Francis Mojica: "The only person who was sure I wouldn't win the Nobel Prize was me."

Nothing could have led Francisco Juan Martínez Mojica to foresee, or Francis MojicaAs is popularly known (Elche, 1963), the hours and hours he spent in the Santa Pola salt flats inspecting microorganisms that gave the water its peculiar pink color would eventually lead him to a discovery that would revolutionize biology.

Indeed, the work of this microbiologist from the University of Alicante has laid the foundations for the development ofCRISPR/Cas9 gene-editing tool for which Emmanuelle Charpentier and Jennifer Doudna were awarded the Nobel Prize in Chemistry in 2020. These genetic scissors have become a fundamental tool for developing personalized therapies to cure babies with ultra-rare diseases or to create plants more resistant to pests and droughts.

Mojica recently participated in the lecture series Greats of science at Cosmocaixa, where he spoke about CRISPR-Cas9. He receives the ARA newspaper the next day at his hotel. Recalling the talk, he explains, with tears in his eyes and visibly moved, that an hour after it ended, people were still queuing to congratulate him and thank him.

What has he told them?

— The history of CRISPR, its beginnings, and how this true revolution came about. But instead of boring you with a long, tedious explanation, I've compiled news stories about advances made possible by CRISPR technology. There are thousands! I've chosen a few, very illustrative ones, such as... cures for babies with illnesses, Because it's amazing. These are the kinds of apps that touch your heart.

Could he have imagined it when he was in the salt flats of Santa Pola, in Alicante, looking for bacteria?

— I wasn't even looking for it! Sometimes things happen and you don't know why. I studied biology in Valencia and I was fascinated by microorganisms. Later, I went to the University of Alicante, which is near my house, because I live in Elche. There, in the Faculty of Medicine, there was a microbiology division, where they worked with pathogenic bacteria, and I thought it was really cool. After all, they're the cause of most deadly diseases. My surprise was discovering that they were biologists. boot, who worked collecting and fishing microorganisms from the salt flats of Santa Pola.

And what did that have to do with health?

— Absolutely nothing! At first, of course. Because now we know that these microorganisms produce carotenoids that give the salt flats their reddish color and also that they slow the growth of tumor cells. However, at that time, that research was pure scientific curiosity. I started a doctoral thesis to find out how these salt flat microorganisms, and one in particular, are able to adapt to changes in the salinity of the environment. But I went to a conference on halophilic archaea, like the ones I was studying, and I saw that this had already been studied, so I had to change my topic. At the university, we had a gene library, a collection of fragments of the microorganism, and we suspected that some of them might be related to adaptation or response to salt. I sequenced them and found very curious repeats, fragments of DNA duplicated many times and perfectly ordered, so the distance between one repeat and the next was always the same, while the sequences in between were different. I remember wondering what on earth that was.

Didn't he think it was a mistake?

— Yes, at first I thought I'd made a mistake in the sequencing. This was in the early 1990s, and those were the first sequences ever done at the University of Alicante. But we repeated the sequencing and got the same results. It was impossible for us to make the same mistake twice with the same accuracy. Initially, I thought those repetitions were involved in the DNA structure, and with the limited bioinformatics tools we had at the time, I analyzed them and saw that they predicted extraordinary flexibility in that region. My first hypothesis was: changes in the salinity of the medium, changes in the ionic strength inside the cell, and this affects the DNA structure. But nothing came of it. After several experiments, we proposed that it was a mechanism for chromosome segregation and distribution.

He was 24 years old at the time and published it in a leading microbiology journal of the era.

— Yes, but it was a lie. That's how science works, progressing slowly. The journal required us to propose some explanation for those repetitions, and you know how it is, to get published, you have to make a deal with the devil... However, we were cautious. We continued working and saw that other groups had described these repetitive regions in bacteria.

Were you the one who coined the term CRISPR?

— First we called them TREPs, an idea from my boss at the time, but Tandem RepeatIn 2001, a Dutch group working with repeats in another bacterium wrote to me to say they had found nothing alongside the repeats. Therefore, they must be functionally related. They told me they didn't like the name I had given them and that they would submit the article with a different title. That didn't make sense. We had to reach an agreement. We were the only two groups working on this, plus another one in Copenhagen. That's when I came up with CRISPR, the acronym for Clustered Regularly Acquired Short Palindromic Repeats. And they liked it very much.

If there were any press headlines about CRISPR...

