The destructive proteins in Parkinson’s and Huntington’s disease
Parkinson’s and Alzheimer’s disease both involve accumulation of misfolded proteins that join together to form soluble protein aggregates and then insoluble fibrils. Highly debilitating, there is no disease-modifying treatment to alleviate patients’ suffering. However, for obvious societal reasons there is intense interest in the field and it is increasingly accepted that the fibrous amyloid state is inert while the soluble oligomeric proteins are the cytotoxic species. The TOPIC project has trained two ESRs to work with these oligomers, which undergo a series of unwanted interactions with proteins and membranes. ‘Despite the vast interest in oligomers, very little is really known about their biochemical interactions apart from the fact that they ultimately kill the cell. We wanted to develop tools to get to know more about these activities. This could pave the way for therapy eventually,’ explains TOPIC coordinator Prof. Daniel Otzen of Aarhus University. Identification of cytotoxic molecular interactions in Parkinson’s The ESRs focused on their own cytotoxic oligomers characteristic of, and associated with, a particular neurodegenerative disease. Their research ran on parallel tracks to achieve synergy in methodology and data analysis. The Parkinson’s disease “culprit” is the protein alpha-synuclein which is relatively easy to work with and forms oligomers spontaneously. However, the oligomer can dissociate again, so a major advance was to introduce a strategy to stabilise the oligomer to allow it to persist in the cell. Once this had been achieved, it was possible to combine immunoprecipitation and mass spectrometry to identify other proteins with which it formed stable complexes in a cellular context. TOPIC therefore established routines for reliable preparation of alpha-synuclein oligomers (alphaSOs) and their impact on the fibrillation pathway of the alphaSO has been described in detail. Significantly, these stabilised oligomers have been observed to bind to one type of primary neuron in culture. However, the researchers are still working to validate the protein partners to which the oligomers attach on the neurons. Formation of stable tau oligomers in Alzheimer’s Another protein, tau, is a key player in Alzheimer’s Disease and it is more tricky to prepare stable oligomers of this protein. However, a new approach was optimised to use naturally occurring sugar complexes (glycosaminoglycans) to oligomerise tau. The next step will be to identify binding proteins in the cell using the same strategy as for alpha-synuclein. Although outside the scope of the present TOPIC project, the ESRs were aware of the long-term goal – to develop small molecule inhibitors to prevent the cytotoxic interactions that fuel disease progression. ‘Once we have identified which protein-oligomer interactions are critical for the development of cytotoxicity, it should be possible to design drug molecules that block these interactions and thus halt pathology.’ Response to the challenges of molecular research Research into protein interaction at the molecular level is full of challenges and the TOPIC researchers had to deal with poor protein expression, inability to control reaction rate and problems with oligomer identification and isolation. As a result, research focus was transferred mid-project from Huntington’s (the protein huntingtin) and Diabetes Type 2 (the peptide amylin) to tau, the protein linked with Parkinson’s and Alzheimer’s. Still the main avenue of research, key results are expected in the near future to see if the tau oligomer can be stabilised to work on. The future for small molecule intervention TOPIC project researchers have provided the neurodegenerative disease research arena with a wealth of data on stabilisation of misfolded proteins that are potentially involved in disease progression, in particular their involvement in cytotoxic reactions. Identification of the binding partners of cytotoxic oligomers and the resulting biological consequences could well pinpoint appropriate therapeutic targets. Development of small molecule disruptors to prevent these damaging reactions is the ultimate goal. ‘TOPIC has demonstrated that it is indeed possible to stabilise oligomers and use them to identify cellular binding partners, paving the way for further cellular analyses to pinpoint weak spots in the cytotoxicity process,’ says Prof. Otzen, summing up the massive societal consequences of successful treatment of neurodegenerative diseases such as Parkinson’s.
Keywords
TOPIC, protein, oligomer, cytotoxic, Huntington’s, Parkinson’s, small molecule, fibril
