In 2020, Juan Alfonzo, a professor of molecular biology, cell biology and biochemistry, began a project out of “pure COVID boredom.”
Now, that project may become the largest scientific undertaking since the Human Genome Project, according to Mukesh K. Jain, dean of medicine and biological sciences and Senior Vice President for Health Affairs.
Alfonzo, a world-renowned RNA biologist who came to Brown in 2023, aims to better understand RNA and its modifications. To do so, Alfonzo and his colleagues are working to lay the groundwork for sequencing all human RNA and its potential modifications as part of an ad hoc committee in the National Academies of Science, Engineering and Medicine, in which he is a member.
The NASEM committee’s stated objective is to “assess the scientific and technological breakthroughs, workforce and infrastructure needs to sequence and ultimately understand the roles RNA modifications play in biological processes and disease.”
The committee plans to highlight the best strategies for developing new RNA sequencing methods. Its findings will be released on Thursday, followed by several congressional briefings.
What does science say about RNA?
RNA, or ribonucleic acid, is a molecule that helps translate information from organisms’ genes into proteins, one of the major building blocks of all living things. RNA comes in several variants which all have specialized roles in cells.
Modifications to the chemical structure of RNA have become a major component of modern health innovation. COVID-19 messenger RNA vaccines, more commonly known as mRNA vaccines, rely on modified RNA. Alfonzo called these modifications a “game changer” for human health.
RNA is often modified in nature in order to function correctly. Incorrect or missing modifications can have serious impacts on the molecules’ functions, which make them an important subject in medicine.
As Alfonzo explains, “If you don’t know that a modification exists … you will never know what to target.”
Still, scientists don’t exactly understand the effects of all RNA modifications.
In the case of vaccines, “the RNA that they inject into you is modified” at several places, Alfonzo said, “but no one knows which (modification) is doing the trick.” That being said, scientists have proven that they are effective and safe, he noted.
Alfonzo says that the NASEM report will inform Congress that RNA sequencing research is of “sufficient relevance” and should be a national priority.
Where did the committee start?
On a Saturday in 2020, Alfonzo was “sitting at home, just doing nothing” when he received an email from Vivian Cheung, a professor at the University of Michigan who specializes in RNA research.
According to Alfonzo, Cheung asked if he, like her, was concerned by the lack of effective technologies available to map RNA and its modifications. In agreement, Alfonzo, Cheung and several colleagues published an article in Nature Genetics that reviewed the state of RNA mapping technologies and called for innovation in the field. Alfonzo attributed Cheung’s efforts as pivotal to the creation of the NASEM committee.
Currently, RNA can be indirectly sequenced through three main methods: sequencing by synthesis, mass spectrometry and nanopore sequencing. While sequencing by synthesis can be relatively accurate, it can only detect a small portion of RNA modifications. Mass spectrometry, which Alfonzo describes as the “gold standard,” is highly accurate but can only be used for small strands of RNA, making it “technically challenging.” Nanopore sequencing, a newer technique, can sequence long strands of RNA, but it is very error-prone.
As a result, the National Academies of Sciences committee set out to understand which existing technologies can be used to better sequence RNA and its modifications and how those technologies can fail. The group states that it will ultimately “develop a roadmap for achieving direct sequencing of modifications of RNA,” which would allow researchers to accurately unlock the RNA code.
The committee will also assess a wide variety of other research inputs for RNA analysis — such as computing capacity, data storage and public policy — to effectively evaluate the scientific, technological and infrastructure improvements needed to achieve RNA research goals. The report has been predominantly supported by funding from the Warren Alpert Foundation of Rhode Island and the National Institutes of Health, Jain explained.
At the same time, an international initiative known as the RNome Project sets out to map every RNA in the human body and all of its possible modifications. Its goal is to create a collection of molecules known as the epitranscriptome.
If the team achieves its goals, the catalog of the RNA epitranscriptome can help produce new diagnostic tools to detect disease, revolutionize our understanding of the human body and ultimately cure illnesses, said Jessica Brown, one of Alfonzo’s former graduate students and research collaborators.
