to sense the chemical mod, uh, the chemical modifications on the DNA, so currently lacking, uh, the, these, uh, set of technology. One thing, uh, scientists are very cautious about is one mode of the CRISPR, uh, function, uh, that’s called off-target cutting, which means that there are homologous regions of the genome, uh, that’s to the region, you programmed it, uh, to, and, uh, there is a very, very, very small chance, but not zero.
And that the CRISPR-Cas12, uh, 9 and 12, and would be able to make, uh, break, and this is what we called off-target cutting. And obviously, it’s, uh, undesired, and it, it is the some, uh, activity one, um, the scientists are paying a lot of, uh, close attention to and to try to eliminate them and to make, uh, the technology more precise. And so, one of our research goals, for instance, is to, uh, to, to achieve that by utilizing, uh, the methylation, uh, markers on the, on the DNA, um, because of this, um, technology, I, I, I, I believe it’s still young. It’s about, uh, 13 years ago, and that’s started this, uh, discovery. And it’s moving very, very fast because of the power of the system, uh, into therapeutics. And so, uh, one of this, uh, CRISPR technology has already been proved by FDI, uh, FDA.
And that is the, uh, CRISPR, um, technology that’s, uh, used in sickle cell disease, uh, uh, therapy, that it allows it to the sickle cell patient to reactivate, uh, their fetal hemoglobin, which is the molecule that carries the oxygen around the body, uh, whereas to, you know, bypass their defective and adult hemoglobin. So, uh, in addition to, uh, this, uh, uh, FDA approved technology, uh, there are many more. And then it’s currently in clinical trials in different, uh, phases of the clinical trial and, and look, um, very promising to be in, uh, future therapeutics. And thank for the, uh, the Innovative Genome Institute and University of California Berkeley, uh, who keep close, uh, uh, close, um, updates on, uh, this information. Uh, so despite they are, uh, you, you know, these technology have to go through all these, uh, um, uh, FDA approval processes, and they, you always hear, um, many of these, what’s called a CRISPR on demand, uh, personalize the medicine.
And it is very exciting. Uh, for instance, uh, this example, uh, just recently reported, uh, on, uh, this, uh, baby, uh, uh, named KJ. It was born with, um, unfortunate but ultra rare genetic disease, which, uh, uh, made his, uh, uh, on this gene called a CPS-1 and carbamoyl phosphate, uh, synthesis one, and the mutation renders this enzyme inactive, and therefore, uh, not able to digest, uh, proteins. And so the baby would have a chance, uh, you know, 50% chance of survival or, uh, to, to have, uh, his liver, uh, transplanted. And so, uh, uh, um, you know, about, uh, about a month period of over a month that, um, multiple scientists and institutions and got together and understood the problem and know exactly what the precise needed genome editing it, it, it was needed. And the, um, the baby KJ received the base editor that I talked about, uh, as those therapy, the, uh, therapy and, uh, is doing very well.
Um, another exciting, uh, discoveries are, for instance, Cleveland Clinic, uh, has, uh, created the off shelf, off the shelf, uh, CRISPR editing therapy that’s targeted to lower the cholesterol and triglycerides. And they use the CRISPR-Cas9 to, uh, uh, remove or silencing the proteins that inhibit what we call lipases. And these are the enzymes that’s responsible for breaking down, uh, these lipid molecules. Uh, so reactivating these, uh, lip lipase, molecules, that’s allow the patient, uh, uh, permanently, uh, because you edited the, the, the genome that lower the cholesterol and, and triglyceride. And so, uh, next slide, please.
Yeah. Uh, so that leads to, uh, that, um, I want to, uh, introduce, uh, in few slides of what we’re, uh, researching here, uh, in Van Andel that, uh, I, um, studied this unique, uh, Cas9 that’s currently not yet available. We’re trying to, uh, to fill the gap that there is no CRISPR-Cas9 that can be sensitive to, uh, DNA, um, modifications in the genome. So my colleague, uh, Peter Jones and Steve Baylin, uh, here at the VAI, uh, were the pioneers actually to study, uh, the DNA chemical modifications and their importance in biology, as well as the hallmark of cancer. So genomes could lose, uh, these modifications. They could also gain, uh, these chemical modifications. So, to developing a CRISPR technology that is able to discern the changes in these DNA modifications in genome, and we argue that they will have, um, enhanced the, say, uh, safety, uh, as well as unique, uh, applications.
