Abstract
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Highlights
Neurodegenerative diseases, such as Parkinson’s disease, Amyotrophic Lateral Sclerosis, and Alzheimer’s disease, are characterized by the appearance of large protein aggregates in cells and in the extracellular space (Selkoe, 2004)
We engineered mammalian cell lines expressing Synphilin1 - a tracer of aggregates in Parkinson’s disease (Chung et al, 2001; Tanaka et al, 2004; Wakabayashi et al, 2000) - fused to a fluorescent protein Dendra2 (Chudakov et al, 2007)
Dendra2 is a green to red photo-convertible protein that enables photo-activation localization microscopy (PALM) (Betzig et al, 2006), a single-molecule based super-resolution (Betzig et al, 2006; Hess et al, 2006; Rust et al, 2006) approach we used previously to study protein clustering in mammalian cells (Cho et al, 2016; Cisse et al, 2013)
Summary
Neurodegenerative diseases, such as Parkinson’s disease, Amyotrophic Lateral Sclerosis, and Alzheimer’s disease, are characterized by the appearance of large protein aggregates in cells and in the extracellular space (Selkoe, 2004). The early steps of aggregate formation have been difficult to study, and may be critical to untangling the relationship between aggregation and disease. Late stages of aggregation can be measured both in vitro and in living cells, but the very early steps of aggregate formation remain elusive due to methodological limitations. The dynamics of nucleation and growth (Fink, 1998; Morris et al, 2009) of protein aggregates were most commonly measured experimentally in vitro (Buell et al, 2014; Jarrett and Lansbury, 1993; Krishnan and Lindquist, 2005; Lomakin et al, 1996; Serio et al, 2000). Aggregate growth dynamics have been studied in living cells, typically by seeding preformed ‘nuclei’ inside the living cells
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