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PhaseAge
Defective ribonucleoprotein (RNP) granules are thought to be a major factor in the development of amyotrophic lateral sclerosis (ALS) and other forms of neurodegenerative disease, while they are also associated with a number of other conditions. We spoke to Professor Simon Alberti about his work in investigating how these granules form and how they cause disease.
Molecular aging of a prion-like RNA-binding protein from a liquid to a solid, aggregated state.
How do RNP granules cause disease?
The aggregation of mis-folded proteins in the body is known to be a major factor in the development of certain neurodegenerative diseases, such as amyotrophic lateral sclerosis (ALS). While younger people are able to essentially clear these mis-folded proteins, this ability declines as we age, which can eventually lead to serious health problems. “The protein quality control (PQC) machinery controls the status of these proteins and keeps them in check. We think that starts to fail as people age,” explains Simon Alberti, Professor of Cellular Biochemistry at TU Dresden. As the Principal Investigator of the PhaseAge project, Professor Alberti is investigating the factors behind this gradual loss of function, looking in particular at the role of ribonucleoprotein (RNP) granules. “These RNP granules are considered to be membraneless organelles as they contain a lot of different factors, such as proteins and RNAs, and they have no membranes around them.” he says. Many different conditions are associated with defective RNP granules, including not just neurodegenerative diseases, but also a number of other disorders, underlining the wider importance of Professor Alberti’s research. These RNP granules become defective when they undergo a transition from a liquid-like state into an aberrant, more solid state. “The important point with this transition is that the dynamics change. The proteins are essentially trapped inside, and there’s little exchange with the outside,” outlines Professor Alberti. The wider aim of his research is to understand the molecular mechanism behind both the formation of these RNP granules and their transition into an aberrant state. “We want to understand the molecular principles of how these granules form, and how they turn into something that can cause disease. And, most importantly, can we then find factors that can prevent these transitions?” says Professor Alberti.
RNP granules The first step towards this wider goal is to understand how these RNP granules are initially formed inside a cell, with researchers looking at the role of proteins in this process. It is thought that initially RNA-binding proteins and other factors assemble into these membraneless organelles, or RNP granules, within a cell. “At first the properties are maintained, but as we age, we lose the ability to control the material properties of these structures,” says Professor Alberti. By analysing single cells, Professor Alberti and his colleagues hope to gain deeper insights into how these RNP granules are formed, how they age, and how this may affect disease development. “We are looking at healthy cells, while we also mimic ageing in cells. Essentially, we stress the cells, as we know that ageing very often increases stress,” he outlines. “We also purify RNA-binding proteins, then using these proteins we try to put together these structures that normally we only see in cells.”
This approach allows researchers to investigate the underlying factors which may affect the formation and overall development of RNP granules. These RNP granules contain information, messenger RNAs that are read in neurons. “Neurons have very complex morphologies. RNP granules are frequently found in neurons, so neurons are very vulnerable to the problems that can arise with these RNP granules,” explains Professor Alberti. Researchers now have the ability to build RNP granules in a test tube, based on the principle of phase separation, a recently described process which is thought to play a major role in the formation of membraneless organelles. “Phase separation is really a new phenomenon that was only described a couple of years ago,” continues Professor Alberti. “Previously we didn’t really understand how large membraneless organelles formed, we didn’t have a concept of how this could work.” A membraneless organelle is thought to be formed through a physical process that can be loosely compared to mixing oil and vinegar. When mixed, oil and vinegar will separate, and Professor Alberti believes that biomolecules work in a similar way in the formation of a membraneless organelle. “That’s how a membraneless organelle is formed. Phase separation is an increasingly important concept in cell biology, and it’s the focus of a lot of attention in research,” he
says. It is now possible to build RNP granules in the test tube, based on this principle of phase separation, from which researchers hope to gain deeper insights which could open up further avenues of investigation. “We’re pretty good at forming these RNP granules. The next step is to look at the transition from these liquid-like, physiological states, to these aberrant, more solid-like states, and
results have been gained. “We have found a few proteins that completely prevent these modifications when you add them to a liquid reconstituted system. So they keep these granules in a liquid state,” outlines Professor Alberti. “It could be interesting to target these with a drug and maybe up-regulate them, as they seem to prevent the transition into an aberrant state.”
