The rockefeller university

Robert Darnell has spent 30 years chasing an unexpected thread from rare brain diseases to the basic mechanisms of how cells read their DNA. (Credit: Lori Chertoff)

When Robert Darnell, MD, PhD, was launching his research career more than thirty years ago, he became fascinated by paraneoplastic neurological disorders (PNDs), a group of syndromes so rare that most doctors had never heard of them. Patients with lung, ovarian, or breast cancer would suddenly show signs of severe brain disease. They would lose sensation in their limbs, become unable to balance, or develop sudden memory loss.

Darnell wanted to know why.

Chasing that mystery led Darnell down a research path he never anticipated: how cells regulate RNA, an intermediate genetic molecule that is used to carry DNA’s message to protein-making machinery. But the foray from PNDs to RNA was just the first twist in Darnell’s career. Since then, he has steered his research in countless new directions. In recent years, his lab has made breakthrough findings about the role of RNA in intellectual disability and autism, where loss of an RNA-binding protein causes Fragile X syndrome—the most common inherited form of intellectual disability and a leading inherited cause of autism. His lab has also studied rheumatoid arthritis and Parkinson’s disease, where RNA signatures in the blood may be able to predict symptoms before they appear, and even the evolutionary origins of spoken language, where a single RNA-binding protein carries a variation unique to humans.

What connects all of it is not only a fascination with RNA regulation, but also with curiosity itself.

We talked with Darnell about why RNA regulation is so much more complex than most people realize, and how his basic research could point toward new diagnostic tests and therapeutics.

You started out as a neuro-oncologist studying rare brain diseases. How did that lead you to RNA-binding proteins?

There’s a general lesson in how I got into this, which is that sometimes the most esoteric science leads to new clinical or scientific implications that you didn’t see coming.

I trained in neuro-oncology at Memorial Sloan Kettering under Jerry Posner, one of the great neurologists of our time, who had been studying paraneoplastic diseases in cancer patients for years. What we figured out by studying blood from these patients is that in these diseases, a tumor—already present elsewhere in the body—turns on a gene that is normally only active in neurons. The immune system suddenly sees this neuronal protein being made in a tumor and starts to attack it. But at some future time, immune cells also start attacking the brain, and that’s where the neurological devastation comes from.

The brain proteins that were being recognized by the immune system turned out to be RNA-binding proteins. They attach to bits of RNA in cells to regulate how and where the RNA is used to make proteins. That set me on the path I’ve been following ever since, trying to understand what these RNA-binding proteins do in the brain, and how this knowledge can be applied to a variety of human diseases.

What have you discovered about why RNA regulation is so critical for human health and for the brain?

Neurons have to be incredibly flexible and fast-acting in terms of their ability to quickly become active or shift into a new state. And so the brain has lots of really complex ways to control how RNA is processed as a way of controlling how and when it uses different genes.

Most people think of genes as the fundamental unit of biological information; you have a gene for this, a gene for that. But what’s less appreciated is how much can happen to that gene between when its DNA code is first copied into RNA, and when that RNA is used to make a protein. A gene isn’t used to make the exact same proteins in the same place and the same way all the time.

In most cases, RNA from a single gene is cut up and only sections of it are used to make protein variants in different conditions. RNA can also be shuttled to one area of the cell—like the far end of a neuron—so that a protein is only made there. And RNA-binding proteins can control whether RNA is used at all, or just stored up for later when translating it into a protein is needed.

We’ve learned is that this regulatory complexity is especially important in the brain.

What other avenues has that led you down?

Many, and often in directions I never anticipated.

When we started studying RNA-binding proteins, we had no reason to expect it would give us insights into a developmental disease, but then we discovered that the protein associated with Fragile X syndrome, called FMRP, is closely related to the paraneoplastic proteins we were already studying, including the neuronal NOVA1 RNA-binding protein.

Remarkably, we recently found that a specific variant in the NOVA1 gene that is unique to modern humans appears to play a role in the brain circuits underlying spoken language. Putting the human form into mice altered both RNA splicing and, in studies with my Rockefeller colleague Erich Jarvis, altered their vocalizations—suggesting that NOVA1 may have contributed to the evolution of neural circuits underlying spoken language. We now have evidence from 10 patients who have NOVA1 mutations that they present with language problems. These discoveries came entirely from basic curiosity about what that protein was doing; we weren’t looking for a language gene.

As we’ve gained an understanding of how important RNA-binding proteins are in health and disease, it’s led us to clinical studies and collaborations as well. For instance, we’ve used blood samples from patients to track how changes to RNA can predict flares of rheumatoid arthritis.

You can use blood samples to track brain RNA?

Yes, and I think it’s one of the most exciting areas in RNA biology right now. We’ve learned that RNA is remarkably dynamic; it’s changing every few minutes in response to what’s happening in a cell. That means RNA can tell you things about the current state of a tissue that DNA simply can’t, because DNA is essentially fixed.

On top of that, it turns out that some of that RNA is released by cells into the blood and saliva. And if you know what to look for, you can read it. In addition to our work on arthritis published in The New England Journal of Medicine, we also published a paper in Nature Communications showing that we can detect RNA signatures in the blood of Parkinson’s patients that correlate with their disease state. The idea is that the RNA could give you an early warning signal that something is about to go wrong, either in the joints or in the brain, before it becomes visible in a patient’s symptoms.

We think this could be useful in a number of diseases, to both diagnose disease and track it over time. It could be used to gauge how well a treatment is working, for instance.

To study RNA regulation in the brain, you had to invent a new tool—CLIP—and you’ve been creating new versions of it for decades now. What does it allow?

The way people were studying RNA-binding proteins before CLIP was that they’d purify protein and RNA separately, mix them together in a test tube, and study their interactions. As a neurologist, that was unsatisfying to me. I wanted to know what was actually happening in a living brain when there’s a whole complex mess of molecules present, not only what two molecules do in the test tube.

What CLIP lets you do is freeze interactions between a protein and RNA exactly where and when they occur in real tissue. You shine UV light on an intact brain, and it locks RNA-binding proteins to whichever RNAs they happen to be touching at that moment. Then you can go in and read exactly what was happening: which proteins were bound to which RNAs and exactly where they interact. You can compare what RNA-binding proteins are doing under two different conditions, such as in healthy versus diseased brains.

We’ve kept pushing this further. Our most recent version can do this in human brain tissue at the level of individual cells, and even in small areas within those cells, which lets us see exactly where in the brain RNA regulation breaks down in disease. Ultimately, that’s the kind of detail we need to start designing drugs that restore healthy RNA regulation and treat disease.

Your lab has followed RNA regulation into so many different areas. How do you decide what to pursue?

Honestly, I follow the science wherever it leads. That’s the privilege of basic research; you’re not obligated to stay in one lane. I’m a strong believer that the value of basic research is in the doors it can open. Clinical medicine often points us toward particularly interesting doors, because disease reveals places where our understanding of biology is incomplete. But you rarely know what you’ll find when you open any door—or where it will lead next. That’s the excitement of basic science.