One of the principal neuropathological features of Parkinson's disease and related dementia is the formation of abnormal clumps of a sticky protein called alpha-synuclein. These protein clumps build up in the brain causing the death of nerve cells, especially the dopamine-producing neurons that control movement. The progressive incorporation of alpha-synuclein into protein clumps is thought to result from impaired handling of this protein in cells.
Cells have developed efficient ways to rid themselves of unwanted proteins. However, the mechanism of destruction of alpha-synuclein has remained elusive. This area of research is challenging because multiple pathways have been implicated in alpha-synuclein turnover without definitive evidence that any of these operate in nerve cells.
Professor George Tofaris and his team have now developed tools to visualise in cells the fraction of alpha-synuclein that is degraded. They have shown that this involves a chain of ubiquitin (a small protein modifier found in most tissues). Typically, when a chain of ubiquitin molecules gets hitched to a protein - usually a damaged one - that protein becomes targeted for destruction.
Reporting in Science Advances, Zenko, Marsh et al. made ubiquitinated alpha-synuclein glow inside living cells using a method called bi-molecular fluorescence complementation. In this way they managed to follow its fate. They found that under normal conditions before alpha-synuclein clumps up, ubiquitination acts as a signal that targets it to intracellular organelles called endosomes. These in turn fuse with lysosomes, which are sacs of digestive enzymes, leading to the breakdown of alpha-synuclein. In contrast, when abnormal protein clumps form, they trap ubiquitinated alpha-synuclein with lysosomes, preventing efficient degradation.
Ubiquitination of alpha-synuclein triggers its endosomal localisation and sorting for degradation by the lysosome
The team used genetic and pharmacological approaches to decipher key factors that shuttle alpha-synuclein to the endosome for lysosomal degradation. Importantly, they produced novel antibodies and demonstrated that the pathway they delineated in living cells also operates endogenously in human nerve cells derived from induced pluripotent stem cells (iPSC) or primary neurons isolated from the mouse brain.
Professor Tofaris said: 'Lysosomes have been implicated in Parkinson's disease through genetics but their molecular link to alpha-synuclein turnover remained unclear. Our study delineates this pathway mechanistically and offers the tools for identifying genetic targets or small molecules that accelerate the destruction of alpha-synuclein'.