The Jennifer Doudna Lab Knocks CRISPR Out of the Park

Jennifer Doudna is a world-renowned biochemist at the University of California, Berkeley, best known for her foundational research on CRISPR, a gene editing technology that has transformed biological research in less than a decade. She has remained a leader in the CRISPR field, becoming a spokesperson for the ethical use of genetic engineering for all applications — from human disease to agriculture, and leading a powerhouse lab that continues to push the boundaries of biology. Together, her lab members are working towards another decade of CRISPR innovation.

Meet the JD Knockouts: Experts in the CRISPR Field

On Sunday evenings in the fall, many members of the Doudna Lab meet up on a softball field at the edge of the Berkeley campus. The lab's intramural softball team, aptly named the JD Knockouts, has been a tradition for "as long as anyone can remember," according to Jenny Hamilton, a postdoc in the lab (and a captain of the team). The team plays against other groups in Berkeley's Molecular and Cell Biology department.

While only about a dozen of the 40+ Doudna Lab members choose to play softball, the team represents the lab's diversity of interests and experiences: it is comprised of undergrads, research assistants, grad students, and postdocs. Some of the players are veterans in the lab, and others have just joined. Each of them study a unique aspect of CRISPR-Cas systems.

"As you can imagine, we have a huge lab," says Haridha Shivram, a post-doc. "We do three broad things: discovery of new systems — new CRISPR-Cas systems, understanding their mechanism — how exactly at the molecular level they work, and then how we apply them — how can we engineer them, how can we make them better as genome engineering tools?"

Haridha's project falls in the discovery bucket; he's interested in finding proteins that inhibit the activity of CRISPR-Cas systems, known as anti-CRISPRs. Anti-CRISPRs are found in bacteriophages; they are bacteriophage's natural defense against bacteria, just as CRISPR is bacteria's natural defense against bacteriophages. Haridha takes advantage of his background in bioinformatics to uncover new anti-CRISPRs, and, once he finds them, he ultimately wants to understand how they work. Though his research is foundational, he and others in the field expect it to impact how CRISPR is applied. For instance, a visiting Master's student named Simon Eitzinger is trying to address the critical problem of off-target effects with anti-CRISPRs. In theory, he says, anti-CRISPRs could be programmed to reduce off-targets by inhibiting Cas proteins after they have cut DNA at intended target loci.

Other Doudna Lab members, such as postdoc Gavin Knott, want to know exactly how canonical CRISPR-Cas systems find and cut their target (or, their off-target) sequences. "My research is very much focused on molecular details — working on how these proteins really function, understanding modes of chemistry, and what they look like at an atomic level." It is no surprise that Gavin and several of his labmates were drawn to the Doudna Lab to study the structure and mechanism of CRISPR-Cas systems. Even before Jennifer Doudna was known for her work on CRISPR, she was well-respected for her work on the structural biology and biochemistry of RNA enzymes. Both Doudna and her lab members recognize that thoroughly understanding CRISPR activity at a molecular level will help others apply the technology. Gavin explains, "It's kind of akin to popping up the trunk of a car and asking how do all these pieces fit together, how do we optimize them so that it's the best machine possible to get you from A to B."

Understanding CRISPR activity isn't a small task, especially when scientists want to use CRISPR gene editing to solve a host of biotechnological problems. Liz O'Brien, another Doudna Lab postdoc, is looking at CRISPR activity from a different lens: she is evaluating the behavior of CRISPR-Cas systems in live cells. Liz fluorescently tags CRISPR effectors (Cas proteins) and tracks both their targeting behavior and the DNA repair outcomes of CRISPR activity. "We also need to understand how CRISPR effectors are behaving in live cell environments. These are proteins we want to use as therapeutics."

Indeed, a large group of Doudna lab members are actively working on optimizing CRISPR-Cas for therapeutic purposes. Jenny, the softball team captain, and Connor Tsuchida, a graduate student (who also happens to be a softball team captain), are interested in delivering CRISPR to human cells. A major problem they're trying to tackle is delivering CRISPR selectively to cells involved in a genetic disease. If they can do that, they will have more control over the gene editing process, and ultimately its outcome. Connor elaborates, "The way that we can make our technologies really, really innovative is by making them specific in the delivery. There are a lot of ways to do this editing in a test tube. But the challenge is getting across biological barriers, getting to specific cells, getting to specific tissues."