So the way, uh, the, the, the sort of the pipeline of our, uh, research is to starting using computing to identify the bacterias, which could potentially, uh, host such, um, interesting CRISPR-Cas9 enzymes. And, uh, once we computationally identify them, and then we could, uh, uh, what we call, uh, biochemically characterize these genes, these proteins, and characterize their enzymatic activities. And then my group is, um, has this unique advantage. We’re using, uh, these powerful, uh, technology called cryo-electron microscopy, which is another, uh, revolution, uh, from the older technology, which called x-ray, uh, diffraction allow us to really visualizing these molecules. So in addition to knowing there’s biochemical properties, but we can also see them, and why is it important to visualize them that allow us to, uh, what we call to engineer them, to modify them, to optimize them to our needs. And I’ll show you an example of that.
And then obviously, finally, uh, we will, uh, use them in the human cells and potentially, um, for therapeutic use. So, an example that we have studied over the last few years is a unique CRISPR Cas9, uh, that’s isolated from this bacteria, uh, called acidothermus cellulolyticus uh, or we call AceCas9. We didn’t go to the Norris Geyser, uh, basin in the Yellowstone National Park, but we were able to, to use its DNA and clone them and study them in a laboratory setting. And we discover, uh, yes, interestingly, this, uh, uh, Cas9 is able to sensing that the DNA is methylated, and so it’ll perform the typical double stranded break, uh, when the DNA has no, uh, methylation or chemical modification, but, uh, it would prevent the Cas9 from cutting. So the key problem for this unique Cas9 is it is very inefficient.
Uh, it’s much, uh, less, uh, efficient than the one that’s previously isolated, for instance, by Professor Jennifer Doudna, uh, called a Spy Cas9. And so, uh, we went on to determine, uh, its, uh, what we call the three dimensional structures of this enzyme, together with, uh, the guide, RNA band in, uh, black, and the two DNA strands that’s shown in red and orange. Um, what’s powerful about this, uh, uh, three dimensional study, uh, structure study is we can actually visualize this enzyme in action. And we know that there is this, uh, pink, uh, parts, and that’s one of the, um, uh, what we call domains that’s responsible for breaking one DNA strand. And we saw that this domain undergo, uh, what we call the hinge motion. Uh, so to, to make the enzyme active, it must move it, uh, to the, to the DNA through this, uh, wide swinging of motion.
Uh, and, uh, we also, uh, determine the structure of this enzyme when the DNA now is methylated. So interestingly, when the DNA is methylated, that’s shown on the right, and this pink domain now is shown, shown in green, uh, no longer moves. It just stays where, uh, it is, and it just doesn’t, that’s why it doesn’t, uh, make the double stranded break. All right? So they gave us the idea that, um, this motion is critical for it to be, you know, uh, activated. So then, um, that allow us to focusing on the site of the motion, or we call the hinge that carries this motion. It’s kind of more like the elbow, the, the, the motion. So, uh, we performed something called, uh, protein directed evolution. So it to, to sort of, uh, using this, uh, analogy of, uh, the athletes that has to jump through this, uh, high bar that we create artificial situation, we make the bar higher and higher and higher, then we ask millions of athletes that who will differ and to, to ask them who can jump over.
So, and that allows us to be able to, to identify winners that we make the, the, the bar a very high. And then we found the winners. And in these winners has to be the, uh, Cas9 that bear some, um, changes in its hinge region. So it makes it stronger and faster. And so you can see the, what we call the wildtype. The native itself doesn’t have the activity, while this mutant allow you to, to, to perform the function very efficiently. And so we can also test these, uh, what we called the catalytic enhanced versions of the enzyme in human cells that indeed, uh, they perform better, uh, than the, you know, the wildtype enzyme. So this shows you an example of how important it is to characterize the biochemistry of these CRISPR enzymes and visualizing, uh, there are three dimensional structures so that you can really have a good, um, basis for you to say what part, which section of the enzyme that one can try to improve and to understand, uh, why it is, uh, making certain, um, uh, functions that it are not supposed to.
