Scientists say they have described for the first time atom-by-atom changes in a protein family linked to amyotrophic lateral sclerosis (ALS). Their study (“Mechanistic View of hnRNPA2 Low-Complexity Domain Structure, Interactions, and Phase Separation Altered by Mutation and Arginine Methylation”) is published in Molecular Cell.
“hnRNPA2, a component of RNA-processing membraneless organelles, forms inclusions when mutated in a syndrome characterized by the degeneration of neurons (bearing features of amyotrophic lateral sclerosis [ALS] and frontotemporal dementia), muscle, and bone. Here we provide a unified structural view of hnRNPA2 self-assembly, aggregation, and interaction and the distinct effects of small chemical changes—disease mutations and arginine methylation—on these assemblies,” write the investigators.
“The hnRNPA2 low-complexity (LC) domain is compact and intrinsically disordered as a monomer, retaining predominant disorder in a liquid-liquid phase-separated form. Disease mutations D290V and P298L induce aggregation by enhancing and extending, respectively, the aggregation-prone region. Co-aggregating in disease inclusions, hnRNPA2 LC directly interacts with and induces phase separation of TDP-43. Conversely, arginine methylation reduces hnRNPA2 phase separation, disrupting arginine-mediated contacts. These results highlight the mechanistic role of specific LC domain interactions and modifications conserved across many hnRNP family members but altered by aggregation-causing pathological mutations.”
The long-term goal is to target this cellular pathway with a drug or other therapy to prevent the disease, said the study's senior author, Nicolas Fawzi, Ph.D., at Brown University. “There is currently no therapy or cure for ALS and frontotemporal dementia. We are pursuing new hypotheses and angles to fight these illnesses.”
Many proteins connected with these diseases contain “low-complexity” domains or pieces, he explains. Compared to a cell's best-understood proteins, which are ordered and static in structure, low-complexity domains are disordered. Instead of a rigid shape, these pieces of protein are flexible and float inside cells until cued into action.
In nondisease situations, low-complexity domains help proteins perform healthy functions, including assembling into liquid-like droplets, where important cellular processes, such as RNA processing, take place, continues Dr. Fawzi.
When low-complexity domains go awry, as in disease, they transform into inclusions, intractable and accumulating knots, or clumps. In certain cancers, low-complexity domains are improperly attached to other proteins that may then incorrectly form droplets in cellular locations, leading to misregulated expression of genes, Dr. Fawzi says, adding that, “We're trying to understand why they change behavior and aggregate, and how we can disrupt those processes.”
In the study, the researchers describe the microscopic physical interactions and chemical changes of proteins associated with several cellular functions, including disease forms, and how still-healthy cells could try to temper it.
“We show how small chemical changes—involving only a few atoms—lead to big changes in assembly and disease-associated aggregation,” explains Dr. Fawzi. “These interactions are more dynamic and less specific than previously thought. A molecule does not take just one shape and bind to one shape, but a molecule is flexible and interacts in flexible ways.”
hnRNPA2 collects in membraneless organelles, where it may use its low-complexity domain to stick together, much in the way that water collects into droplets on the outside of a cold soda bottle on a humid summer day, as described by Dr. Fawzi. Until the publication of this study, several mechanistic details of how the low-complexity domain of hnRNPA2 worked and how it changed into aggregates in disease were unknown, he says.
Using nuclear magnetic resonance (NMR) spectroscopy, computer simulations, and microscopy, the researchers showed how disease mutations and arginine methylation, a functional modification common to a large family of proteins with low-complexity domains, altered the formation of the liquid droplets and their conversion to solid-like states in disease.
These findings explain several threads of research conducted over the last 20 years on the role of hnRNP family proteins in neuron function and neurodegeneration, notes Dr. Fawzi, who is an assistant professor in the department of molecular pharmacology, physiology, and biotechnology.
Previously, he and colleagues described the structure and biophysics of a related protein—how ALS-associated genetic flaws interfered with its proper function and behavior of another member of the protein family, causing it to aggregate. A separate study revealed a possible means of preventing those clumps from forming.
“Because these low-complexity domains are too flexible to be directly targeted by standard drugs, finding out how cells use and tame these domains is a potential route to stopping their unwanted assembly in disease,” Dr. Fawzi says.