— If I got paid a penny every time CRISPR was used in an article, I'd be rich by now. As a name, it sounded ridiculous to me. You wouldn't believe the jokes I got... When I ran into someone at university, they'd say, "Look, it's the guy with the crisps." Or: Mr. Crisps, Mister Crisps, all sorts of things. Even my wife said it was okay as a dog's name, but as a scientific name... not so much. But it was easy to pronounce, catchy, and look.

When did he realize that the repetitions were an immune system?

— Ever since we found them, I'd wondered where they came from. The first complete bacterial genome wasn't sequenced until 1995. Therefore, there was very little genetic information in the databases. But by 2003, the situation had changed considerably. And we found, by sequencing the CRISPR repeats ofE. colia spacer that was identical to that of a virus that infects this bacterium. We found in the literature that this particular strain was resistant to infection by phage p1, a virus. We continued to look E. coli And we found some more spacers, then we analyzed all the spacers in 86 genomes. It was all very manual, because I had no idea about bioinformatics: I would copy the entire sequence, replace the CRISPR repetitive sequence with nothing or with a text sequence, and separate the spaces. I would take that, upload it to the database, and compare each spacer one by one with what was there. When I found a match, I saw that it was a virus that infected that species of bacteria.

They are, therefore, a microbial protection mechanism.

— The most surprising case was that of Fehleisen's streptococcus. There were six sequenced genomes, and only one had CRISPR; it was precisely this CRISPR region that was resistant to the virus. We proposed that these CRISPR regions were a memory of infections, fragments of the invading virus's DNA. Later, the associated proteins, Cas, were identified. Prokaryotic cells store fragments of genetic material from invaders in the CRISPR regions, which they use to recognize and destroy future threats thanks to the action of the Cas proteins.

Why was it so revolutionary? Was it really so impossible for microorganisms to also have a defense system?

— In 2012, what wasn't known was that they could have an acquired immunity system. Defense systems or defense strategies against a virus are known, but CRISPR is the only one with adaptive capacity. As if vaccinated, it acquires genetic memory. And this is absolutely unique, a challenge to Darwinian evolution.

Because?

— Because Darwin said that changes occur randomly, and then those that confer the greatest evolutionary advantage are selected. In contrast, [Jean Baptiste] Lamarck thought that the giraffe, for example, grows a longer neck because it stretches it to reach the trees. Today we know that this isn't the case; rather, in the face of change, the giraffe with the longest neck will be the one that feeds best because it can reach higher; it will survive, have a greater chance of reproducing, and pass on its genes.

And what about microorganisms and CRISPR repeats?

— They are an intentional adaptation. The microorganism takes foreign genetic material and integrates it into its own. It's not a random change. While it's true that the fragment isn't selected, the bacterium is intentionally, voluntarily, modifying its own genetic information. There is a mechanism for integrating this genetic material. And this is a paradigm shift.

Five magazines refused to publish his findings.

— Years later, a reviewer of my article confessed that he rejected it simply because he couldn't believe it. It was the sixth time we sent it that they finally accepted it. And it still took a year to be published.

At that time, did you imagine the applications that CRISPR would have later?

— No and yes. [Laughs] If you want them to publish your article, you have to sell it as if it's going to be amazing! We argued so they'd publish it. Nature which would have a huge impact on clinical practice, biotechnology, biology, etc.

Were you surprised that Jennifer Doudna and Emmanuelle Charpentier won the Nobel Prize in Chemistry for developing a gene-editing method based on CRISPR?

— When Doudna and Charpentier's article was published proposing the use of CRISPR as a gene-editing tool, I remember a colleague coming into my office and saying, "Can CRISPR really be used for editing?" I had no idea at the time.

Did it hurt him not to be recognized?

— The only person who was certain I wasn't going to beat him was... me. Back in 2020, I was in my office working when a journalist from the EFE news agency came in. He reminded me that the Nobel Prizes were being announced that morning and that they might call me. I was sure there was no chance, but he logged on to the ceremony on his phone and we watched it live. When they announced that the chemistry prize was going to Doudna and Charpentier, the journalist was utterly dejected. Right at that moment, one of my colleagues came into the office, indignant because they hadn't given the prize to Feng Zhang, a biochemist from MIT, who was the first to apply gene editing to human cells. It was funny to see the EFE news agency repeating "I can't believe it" and my colleague saying the same thing about Zhang!

How do you feel when you think about the countless applications that your discovery has enabled?

— It's incredible. For me, it was already amazing, personally, to be able to answer a question I'd been asking myself for ten years, and even more so when the answer is as wonderful as this one. Everything that has come after is unimaginable and impossible to grasp. There are so many applications, so diverse, that they're making every other tool or discovery in molecular biology look like a distant memory.