So, uh, next slides, please. Okay, so how do we harness the power of this, um, methylation-sensitive Cas9, for instance, we know the normal cells, many sites, uh, are chemically methylate methylated as this red dot shows, but a corresponding site in cancer cell could have lost it. And we can imagine that we can direct the Cas9 to the site where it lost the methylation in cancer cells, but not in normal cells, and therefore killing the cancer cell. Um, we can also imagine to drive this Cas9, to have the base editor, uh, base editing, um, ability to change the, the nucleotide, uh, in a methylation sensitive manner, and therefore, uh, you know, apply the, the function in the cancer cell, uh, specific manner. So we’re very excited about, uh, this discovery. Uh, so the future, uh, how do we, uh, continue to, to harness the, the, you know, this technology?
Uh, we definitely should continue to expand the various, uh, CRISPR, uh, tools. Uh, we definitely should further optimize the, uh, you know, efficiency. Uh, we really need to evaluate and improve the safety of these enzymes. Uh, before we introduce into the human cells, uh, one of the critical improvements is to improve how to deliver, uh, the enzymes, uh, into the, um, the human cells at the site, uh, of desired site. And so, uh, and, and the, in conclusion, I hope I have convinced you, uh, CRISPR technology is really, really easy to use and it is powerful and has been adapted by nearly all biomedical research labs. And there’s, you can talk to any research labs, and there’s probably hardly any who don’t use this technology compare, sorry, compared to the other previous genome editing technology, uh, CRISPR is RNA guided or programmable. And, um, and it’s very powerful and it can also, uh, can avoid, uh, the immune response.
Uh, we also anticipating that in the future, the, uh, emerging artificial intelligence, it will make it a very important role in further developing, uh, in the CRISPR technology. Uh, and you may or may not know that these, uh, technology, uh, in CRISPR based, which is not characterized as a GMO has been already used in, uh, the changing in the agriculture, uh, in the food, uh, that we actually eat. So, uh, to conclude, I would like to really acknowledge the, who actually did all these work. And, uh, a group of scientists that’s very inspiring, uh, or bring you from a graduate student to postdoctoral follows. Uh, some of them are undergraduate student, and I also are grateful for the, uh, what we called, uh, cryo-EM core facility here at, uh, uh, Van Andel, and as well as at FSU before I moved to here, and some national, um, centers of, uh, the Micros micro mic microscopy that allow us to perform, uh, these study. Uh, in addition, Van Andel is just a very supportive, very inspiring place to do research and, uh, the, um, all the departments, supporting departments, including all the board that’s listed here, uh, provided, uh, really, um, um, you know, uh, a strong support for our research. And I listed some of the, uh, collaborators that I currently, uh, working with. Uh, well, I guess I thank you for your attention and looking forward to some discussions.
Dr. Mary Winn [44:01]
All right. Thank you, Dr. Li for the wonderful introduction to CRISPR, gene editing and your research. Shall we get into some questions? All right. As a reminder to our audience, you may submit questions for Dr. Li in the chat, which I see we already have a couple in there. So let’s get started. You talked about it a little bit, Dr. Li, and you talked about the different functional modes of CRISPR.
Dr. Hong Li [44:27]
Yes.
Dr. Mary Winn [44:27]
And what is mode you, you mentioned is how you can influence a gene without, um, actually changing the sequence.
Dr. Hong Li [44:37]
Yes.
Dr. Mary Winn [44:37]
Can you give a specific example of how you or others are using that mode?
Dr. Hong Li [44:42]
Yes. So for instance, you could attach the CRISPR-Cas9 protein, when I say attach you, you, you have to develop what we call a fusion protein. So you make a CRISPR-Cas9 much longer, so it, it bears additional protein. And that protein has been, for instance, well studied in human or in other organisms that play a function, for instance, what we call the transcription activation. And that you can bring that molecule to wherever you want, and that to allow that gene to be, uh, activated. Or you can, if that part of the protein is what we call a repressor, for instance, and those are well studied roles of these proteins that they can block, um, the transcription, for instance, that is the step of making from, uh, you know, go from DNA to make RNA, so that that way you can, you know, stop making, uh, that gene so you can turn the gene off.
