Protein clumps puncture brain cells
Danish researchers have uncovered a toxic mechanism that may lie behind Parkinson’s disease. With a newly developed test platform, they have for the first time shown how clumps of toxic protein can puncture brain cells and drain them of vital substances.
By the time the first trembling hand movement reveals a new case of Parkinson’s disease, the chronic condition has already been active for a long time. Brain cell after brain cell has been attacked and killed. And the “villain” is toxic protein accumulations in the brain cells. That much is well known. The mechanism behind it, however, has until now been unknown. But now, a team of researchers from iNANO at Aarhus University has for the first time shown how some of the protein clumps can kill brain cells by forming holes in them, draining their contents.
“There are some very fundamental things we don’t know about this subject, which I am truly passionate about and would like to explore further. And I believe I can contribute with my biophysical perspective and some slightly different questions than those asked by doctors and molecular biologists in this field of research.”t.”
“We have for the first time shown how some of the toxic protein clumps can attach themselves to the brain cell’s protective membrane and form small pores or holes in the membrane,” explains postdoc Mette Galsgaard Malle, senior author of the study.
The discoveries were made using a new advanced test platform with cell models, which also made it possible to record a video of three of the “cells” under attack.
The research is supported by the Lundbeck Foundation, and the study is published in the renowned journal ACS Nano, issued by the American Chemical Society.
A Surprising Discovery
Parkinson’s disease is a progressive chronic disease that breaks down nerve cells in the brain, primarily those that produce dopamine. This leads to motor and cognitive impairments. According to WHO, twice as many people suffer from the disease today compared to 25 years ago. There is currently no cure, only symptomatic treatment.
The disease mechanisms are only partially mapped, but one important factor is thought to be the protein alpha-synuclein, which clumps together in the brain’s nerve cells (neurons) and destroys them. Normally, alpha-synuclein is involved in regulating dopamine and other neurotransmitters, but in Parkinson’s disease, the proteins begin sticking together in various small or larger clumps. The small clumps are called oligomers, and it is these that the researchers have studied.
“We don’t have many studies of oligomers, even though they are considered the most toxic form of alpha-synuclein because they can disrupt membranes and also synapses – the contact points where cells communicate with each other,” explains Mette Galsgaard Malle.
A prevailing theory about oligomers is that they create holes in cells, and it is this theory that the Danish researchers have now supported for the first time in an experiment. At the same time, they made another – surprising – discovery.
The expectation was that the protein clumps would attach to the cell membrane, make a hole, and then the membrane would break. But the researchers found that oligomers attack the cell in multiple rounds and in three stages: they alternate between binding to the membrane’s surface and drilling halfway or completely into the membrane. And the holes are not static, as expected, but open and close rapidly depending on the membrane’s composition, electrical charge, and the molecules that bind to the protein clumps. So even though the membrane is punctured, it remains intact for a longer time before the cell is drained of its contents and dies.
Bo Volf Brøchner, PhD student and first author of the study, describes the discovery:
“It happened many times that the oligomers made holes and then pulled back again. In one case, we counted 37 times. It really surprised us that it was such a dynamic process.”
He believes this may explain why cells don’t die immediately but become dysfunctional before they perish.
“This is an important discovery for understanding why and how these protein clumps are so toxic to our cells,” says Mette Galsgaard Malle.
Platform Reveals Details and Variations
To investigate the effect of oligomers on cell membranes at an unprecedented level of detail, the researchers developed an entirely new test platform with thousands of liposomes – small fat bubbles that serve as cell models – placed on a glass plate.
The artificial cells can be constructed in varying shapes and sizes, and the researchers can fill them with different substances, as well as add compounds to the membranes’ surfaces. Using advanced microscopy, measurements of electrical signals across the membranes, and specially developed computer programs, the researchers can study and analyze how membranes and substances interact under different conditions. Crucially, they can explore the “cells” one at a time and in real time.
“We capture all the variations and can see how each individual molecule interacts, instead of looking at a billion cells at once and working with averages, as is traditionally done with other methods,” explains Mette Galsgaard Malle.
In the specific experiment, the researchers got bacteria to produce alpha-synuclein and then induced the proteins to clump together as oligomers.
They filled some of the artificial cells with fluorescent dye and added oligomers to the membranes. This allowed them to uncover which membranes the oligomers bound to most strongly, and how they created holes in them. In video recordings, they could after a while see dye leaking out of some of the liposomes as a result of the oligomers forming holes in the membranes.
It turned out that oligomers prefer some membranes over others.
“Strangely enough, they most readily bind to the most curved membranes, but they are best at making holes in the larger, flatter membranes. And that’s important to know, because we want to take a step closer to stopping this negative process,” explains Mette Galsgaard Malle.
Another important discovery is that oligomers prefer membranes resembling those around mitochondria – the so-called powerhouses of the cells.
The researchers believe this may indicate that the damage starts there.
In total, the researchers screened 500,000 individual interactions between liposomes and oligomers.
The next step is to test the results in cell cultures.
“Now we need to see if the results from the platform can be transferred to more complex biological systems,” says Mette Galsgaard Malle.
In principle, the platform could serve as a springboard for testing many other substances’ interactions with membranes. And Mette Galsgaard Malle already has plans to further develop the platform and is actively applying for funding for new projects.
“This platform opens up so many new possibilities. For example, studying other toxic mechanisms in Parkinson’s disease and in other neurodegenerative diseases such as Alzheimer’s, where protein accumulation also plays a role.”
