Fred Hutchinson Cancer Center RNA expert Arvind “Rasi” Subramaniam, PhD, has teamed up with investigators at the University of Pittsburgh to better understand microproteins, long overlooked short strings of amino acids. The group received a Transformative Research Award from the National Institutes of Health to uncover the roles that microproteins play in biology.
“We know microproteins exist. The question has been, do they have any function?” Subramaniam said. He and his collaborators will seek to answer this question, as well as determine whether microproteins are subject to the same biological rules that shape the evolution of other proteins.
He has joined Pittsburgh evolutionary biologist Anne-Ruxandra Carvunis, PhD, and immunologists Maninjay Atianand, PhD, and Alok Joglekar, PhD. The team hopes to reveal why microprotein genes evolve so quickly and whether this reflects a role in immunity and autoimmunity.
Part of the NIH Common Fund’s High-Risk, High-Reward Research Program, Transformative Research Awards are designed to support unconventional research with the potential to overturn or create new scientific paradigms. The team will receive $7.5 million over five years to investigate how microproteins may contribute to immunity and autoimmunity, as well as the biological rules governing their evolution. These insights could help scientists seeking new treatments for autoimmune diseases, such as multiple sclerosis, lupus and rheumatoid arthritis.
Microproteins have 100 amino acids or fewer. For a long time, scientists dismissed stretches of DNA that appeared to encode smaller proteins. To make proteins, molecular machines “read” DNA sequences and turn them into RNA molecules. Different molecular machines then “translate” the RNA molecules and create proteins based on information within the RNA. Scientists assumed that the information encoded in super-short segments of DNA never made its way into proteins.
“Larger than 100 amino acids: That was the definition of a protein,” said Subramaniam, who studies RNA translation. “Anything else was ignored.”
But about 15 years ago, technological advances that allowed scientists to get a more detailed picture of which RNAs were being turned into proteins upended assumptions about protein sizes. Researchers confirmed that thousands of microproteins are made across many species.
“People have shown specific instances where they seem to have roles in cancer, or development or the immune response," he said.
Now, scientists are working to discover what they do.
They have one major clue to follow: Microprotein genes flash in and out of existence within a single species — a blink of an eye in evolutionary terms.
“Microproteins are not super evolutionarily conserved,” Subramaniam said. “There are often multiple variants of them on a single RNA transcript and they don’t quite behave like normal proteins.”
Even closely related species usually don’t share microprotein genes, which can make it harder to pin down what they do. While scientists have revealed what a handful of individual microproteins do, they don’t have a big picture idea of microprotein function. However, Subramaniam and his colleagues believe the pace of microprotein evolution can guide their investigations.
Microproteins’ rapid appearance and disappearance hints at the forces shaping their existence. Quick evolution is a hallmark of genes and proteins locked in arms races with other molecules. Genetic adversaries must evolve quickly to sidestep their opponents. The immune system is an evolutionary arena in which protective immune molecules are clinched in an evolutionary wrestling match with fast-evolving microbes. The quick rate at which microproteins evolve suggests that some of them may play roles in the immune system, Subramaniam said.
The immune system may also shape microprotein evolution in a way that links the tiny peptides to autoimmunity, in which immune cells attack healthy tissue. Our immune system has evolved ways to build “tolerance” against normal tissue. Certain developing immune cells learn to distinguish between what is self and safe, and what is non-self (like a pathogenic virus) and unsafe. If these immature immune cells react too strongly to their environment — running the risk of attacking healthy tissue when they’re mature — they get killed off.
The immune cells that survive this weeding-out process “tolerate” their environment and only target microbes or diseased cells like cancer cells.
His Pittsburgh collaborators “were looking at whether microproteins are subject to the same rules of immune tolerance as normal proteins,” Subramaniam said. “For a protein to be functional in our body, it needs to not be acted against by the immune system — it needs to be recognized as self.”
In addition to testing whether microproteins help us fight off infection, the team will investigate whether microproteins are part of immune cells’ tolerance training, and if any are not recognized as self, and whether this means they contribute to autoimmunity. First, the scientists will create a catalog of microproteins which can be assayed for roles in immunity and autoimmunity.
“That’s where we come in,” said Subramaniam, who has led the development of high-throughput methods to study RNA translation and protein function. “We are good at doing these translation assays and figuring out what regions are translated, and then we have these high throughput assays where we do screens for antiviral activity and antibacterial activity. And then the idea is to put them together to try to figure out what are the large-scale principles of evolutionary selection [on microproteins].”
The investigators’ ideas are big and new — too new to have a foundation of solid research backing them up. This made their collaboration a perfect fit for the NIH’s Transformative Research Award, which was developed to fund exactly this kind of novel science.
“It’s a compelling idea that this evolutionary immunological nexus could provide an overarching framework to understand how microproteins arise and how they function,” Subramaniam said.
Sabrina Richards, a staff writer at Fred Hutchinson Cancer Center, has written about scientific research and the environment for The Scientist and OnEarth Magazine. She has a PhD in immunology from the University of Washington, an MA in journalism and an advanced certificate from the Science, Health and Environmental Reporting Program at New York University. Reach her at srichar2@fredhutch.org.
Read more about Fred Hutch achievements and accolades.
Every dollar counts. Please support lifesaving research today.
For the Media