Dr. Hong Li [45:47]
Uh, so these are the examples of not breaking. Another example are these, um, uh, CRISPR uh, derived a technology where, uh, scientists have attached, uh, a DNA methyltransferase, that’s the enzyme that’s putting a methyl group onto the DNA, for instance. And then you can guide your, uh, this, uh, you know, fusion Cas9 as well as this, uh, uh, DNA methyltransferase to allow that section of the DNA to be putting on a chemical modification that otherwise lost the methylation, which will influence the, you know, silencing the gene, for instance, uh, if you remove it and you can activate the gene. So these are, um, some of the few examples that you don’t even have to make a cut. Simply Cas9 will act like as a, a vehicle to deliver these other machineries to the genome of your choice.
Dr. Mary Winn [46:54]
Excellent. So you mentioned that CRISPR is being used in a variety of different, um, tools and approaches. You’ve got the, you had the two examples from, um, the rare gene, um, and then also the, the more over the counter example, once a, a new treatment is identified using CRISPR or using a CRISPR tool, how fast does it take to get that into patients? Hmm. For example, if you’re gonna do something in cancer or, um, the example of that rare genetic disease, right? How long does it take to get to the, to the clinic?
Dr. Hong Li [47:41]
Yeah. I, I would not say that I, I’m the expert of the, the, you know, whole clinical trial process, but I was thoroughly impressed with the, for instance, the example that I showed that baby KJ was born with genetic disease, where, you know, I think within a month they, he received the, the therapy. Uh, that’s, that’s very exciting. But, uh, the whole FDA approve process and all that, that needs to be, um, you know, go through all the regular clinical trials and, and et cetera. But the, the technology itself is relatively simple to to be, you know, put together once you know what site that you wanted to, you know, you, you wanted to change.
Dr. Mary Winn [48:43]
What are some common misconceptions about gene editing that you run into when talking with your colleagues or friends and family?
Dr. Hong Li [48:53]
Some of them, I guess, uh, for instance, I, I think they are questioning about what is CRISPR <laugh>? They did not know it’s actually a bacteria, uh, repurposed protein, uh, and, and RNA, so the, you know, study of the bacteria led to the repurpose of these molecules that we didn’t, you know, even knew they exist before. Hmm. So <laugh>, that might be my one misconception. Um, I don’t know others, it could, could be that, uh, I mean, it depends on the person’s background. They, they don’t know what genome it is, then that’s much more, uh, there will be much more questions on what the technology is about. Um, perhaps they are uses of this CRISPR technology used in RNA that may maybe people don’t know. Mm. That you don’t have to just target DNA. You could, uh, you know, engineer the CRISPR-Cas9 or related, uh, CRISPR technology that can, don’t touch the DNA, so not permanently changing the genome, but to, you know, temporarily changing the message itself, the RNA and that, that’s another big use of the CRISPR technology too.
Dr. Mary Winn [50:41]
Yeah, that’s an exciting, uh, approach and yes, exciting to see where, where we go with that in That’s right. <crosstalk> to human health.
Dr. Hong Li [50:49]
Yes. Mm-hmm.
Dr. Mary Winn [50:50]
So one of the things that you are trying to do is identify CRISPR um, tools that read those methylation marks or other marks on our DNA. And you talked a lot about methylation. Can you explain for the audience what methylation specifically is and how that functions?
