Reduction of nemo-like kinase increases lysosome biogenesis and ameliorates TDP-43–related neurodegeneration
Leon Tejwani, … , Paymaan Jafar-Nejad, Janghoo Lim
Leon Tejwani, … , Paymaan Jafar-Nejad, Janghoo Lim
Published June 29, 2023
Citation Information: J Clin Invest. 2023;133(16):e138207. https://doi.org/10.1172/JCI138207.
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Research Article Genetics Neuroscience

Reduction of nemo-like kinase increases lysosome biogenesis and ameliorates TDP-43–related neurodegeneration

Abstract

Protein aggregation is a hallmark of many neurodegenerative disorders, including amyotrophic lateral sclerosis (ALS). Although mutations in TARDBP, encoding transactive response DNA-binding protein 43 kDa (TDP-43), account for less than 1% of all ALS cases, TDP-43–positive aggregates are present in nearly all ALS patients, including patients with sporadic ALS (sALS) or carrying other familial ALS–causing (fALS-causing) mutations. Interestingly, TDP-43 inclusions are also present in subsets of patients with frontotemporal dementia, Alzheimer’s disease, and Parkinson’s disease; therefore, methods of activating intracellular protein quality control machinery capable of clearing toxic cytoplasmic TDP-43 species may alleviate disease-related phenotypes. Here, we identify a function of nemo-like kinase (Nlk) as a negative regulator of lysosome biogenesis. Genetic or pharmacological reduction of Nlk increased lysosome formation and improved clearance of aggregated TDP-43. Furthermore, Nlk reduction ameliorated pathological, behavioral, and life span deficits in 2 distinct mouse models of TDP-43 proteinopathy. Because many toxic proteins can be cleared through the autophagy/lysosome pathway, targeted reduction of Nlk represents a potential approach to therapy development for multiple neurodegenerative disorders.

Authors

Leon Tejwani, Youngseob Jung, Hiroshi Kokubu, Sowmithra Sowmithra, Luhan Ni, Changwoo Lee, Benjamin Sanders, Paul J. Lee, Yangfei Xiang, Kimberly Luttik, Armand Soriano, Jennifer Yoon, Junhyun Park, Hannah H. Ro, Hyoungseok Ju, Clara Liao, Sofia Massaro Tieze, Frank Rigo, Paymaan Jafar-Nejad, Janghoo Lim

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Figure 3

Genetic reduction of Nlk reduces TDP levels in vitro.

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Genetic reduction of Nlk reduces TDP levels in vitro. (A and B) Western ...
(A and B) Western blots showing coexpression of Nlk-WT with GFP-tagged TDPWT or TDPA315T in NSC-34 cells (A) increased levels of TDP-35, while Nlk-KN did not. V-ATPase inhibitor BafA1 treatment similarly increased TDP-35 levels in the absence of Nlk overexpression to an even greater extent. Quantification shows TDP-35 levels in B (n = 4 biological replicates). (C and D) Western blots of sequentially extracted proteins based on detergent solubility from subcellular fractionated NSC-34 cells (C). Quantification of TDP-43 species shows Nlk WT overexpression increased cytosolic insoluble TDP-43 (sarkosyl and urea fractions) (D) (n = 3 biological replicates). LS, low salt; SK, sarkosyl; UR, urea. (E and F) Representative immunostaining of NSC-34 cells cotransfected with GFP-tagged TDP-43 and Nlk showing that WT Nlk increased formation of GFP-positive cytosolic TDP-43 aggregates (E), quantified in F. Nuclei are outlined with dotted lines (n = 4 biological replicates). (G and H) Western blots of cell lysates from WT or Nlk KO N2a cells transfected with GFP–TDP-43A315T (G), quantified in H (n = 12–24 replicates pooled from 4 individual experiments). Levels of exogenous GFP–TDP-43A315T, endogenous TDP-43, and truncated fragments of TDP-43 were significantly reduced. One-way ANOVA analyses (B, D, and F) or 2-tailed t tests (H) were performed to compare all listed genotypes/treatments unless otherwise noted, and data are represented as mean ± SEM. *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001.