Aggregation of amyloid proteins is believed to play a central role in many neurodegenerative diseases, including Parkinson’s disease. Now, a collaborative study by Indian researchers has explored certain key biophysical processes that are involved in the initial steps of this process, providing us with an important clue about the early stages of Parkinson’s disease progression.
In a recent study, scientists led by Samir K Maji from the Indian Institute of Technology (IIT) Bombay, Mumbai, working in collaboration with researchers from the National Centre of Biological Sciences, Bengaluru, Anna University, Chennai, and ETH Zürich, have mimicked the aggregation of a certain protein under laboratory conditions to understand early events in Parkinson’s disease – a progressive neurobiological disorder.
Parkinson’s disease is often characterised by tremors in the extremities. Other symptoms include restricted mobility, loss of body control, and stiffness. One of the major contributors to this disease are clumps of proteins called amyloids. These amyloids are formed within specific brain cells, leading to their deterioration and death. Amyloids are the leading cause of several neurodegenerative diseases, including Parkinson’s disease, Alzheimer’s disease, Huntington’s disease, and amyotrophic lateral sclerosis.
Maji’s team unravelled the early changes in protein structure and interactions that lead to amyloid protein accumulation in Parkinson’s disease. They found that these changes occur due to liquid-liquid phase separation (LLPS), the same phenomenon that prevents oil from mixing with water.
α‑Synuclein, a protein that can clump and form amyloids, is associated with Parkinson’s disease. It undergoes several structural and functional transitions during the progression of the disease. α‑synuclein is initially an unstructured protein that eventually takes a fibrous form, transitioning from a non-toxic protein to a toxic amyloid. However, the early events that cause α‑synuclein to aggregate were hitherto unknown.
Earlier studies have shown that biomolecules like proteins or nucleic acids often come together to form a concentrated liquid core separated from the surrounding liquid medium, somewhat like an oil droplet suspended in water. “We were curious to know whether liquid droplet formation is the initial mechanism for α‑synuclein aggregation and amyloid formation as several proteins form liquid droplets during the process of liquid-liquid phase separation (LLPS),” says Maji.
Maji and his team began by using in silico (computational) tools to check for the presence of certain characteristics like disordered and low complexity regions in the sequence of α‑synuclein. These characteristics usually indicate an ability to undergo liquid-liquid phase separation. Upon finding such regions, they experimentally induced α‑synuclein crowding under laboratory conditions, which led to the formation of liquid droplets. Probing the physical properties of the droplet revealed that the droplets became more rigid over time, suggesting the formation of protein aggregates.
The researchers followed this up with studies in the presence of Parkinson’s disease-promoting factors like metal ions, lipid membranes, and genetically mutated forms of α‑synuclein. These experiments showed that these factors accelerated the formation of liquid droplets and subsequent protein aggregation.
According to Maji, many proteins that are associated with the normal functions of cells undergo liquid-liquid phase separation. However, phase separation associated with specific disease often leads to a further transition from liquid droplet state to solid state, usually as a result of amyloid formation.
The researchers next performed biochemical, microscopic, and rigidity studies that confirmed that the droplets underwent liquid to solid transition and that this was due to the formation of amyloids within these droplets. The researchers also identified the specific domains in the protein that were responsible for the phase separation.
The present study may have significant implications for understanding the mechanism of how Parkinson’s disease-associated protein aggregation might initiate in cells. Aurnab Ghose, Associate Professor at Indian Institute of Science Education and Research (IISER) Pune, who was not associated with this study, points out the significance of this work. “It may be possible to leverage these observations to develop screening/early diagnosis strategies. This is not a paper that explains mechanisms of cellular toxicity of cell-cell transmission; instead, it focuses on biophysical mechanisms initiating the formation of aggregates,” says Ghose.