Dr. Hong Li [51:13]
Yeah. Um, so in our genome, so these DNAs comprised of these nucleotides, these are the, the, you know, ring-shaped, uh, molecules that we have four different types on our DNA, uh, called adenine and guanine, cytosine uh, thymine, these DNA, these nucleotides, normally they are just as these chemical structures, they maintain these chemical structures, but in, uh, mostly in eukaryotic cells such as plants, animals, uh, some of these, uh, cytosine, for instance, the nucleotide can be added an extra chemical group called a methyl methylation. So, uh, the addition of a chemical, uh, of the methyl group on the C in the genome in multiple positions, multiple locations in the genome, uh, have a profound impact in altering the message that the DNA codes, uh, you know, you could, uh, making it, um, uh, mostly is for, uh, making suppress certain genes or, uh, turn on certain genes that’s they, otherwise, the message alone, uh, cannot, cannot direct that message to, uh, so it’s, it’s, and then, you know, uh, our colleagues, this study has found that this methylation can go wrong too. Um, and when they go wrong, and when they just lost, they’re normally methylated pattern or they, uh, normally gain of methylation that allow the, you know, cell to behave differently, for instance, in cancer, that you have a massive loss of, uh, DNA methylation, uh, and, and then that is a, a reason to turn on, for instance, some genes and et cetera. So it’s a very, very important, uh, biology that regulates our, uh, basically the information highway.
Dr. Mary Winn [53:37]
Wonderful. Um, I think that helps explain, right, yeah. The importance of those marks and modifiers on, on DNA beyond just the A’s G’s, T’s and C’s.
Dr. Hong Li [53:48]
Yes.
Dr. Mary Winn [53:49]
So you talked about kind of the next steps, the future, and one of those was AI, right? And these new AI models and, um, for protein folding and structure, how do you see, or how are you leveraging those already, um, to impact the landscape of what CRISPR can do?
Dr. Hong Li [54:13]
That That’s a great question. Uh, so, um, we have already seen some of the AI, uh, technology being applied in engineering of the enzymes that they, um, for instance, you can take the existing CRISPR Cas9 enzyme, then apply the, you know, AI tools to them and to allow them to be functioning, you know, stronger, be better and safe. Uh, you know, that, that’s much, I guess, computationally much more easily to be realized than we do all these, uh, protein directly evolution, uh, to, to come up with a computational model to allow these, uh, enzymes to behave better. That’s, that’s definitely one of a good example of how AI can be used. And I’m sure that AI could also be used to predict, uh, and avoid perhaps these off target that, uh, that can be, um, you know, that causes the safety concern, um, because there’s a lot of genome, um, data that’s there that can be put together, you know, in a language model to, to, to learn.
Dr. Mary Winn [55:48]
So we don’t have a lot of time left. There’s a ton of really great questions, um, in the chat that we won’t be able to get to, unfortunately. But just to end up kind of what, what is it the most exciting to you about your research and kind of the next steps?
Dr. Hong Li [56:05]
Mm. Yeah. This is, there’s really no ending in this research because the bacteria genome, so much bacteria in the world. So in fact, the bacteria outnumber, uh, the, the, you know, even the bacteria in our human body outnumber, uh, our human cells. So there are so much diversity in CRISPR or related technology that can be mined from these microbes, uh, that you never know what is the new next, uh, CRISPR like technology. Uh, we have seen some of them in early publications now of these new unexpected enzymatic, uh, molecules that can perform things we don’t yet know yet. So, um, and another, I guess excitement is the, the education. I think, um, it’s, it’s very satisfying to see, you know, like students all the way from K–12 level. They are excited about the CRISPR technology that we are, in fact, we are actually making a field trip for our Van Andel Education Institute to allow the, you know, the middle schoolers or high schoolers to be able to experiment in, uh, CRISPR technology. And that shows how simple it is to do and how powerful it can be, and how easy is to understand for these young, uh, generation of scientists and, and, you know, hopefully they can come and make new discoveries. I’m excited about that.
Dr. Mary Winn [57:51]
That’s, that’s really exciting.
Dr. Hong Li [57:51]
Yes.
Dr. Mary Winn [57:53]
Well, That is the end of our time. Thank you Dr. Li for sharing your insights into this incredible topic. And thank you to our audience and the VAI community for taking time to learn more about this impactful research, you can visit vai.org to learn more about Dr. Li’s work and more initiatives taking place across the Institute. Make sure to sign up for our mailing list and follow us on social media to stay up to date with the latest news.