The Explorer Who Went Small

The cows had to be milked twice a day. Holiday, birthday, funeral, wedding, it didn't matter. If nothing else, Ronald Breaker, PhD, knew that, growing up on a dairy farm 25 miles outside of Steven's Point, Wisc.

That same farm kid would become a Sterling Professor at Yale University, chair of two departments, and a Howard Hughes Medical Institute Investigator. He would discover riboswitches—ancient RNA devices that sense molecules and control genes—and solve a 150-year-old mystery about an explosive chemical found in bird droppings.

But perhaps most remarkably, he would maintain the same relentless work ethic and unflinching sense of duty that he learned while milking cows at dawn.

"There's a responsibility that is bigger than you," reflects Breaker, chair of the Department of Molecular Biophysics and Biochemistry. "I didn't know any other view. I took that same approach in tackling the challenges in science in general."

Growing up, there wasn't much in the way of scholarly materials in the Breaker family farmhouse. But there was one thing: an old globe so outdated that the North Pole was labeled “Unexplored Territory.” Breaker would look at those words and think, "I want to explore. I want to go there."

He later realized that the North Pole had, in fact, been fully mapped. For many children, this would be disappointing. For Breaker, it was redirecting.

"I'm absolutely an explorer," he says now. "I've just gone small."

Breaker earned his bachelor’s degree in science and chemistry from the University of Wisconsin-Stevens Point and his doctorate in biology and biochemistry from Purdue University. As a postdoc at The Scripps Research Institute in San Diego, he had a breakthrough so stunning that he told his wife Michelle, "I don't care how long of a career I have in science. I will never do anything more spectacular than what I just did today."

"I created a test tube filled with trillions of different DNA molecules," he says. "And I had a method to identify that crazy rare sequence—maybe one in a trillion—that could fold up and cut an RNA molecule." This was remarkable because, at the time, DNA was thought only to store genetic information; it was not the action molecule of the cell. "I was able to prove for the first time that DNA could fold and act like a protein or act like some RNAs."

So by the time Breaker arrived at Yale in 1995, he was already making DNA do things it wasn't known to do. In fact, he continued focusing on making DNA perform other chemical reactions. "We got really good at that," he says. "In fact, I probably got tenure at Yale because we got really good at forcing DNA to fold up and act like enzymes."

The RNA World theory is the idea that before DNA and proteins dominated biology, RNA ruled everything, storing all the genetic information needed for organisms to function. Breaker explored this theory as a professor at Yale. "I thought that if the RNA World theory is correct and organisms built out of RNA became really complicated, then there had to be RNA sensors and switches," he says.

RNA sensors are the RNA that organisms need to sense and interpret their environment. A switch is a mechanism that turns certain genes or enzymes “on” or “off.” These capabilities enable RNA to monitor how things are going and then activate or deactivate processes needed to maintain all the activities required for these complex creatures to thrive.

And so—using the same methods he used at Scripps—his lab went about engineering those RNA switches in test tubes, getting them to fold, bind targets, and turn enzymes on and off. Just as he had discovered how to fold DNA at Scripps, Breaker was repeating that at Yale to master the art of folding RNA.

Then came a fateful lab meeting that Breaker remembers clearly.

A postdoc named Garrett Soukup, PhD—now at Creighton University in Omaha—was presenting data on their latest engineered RNA switch. The results looked really good. Breaker stopped the presentation and announced, “We created this technology in the laboratory! But you've got to believe that this technology was once dominant in biology, and that it’s good enough to not have been driven to extinction. It's going to still be with us today. We just have to be smart enough to know where to find it."

Then he described exactly how they would find it: Geneticists studying gene control would have encountered these switches without knowing what they were studying. They'd be frustrated that they couldn't find the protein that regulated their gene—because there was no protein. It was RNA all along.

The very next day, another postdoc, Narasimhan Sudarsan, PhD, walked into Breaker's office holding a paper. "I think this is what you're looking for," Sudarsan said. It was. His team ultimately found seven papers describing what Breaker had anticipated. Seven hidden switches in bacteria that researchers had studied without recognizing them.

"Science is a series of alleys that need to be explored, and sometimes those alleys are blind alleys," Breaker often tells his trainees. "If I had to pride myself in anything that we do, it is that we want to identify the strangest RNAs and DNAs and figure out what they do."

Breaker's team systematically proved that every switch was real. They called them riboswitches. The name stuck.

Breaker's lab doesn't look like a movie set. "It doesn't look much like a 1930s Flash Gordon laboratory," he says. "It looks much more like a Tupperware party."

Plastic tubes, pipette tips, plexiglass shields. People in lab coats peering at bacterial plates, manipulating trillions of molecules. "It's plastic," Breaker explains, "and hopefully a bunch of smart, eager people asking really important questions."

Getting others to see the importance of his work was a constant struggle. Breaker remembers standing in a hallway near the mailboxes at Yale after opening a letter. His first riboswitch paper had been rejected by a top journal.