Building a Team with Benchling

In a lab as large and interdisciplinary as Jennifer Doudna's, capturing experimental notes and results and sharing that data becomes a significant challenge. For that, the lab has turned to Benchling. Abby Stahl and Enrique Lin Shiao, both postdocs who joined the lab in the last six months, were introduced to Benchling when they started. For Abby, it's the first time she's used an electronic lab notebook. Both Abby and Enrique quickly learned that Benchling helps them organize their initial ideas and experiments chronologically, which they can now more easily revisit or reference.

For members of the lab who've been around a bit longer, Benchling is a key part of the way they collaborate on a daily basis. This is particularly helpful for lab members who mentor others in the lab, such as Gavin and another postdoc, Brady Cress. Both Gavin and Brady work with small teams, including technicians and graduate students. Within their teams, they send notes, protocols, and results back and forth through Benchling, centralizing their workflow at the bench.

This centralization also helps them take advantage of protocols and data that were produced by people who have since left the lab. For Gavin, the ability to search through Benchling is critical. "The feature that I like the best in Benchling hands down is the 'control F' function. Being able to search through protocols, projects, directories for something that was written back in 2012 — and it's legible — is amazing. It beats going through Tupperware boxes full of notebooks, trying to find that protocol that you really, really need."

I really appreciated that there were experts from a large variety of different fields so that you could have one expert in neurology and another expert in structural biology. I think that's where most collaboration starts.

Collaboration in the Doudna Lab extends beyond Benchling. Collaboration is part of each lab member's everyday, whether at the bench, in the break room, on the way out in the evening, at softball. And, in any of these settings, it's palpable. In fact, if you ask any member of the lab why they joined, they are almost guaranteed to first describe the way lab members work together. "I really appreciated that there were experts from a large variety of different fields so that you could have one expert in neurology and another expert in structural biology," says graduate student Joy Wang, describing why she chose the Doudna Lab. "I think that's where most collaboration starts."

The sense of collaboration helps to explain why Jennifer Doudna's students and postdocs have been so successful. They are certainly fortunate to work on some of the most cutting-edge science, but they also seem to benefit from working as a team. To the backdrop of the JD Knockouts' softball game, Connor explains, "I think the camaraderie in a lab makes a really, really big difference. A lot of people's best ideas are not necessarily at their desk or at their bench, but just chatting with people at softball or talking with people at the bar."

CRISPR-Cas Innovation On Deck

For obvious reasons, Jennifer Doudna's lab is widely acknowledged as one of the best CRISPR labs in the world. But it's not just the reputation that draws new students and postdocs to the lab. It seems to be either the interdisciplinary and collaborative nature of the lab, or the desire to apply science to a meaningful set of problems, to see real-world impact. CRISPR "has huge potential for a lot of global challenges that we are facing to date — some of them from climate change to human diseases, loss of biodiversity," says Enrique, synthesizing the ambitions of many of his labmates. "There are all these very global fundamental problems in the world right now, and I think gene editing can provide some of those solutions."

Each lab member's ultimate vision for CRISPR — or gene editing in general — often directly motivates the work they are doing. Brady, for example, is developing CRISPR-Cas systems that can be more precisely controlled. Using protein engineering strategies, he is designing on- and off-switches for CRISPR, a technology that could help control metabolic pathways in various organisms. He ultimately envisions this being used to creatively address medical problems: perhaps, to detect physiological changes associated with disease states or, maybe, to optimize the biosynthetic capacity of microbes for drug production.

CRISPR has huge potential for a lot of global challenges that we are facing to date — some of them from climate change to human diseases, loss of biodiversity. There are all these very global fundamental problems in the world right now, and I think gene editing can provide some of those solutions.

Many lab members are motivated to solve problems in human health and disease. Liz's big dream for CRISPR is to develop personalized, preventative cancer therapeutics. For Abby, the most incredible application of CRISPR medicine would be to delay the onset of age-related diseases, especially in elderly populations. Jenny wants to treat genetic diseases with targeted CRISPR therapeutics. She, like several others in the Doudna Lab, is confident that CRISPR will be deployed broadly in the clinic in ten years.

Jennifer Doudna's lab and the CRISPR field still have a lot of work to do to make these ideas a reality. As any scientist knows, not all of their ideas are a sure bet. But there is one thing that the Doudna Lab — and the entire scientific community, for that matter — knows definitively. In Gavin's words, CRISPR-Cas is "a tool that will and has already changed molecular biology."