Scientists have unraveled the mystery behind the mechanism behind a type of protein that leads to a progressive nervous system disease.
In a study, recently published in the journal Physical Chemistry Chemical Physics, the researchers say that the RNA-binding ‘fused in sarcoma’ (FUS) protein transitions between liquid and solid phases inside cells.
The scientists from Japan explained in the study that this transition may lead to the formation of riboprotein granules that cause neurodegenerative diseases like familial amyotrophic lateral sclerosis (ALS), which affects nerve cells in the brain and spinal cord, causing loss of muscle control, the mechanisms of which remain unclear.
The researchers revealed what is behind FUS protein aggregation that leads to ALS. They also report that arginine suppresses the aggregation and could be a potential drug candidate for ALS-causing FUS.
FUS is a multifunctional RNA-binding protein that regulates transcription and mRNA processing inside cells. It is mainly present in the nucleus with traces in the cytoplasm. The researchers found that alteration of the FUS protein and its abnormal aggregation has been associated with the development of neurodegenerative diseases like familial ALS and frontotemporal lobar degeneration.
The scientists explained that FUS, like many other proteins, has been noted to undergo liquid-liquid phase separation (LLPS) both in artificial lab conditions and in its natural conditions in the body, to form liquid-like droplets containing ribonucleoprotein aggregates.
LLPS separates solutions of macromolecules like protein and RNA into two distinct liquid phases that have various biological functions. Although LLPS of FUS, leading to aberrant liquid-solid transition, is suggested to cause ALS, the underlying mechanism of the process remains unknown.
To this end, a group of researchers from the Ritsumeikan University have investigated the dynamics of the FUS-LLPS liquid condensates. The team led by Professor Ryo Kitahara discovered how FUS undergoes phase transition, and also identified a new therapeutic target for ALS.
Kitahara said: “The mechanism of liquid-to-solid phase transition of FUS must be understood in order to comprehend the onset of ALS and develop therapeutic agents targeting FUS.”
He added that previous studies have established that LLPS of FUS forms two distinct, reversible types of liquid condensates in equilibrium, the LP-LLPS (normal type) and the HP-LLPS (aberrant type), each with different partial molar volumes.
This information provided the team with a base to build upon. They examined the FUS solution using fluorescence recovery after photobleaching (FRAP), UV-Vis spectroscopy, and microscopy combined with hydrostatic pressure variations.
Aging of droplets
The researchers say the result of FRAP experiments provided some insights into the dynamics of FUS phase transitions. HP-LLPS accelerated the ‘aging of droplets’, which is the time-dependent transition of liquid condensates into solid aggregates.
Motivated by this discovery, the researchers used UV-Vis spectroscopy to investigate the reversibility of the phases. They found that the time-dependent formation of irreversible FUS aggregates was greater with HP-LLPS when compared to LP-LLPS.
They said that HP-LLPS promoted formation of irreversible solid aggregates with time, which could possibly be the reason behind the aging or degeneration of nerve cells.
The researchers said the results were the key to understanding the mystery behind liquid condensates transitioning to solid phase.
Kitahara added: “We strongly suggest that the formation of HP-LLPS via one-phase or LP-LLPS might be the physiological pathway underlying the aberrant liquid-to-solid conversion of FUS liquid condensates.”
Upon confirming that LLPS dynamics is the key to neurodegenerative disorders, the scientists wanted to search for potential therapeutic agents.
Since FUS-LLPS is governed by interactions between tyrosine and arginine residues, the team evaluated the effect of small molecules like arginine, dopamine, and pyrocatechol on LLPS and droplet formation. Interestingly, they found that all three suppressed HP-LLPS formation more strongly than LP-LLPS.
Kithara added: “We were thrilled to find out that arginine suppressed FUS aggregation even at very low concentrations. Nerve cells gradually age due to abnormal protein aggregation but can remain healthy up on arginine consumption. Therefore, arginine could be a plausible drug candidate for ALS-causing FUS.”
While arginine is already marketed as a supplement, its effects on ALS prevention and progression delay should be confirmed by future clinical trials.
Discussing their long-term goals, Kitahara said: “This study is the first to identify aberrant LLPS as a therapeutic target. We hope that our findings will not only help in identifying new treatments for ALS, but also stimulate further studies into the relevance of aberrant LLPS in various human diseases.”