Dr. Jacob Ayers, the Borchelt lab & colleagues publish in Jan 2017 Journal of Neurochemistry

Ayers JI, McMahon B, Gill S, Lelie HL, Fromholt S, Brown H, Valentine JS, Whitelegge JP, Borchelt DR.   Relationship between mutant Cu/Zn superoxide dismutase 1 maturation and inclusion formation in cell models. J Neurochem 2017; 140(1):140-150.
A common property of Cu/Zn superoxide dismutase 1 (SOD1), harboring mutations associated with amyotrophic lateral sclerosis (ALS), is a high propensity to misfold and form abnormal aggregates. The aggregation of mutant SOD1 has been demonstrated in vitro, with purified proteins, in mouse models, in human tissues, and in cultured cell models. In vitro translation studies have determined that SOD1 with ALS mutations is slower to mature, and thus perhaps vulnerable to off-pathway folding that could generate aggregates. The aggregation of mutant SOD1 in living cells can be monitored by tagging the protein with fluorescent fluorophores. In the present study, we have taken advantage of the Dendra2 fluorophore technology in which excitation can be used to switch the output color from green to red, thereby clearly creating a time-stamp that distinguishes pre-existing and newly made proteins. In cells that transiently over-express the Ala 4 to Val variant of SOD1-Dendra2, we observed that newly made mutant SOD1 was rapidly captured by pathologic intracellular inclusions. In cell models of mutant SOD1 aggregation over-expressing untagged A4V-SOD1, we observed that immature forms of the protein, lacking a Cu co-factor and a normal intramolecular disulfide, persist for extended periods. Our findings fit with a model in which immature forms of mutant A4V-SOD1, including newly-made protein, are prone to misfolding and aggregation. This article is protected by copyright. All rights reserved.

This article is protected by copyright. All rights reserved.

KEYWORDS: Amyotrophic Lateral Sclerosis; aggregation; live-imaging; oxidation; superoxide dismutase 1