"I remember just leaning back against the wall in the hallway near the mailboxes and thinking, 'I know how to get my papers accepted. I just have to change my science and do what a lot of other people are doing, choose topics or fields that seem to be more well appreciated by my fellow scientists or journal editors.'

"But then, I thought, 'No, no, every part of my scientific training is screaming that you're doing the right thing!'"

He did not change course. When he did start publishing on bacterial riboswitches, he says some people responded with a patronizing "how cute." Simple organisms using simple mechanisms. They didn’t think riboswitches would apply to human biology, as humans would clearly use proteins instead.

Breaker disagreed. “If these RNAs were not good at what they did, evolution would have thrown them into the evolutionary trash heap," he argues. "They're here today because they're actually very good at what they do."

Breaker kept finding more riboswitches in bacteria and insisting that these ancient RNA devices existed in humans, too. His colleagues kept asking: "Breaker, where are the human riboswitches?" His answer never changed: They're out there. We’ll find them in bacteria first, he said, where genomes are smaller and cleaner. Then we’ll look for similar sequences in humans.

"My view was that humans could have the same technology, but it was going to be much more difficult to find,” he says.

In the 1850s, the German chemist Adolph Strecker heated bird guano to high temperatures. From that steam emerged guanidine, a small, nitrogen-rich molecule that reacted violently with oxygen. By the early 20th century, it was being used in slow-burning explosives to propel shells from battleship cannons. Today it's in airbag propellants.

In the 1870s, researchers noticed something odd: Guanidine made frog muscles twitch. Over the next century, it was sporadically tested as a treatment for neuromuscular disorders. Sometimes it worked, but no one knew why. For 150 years, guanidine's effects on muscle function remained "something of a mystery," as Breaker puts it.

Breaker’s lab went on to discover four classes of bacterial riboswitches that specifically bind and sense guanidine. Then came the leap. Using computational tools, Breaker's team searched vertebrate genomes for similar RNA structures. They found dozens in elephants, dolphins, birds, reptiles—and humans.

These guanidine-sensing RNAs clustered near genes involved in calcium signaling and neuromuscular function, such as CA8 or ITPR1. Mutations in either gene can cause spinocerebellar ataxia, a devastating disorder that leaves some patients so impaired they can only walk on all fours.

The team biochemically validated that these vertebrate RNAs actually bind guanidine. Some even more tightly than their bacterial counterparts. When they expressed human CA8 protein in bacteria, it increased cellular guanidine levels. The mystery that started with twitching frog muscles in the 1870s was finally making sense.

“Guanidine may be a long-overlooked signaling molecule that plays a critical role in neuromuscular health,” Breaker explains. “And RNA molecules—once thought to be simple genetic messengers—may be active participants in one of the body's most complex systems controlling muscle function.”

Breaker is protective of the freedom to explore for its own sake. "That’s the one beauty I think, which I would defend to the end. The idea that academic scholars, we're expected to look in unusual places, think about unusual things."

This freedom to spend years studying "strange RNAs" and follow hunches about ancient molecular devices is what makes breakthroughs possible. "Scientists pursuing basic knowledge, basic understanding of biology, can make advances that can revolutionize fields and create new companies and new therapeutics,” Breaker says.

Anna Marie Pyle, PhD, Sterling Professor of Molecular, Cellular, and Developmental Biology in Yale’s Faculty of Arts and Sciences, has known Breaker since the early 90s and now teaches a seminar for advanced students with him. She says Breaker is "an amazingly effective leader in academia."

"It's not just the rigor of his work," she says. "He has a profoundly creative imagination for envisioning what's possible and designing experiments to understand it. He's not just a scientist. I like to think of Ron and people like him more like explorers. He's one of these people who has found fundamentally new knowledge about matter and about biology."

Breaker and his wife Michelle, a mathematics professor, have a rule: only one department chair in the house at a time. When Breaker was chair of the Department of Molecular, Cellular, and Developmental Biology for six years, Michelle couldn't be a chair. When she became chair for seven years, he stepped down. Now he's back, chairing his second Yale department.

They have two children, both in STEM fields. Their son designs aircraft engines and their daughter works at the Centers for Disease Control and Prevention, where she ran some of the first COVID tests during the pandemic. In fact, when she visits for holidays, she and her father speak "English microbiology," as Breaker calls it.

Before college, Breaker raised chickens. Not just any chickens. He was doing genetics, selecting for "fierce roosters and unusual plumage and odd-looking eggs." His father finally banned him from raising any more chickens because "they got a little bit strange," he says.

It's almost too perfect: The future biochemist still in high school, already pushing the boundaries of what biology could do, already willing to explore the unusual. Though the fierce rooster experiments are no more, the impulse that created them—the willingness to select for the unusual, to see what happens when you explore an unexpected direction—is still very much alive.

In going small, Breaker found something vast: ancient molecules that still control how our muscles work, how our neurons fire. It's a journey that required the persistence of someone who learned early that the cows need milking twice a day, no matter what.

Read more about Yale School of Medicine's groundbreaking RNA research.