According to a recent University press release, that process will likely be “bigger, more expensive and more daunting than the Human Genome Project.”
“You have to think big!” Alfonzo says.
The RNome Project’s first international meeting was held this January at Brown’s Warren Alpert Medical School and convened RNA experts from around the globe. The group set out to “strategize on key scientific issues, including which human cells should be sequenced first, the need for technology development, informatics and databases for RNA sequences and the need to establish RNA standards,” Jain noted.
Jessica Brown describes the RNome project as a “gigantic challenge,” noting that sequencing RNA modifications is particularly difficult. Because modifications can be added on to RNA in layers and each layer can interact with the preceding layer, scientists must be able to capture the modified RNA molecules at the correct points in time to understand the order in which modifications are added, she said.
Though the impacts of RNA research are well understood today, they were ignored for decades, says Alfonzo, who first became interested in RNA while studying at Indiana University, Bloomington. An undergraduate student from Venezuela, Alfonzo explained that studying science helped him with his immigration visa, enabling him to continue studying in the country. As a result, Alfonzo worked as a research technician in a former professor’s lab. There, he had his own project studying viruses and eventually began his PhD in the lab. It was at this time that Alfonzo met renowned RNA biologist Norman Pace, whose lab was in the same building as Alfonzo’s.
Alfonzo explains that Pace’s “larger-than-life personality” and passion for RNA got him excited about the field, leading him to undertake post-doctoral study on RNA editing in organisms called trypanosomes in Professor Larry Simpson’s lab at the University of California at Los Angeles. “That was really the beginning of RNA editing for me,” Alfonzo said.
At Simpson’s lab, Alfonzo came across a hypothesis that transfer RNA, known as tRNA, was modified by editing inside the cell. Alfonzo explained that in the early years of RNA research, RNA modifications were largely deemed “unimportant” by the scientific community. Though the tRNA modification hypothesis was “completely beyond the lab’s interest,” Alfonzo found it interesting and decided to test the theory while Simpson was on vacation. To Simpson’s disbelief, Alfonzo was able to prove the hypothesis and then completely switched his scientific focus to RNA.
Throughout his early years of research, Alfonzo recalled his colleagues saying that he was “wasting his time.. But when it was later found that mRNA modifications were associated with disease, the RNA modifications that Alfonzo studied “became the next big thing.” While human health is an important research application, Alfonzo said his own end goal for research is not to produce therapies, but rather to better understand RNA itself.
“Science builds on grains of sand,” Alfonzo said.
Alfonzo’s recruitment to Brown is part of the University’s broader push toward RNA research. He joined the University in 2023 to head the new Brown RNA Center. Having previously served as Director of The Ohio State University Center for RNA Biology, Alfonzo said he developed a vision for RNA research at Brown that involves attracting the top RNA researchers and facilitating collaborations between their research labs. When “you toss (scientists) in a bowl and mix them … they’ll come up with some good stuff!” he said.
“I like science to be collaborative,” he added. Reflecting on this interdisciplinary drive, Alfonzo’s own lab combines cell biology, cell imaging, molecular biology and biochemistry to achieve its research objectives.
While the OSU Center for RNA Biology housed around 45 RNA research labs, Alfonzo calculates that when he was recruited, Brown had roughly eight RNA research labs. Alfonzo said he thinks the University needs to build at least 14 RNA research labs to achieve its goals, and is in the process of bringing in six new faculty to Brown to achieve this.
Alfonzo envisions the center as an interdisciplinary hub for RNA research that “anybody who’s interested … can be part of” regardless of their background. At the center, Alfonzo hopes to attract researchers who study all aspects of RNA, not just modification, to develop comprehensive insights. The Center will initially operate out of Brown’s Dyer 225 building, which it will share with the University’s Center on the Biology of Aging.
Jessica Brown described Alfonzo’s creative, application-based teaching methods as “eye-opening,” adding that Alfonzo is “an outstanding scientist” and “definitely a well-picked person” for the job.