we’ve achieved this already to some degree,” explains Professor Alberti
Researchers have identified several important protein components during the course of this work, and are also looking at the possibility of preventing the formation of these aberrant, solid-like states. One part of this work involves looking for drugs that would dissolve these membraneless organelles, and Professor Alberti says there have been some promising results in this area. “We have found a few interesting drug candidates,” he says. Another part of the project’s research centres on investigating candidate proteins and the PQC machinery, and again some interesting
PCQ machinery The aim with a drug would be to essentially enhance the PQC machinery and strengthen it in some way, so as to prevent the formation of defective RNP granules. It is therefore important to identify the components of the PQC machinery, which is a major priority in Professor Alberti’s research. “The PQC machinery is a large system, it has many components. We’re trying to figure out which of those components are important for the control of RNP granules, then in future we can look to strengthen them,” he explains. While this is an exciting prospect, opening up the possibility of improved treatment of neurodegenerative
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disease, the focus in research at this stage is more on understanding the basic mechanisms involved in the formation of defective RNP granules. “The molecular mechanisms behind the formation of these structures and how they become defective has not yet been described,” says Professor Alberti.
This remains the focus of attention in research, yet Professor Alberti is also aware of the wider picture beyond the project’s immediate agenda. Defective RNP granules are associated with a wide variety of conditions, including schizophrenia, autism and many other examples. “Many different brain and developmental diseases are associated with defective RNP granules,” stresses Professor Alberti. Evidence from the brains of patients with ALS shows that changes in RNP granule properties are an important consideration in terms of the development and progression of these diseases. “We often see these RNAcontaining aggregates in stressed neurons, or clumps of protein and RNAs. We also find a certain set of proteins clumped in diseased cells,” continues Professor Alberti. “These proteins stick together and adopt abnormal structures. These structures are very difficult to dispose of – they could be toxic and eventually kill the cell.”
With evidence mounting of the involvement of RNP granules in different types of disease, Professor Alberti is keen to continue his
work in this direction. “I’m in touch with many other researchers, as I think there is a lot of potential to look at other diseases, to investigate what’s happening there and what’s going on,” he says. “Can we then again identify the key factors? Will it be possible to identify targets for therapeutics later on?”
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PhaseAge The chemistry and physics of RNP granules: how they form, age and cause disease Project Objectives Many debilitating age-related diseases affect the nervous system but the molecular causes of these diseases have remained largely unknown. In recent years, deficiencies in RNA metabolism mediated by RNA-binding proteins (RBPs) have come into the focus. In PhaseAge, researchers; • Investigate the molecular mechanisms of RNP granule formation • Study the molecular events that lead to aberrant RNPs, focusing on disease-associated mutations, changes in environmental conditions, post-translational modifications and molecules with fluidizing or solidifying effects • Define the mechanisms of RNP granule quality control, which prevent aberrant phase transitions or reverse RNP granule aggregates to their normal state Project Funding Funded by an ERC Consolidator Grant (PhaseAge) and JPND (CureALS) Project Partners • Anthony Hyman MPI-CBG Dresden • Jared Sterneckert CRTD Dresden • Rohit Pappu and Alex Holehouse Washington University, USA) • Serena Carra, University of Modena
Contact Details Simon Alberti Professor of Cellular Biochemistry Technische Universität Dresden Center for Molecular and Cellular Bioengineering (CMCB) Biotechnology Center (BIOTEC) Tatzberg 47/49, 01307 Dresden, Germany T: +49 351 46340236 E: Simon.Alberti@tu-dresden.de W: http://www.biotec.tu-dresden.de/ research/alberti.html
Alberti S, Dormann D (2019). Liquid-liquid phase separation in disease (2019). Annual Reviews in Genetics, doi: 10.1146/annurev-genet-112618-043527.
Alberti S, Gladfelter A, Mittag T (2019). Considerations and Challenges in Studying Liquid-Liquid Phase Separation and Biomolecular Condensates. Cell, 176(3):419-434. doi: 10.1016/j.cell.2018.12.035.
Professor Simon Alberti
Simon Alberti is a Professor of Cellular Biochemistry at BIOTEC, Technische Universität Dresden in Germany. He received his PhD in Cell Biology from the University of Bonn in 2004 and has since held research positions in Europe and America. His recent work shows that cells form many membraneless compartments via a biophysical process known as phase separation.
