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Cysteine cathepsins are essential in lysosomal degradation of ?-synuclein.

ABSTRACT: A cellular feature of Parkinson's disease is cytosolic accumulation and amyloid formation of ?-synuclein (?-syn), implicating a misregulation or impairment of protein degradation pathways involving the proteasome and lysosome. Within lysosomes, cathepsin D (CtsD), an aspartyl protease, is suggested to be the main protease for ?-syn clearance; however, the protease alone only generates amyloidogenic C terminal-truncated species (e.g., 1-94, 5-94), implying that other proteases and/or environmental factors are needed to facilitate degradation and to avoid ?-syn aggregation in vivo. Using liquid chromatography-mass spectrometry, to our knowledge, we report the first peptide cleavage map of the lysosomal degradation process of ?-syn. Studies of purified mouse brain and liver lysosomal extracts and individual human cathepsins demonstrate a direct involvement of cysteine cathepsin B (CtsB) and L (CtsL). Both CtsB and CtsL cleave ?-syn within its amyloid region and circumvent fibril formation. For CtsD, only in the presence of anionic phospholipids can this protease cleave throughout the ?-syn sequence, suggesting that phospholipids are crucial for its activity. Taken together, an interplay exists between ?-syn conformation and cathepsin activity with CtsL as the most efficient under the conditions examined. Notably, we discovered that CtsL efficiently degrades ?-syn amyloid fibrils, which by definition are resistant to broad spectrum proteases. This work implicates CtsB and CtsL as essential in ?-syn lysosomal degradation, establishing groundwork to explore mechanisms to enhance their cellular activity and levels as a potential strategy for clearance of ?-syn.

SUBMITTER: McGlinchey RP

PROVIDER: S-EPMC4522768 | biostudies-literature | 2015 Jul

REPOSITORIES: biostudies-literature

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Cysteine cathepsins are essential in lysosomal degradation of α-synuclein.

McGlinchey Ryan P RP Lee Jennifer C JC

Proceedings of the National Academy of Sciences of the United States of America 20150713 30

A cellular feature of Parkinson's disease is cytosolic accumulation and amyloid formation of α-synuclein (α-syn), implicating a misregulation or impairment of protein degradation pathways involving the proteasome and lysosome. Within lysosomes, cathepsin D (CtsD), an aspartyl protease, is suggested to be the main protease for α-syn clearance; however, the protease alone only generates amyloidogenic C terminal-truncated species (e.g., 1-94, 5-94), implying that other proteases and/or environmenta ...[more]

PMID: 26170293

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Project description:BackgroundAutosomal dominant mutations in α-synuclein, TDP-43 and tau are thought to predispose to neurodegeneration by enhancing protein aggregation. While a subset of α-synuclein, TDP-43 and tau mutations has been shown to increase the structural propensity of these proteins toward self-association, rates of aggregation are also highly dependent on protein steady state concentrations, which are in large part regulated by their rates of lysosomal degradation. Previous studies have shown that lysosomal proteases operate precisely and not indiscriminately, cleaving their substrates at very specific linear amino acid sequences. With this knowledge, we hypothesized that certain coding mutations in α-synuclein, TDP-43 and tau may lead to increased protein steady state concentrations and eventual aggregation by an alternative mechanism, that is, through disrupting lysosomal protease cleavage recognition motifs and subsequently conferring protease resistance to these proteins.ResultsTo test this possibility, we first generated comprehensive proteolysis maps containing all of the potential lysosomal protease cleavage sites for α-synuclein, TDP-43 and tau. In silico analyses of these maps indicated that certain mutations would diminish cathepsin cleavage, a prediction we confirmed utilizing in vitro protease assays. We then validated these findings in cell models and induced neurons, demonstrating that mutant forms of α-synuclein, TDP-43 and tau are degraded less efficiently than wild type despite being imported into lysosomes at similar rates.ConclusionsTogether, this study provides evidence that pathogenic mutations in the N-terminal domain of α-synuclein (G51D, A53T), low complexity domain of TDP-43 (A315T, Q331K, M337V) and R1 and R2 domains of tau (K257T, N279K, S305N) directly impair their own lysosomal degradation, altering protein homeostasis and increasing cellular protein concentrations by extending the degradation half-lives of these proteins. These results also point to novel, shared, alternative mechanism by which different forms of neurodegeneration, including synucleinopathies, TDP-43 proteinopathies and tauopathies, may arise. Importantly, they also provide a roadmap for how the upregulation of particular lysosomal proteases could be targeted as potential therapeutics for human neurodegenerative disease.

Dataset Information

Cysteine cathepsins are essential in lysosomal degradation of ?-synuclein.

Publications

Cysteine cathepsins are essential in lysosomal degradation of α-synuclein.

Similar Datasets

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