N-acetyl-l-leucine lowers α-synuclein levels and improves synaptic function in Parkinson’s disease models
Pingping Song, Chuyu Chen, Rossella Franchini, Bryan Duong, Yi-Zhi Wang, Robert Coukos, Zhong Xie, Jeffrey N. Savas, Yueqin Zhou, Mariarita Bertoldi, D. James Surmeier, Loukia Parisiadou, Dimitri Krainc
Pingping Song, Chuyu Chen, Rossella Franchini, Bryan Duong, Yi-Zhi Wang, Robert Coukos, Zhong Xie, Jeffrey N. Savas, Yueqin Zhou, Mariarita Bertoldi, D. James Surmeier, Loukia Parisiadou, Dimitri Krainc
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Research Article Cell biology Neuroscience

N-acetyl-l-leucine lowers α-synuclein levels and improves synaptic function in Parkinson’s disease models

Abstract

N-acetyl-l-leucine (NALL), a derivative of the branched-chain amino acid leucine, has shown therapeutic potential for neurodegenerative diseases, including in prodromal stages of Parkinson’s disease (PD). However, the mechanism of its protective effects has been largely unknown. Using human induced pluripotent stem cell–derived dopaminergic neurons from patients carrying GBA1, LRRK2, or VPS35 mutations, as well as from sporadic PD cases, we found that NALL treatment markedly reduced Ser129 phosphorylated α-synuclein (pS129-syn). Discovery-based proteomic analysis revealed that NALL treatment upregulated lysosomal, mitochondrial, and synaptic proteins without inducing cytotoxicity. The reduction of pS129-syn was dependent on serine protease HTRA1, which was robustly induced by NALL. Moreover, NALL increased the expression of wild-type parkin in mutant dopaminergic neurons, leading to increased glycosylated dopamine transporter, elevated synaptic membrane-associated synaptojanin-1, and accelerated synaptic vesicle endocytosis, suggesting improved synaptic function. Furthermore, in LRRK2R1441C knockin mice, NALL administration decreased pS129-syn, elevated parkin levels, and ameliorated dopamine-dependent motor learning deficits. These findings highlight the therapeutic potential of NALL for PD by its protective effects on α-synuclein pathology and synaptic function in vulnerable dopaminergic neurons.

Authors

Pingping Song, Chuyu Chen, Rossella Franchini, Bryan Duong, Yi-Zhi Wang, Robert Coukos, Zhong Xie, Jeffrey N. Savas, Yueqin Zhou, Mariarita Bertoldi, D. James Surmeier, Loukia Parisiadou, Dimitri Krainc

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Figure 3

NALL increases functional dopamine transporter and synaptic function through parkin.

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NALL increases functional dopamine transporter and synaptic function thr...
(A and B) Representative Western blot (A) and quantification (B) of glycosylated and nonglycosylated dopamine transporter (DAT) in GBA1 L444P mutant dopaminergic neurons treated with increasing NALL concentrations. (C and D) Western blot (C) and quantification (D) of parkin expression in NALL-treated GBA1 L444P neurons. (E–J) Western blots and corresponding quantification of SYNJ1 redistribution from cytosolic (Cyto) to synaptic membrane (SM) fractions following NALL treatment in GBA1 L444P (E and F), GBA1 N370S (G and H), and parkin mutant (I and J) dopaminergic neurons. Synaptophysin (SYP) and GAPDH served as synaptic and cytosolic markers, respectively. (K) Scatter plot showing time constants of pHluorin fluorescence recovery following exocytosis in GBA1 L444P neurons. NALL-treated synapses (105.3 ± 5.8 s; n = 59 ROIs) recovered significantly faster than NT (133.2 ± 5.3 s; n = 89 ROIs) (Mann-Whitney test). Lines represent means ± SD. (L) Representative traces of pHluorin fluorescence intensity from 50 s before to 300 s after exocytosis in GBA1 L444P mutant neurons. For Western blots, β-III-tubulin and GAPDH served as loading controls (n = 3 independent experiments). Data are expressed as mean fold-change relative to 0 mM (DMSO) or nontreated (NT) groups. Statistical significance was determined by 1-way ANOVA (B and D), Student’s t test (F, H, and J), or Mann-Whitney test (K). Data represent mean ± SEM, except in K (mean ± SD). *P < 0.05, **P < 0.01, ***P < 0.005, and ****P < 0.001.