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Loss of MAGEL2 in Prader-Willi syndrome leads to decreased secretory granule and neuropeptide production
Helen Chen, … , Lawrence T. Reiter, Patrick Ryan Potts
Helen Chen, … , Lawrence T. Reiter, Patrick Ryan Potts
Published September 3, 2020
Citation Information: JCI Insight. 2020;5(17):e138576. https://doi.org/10.1172/jci.insight.138576.
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Research Article Cell biology Neuroscience

Loss of MAGEL2 in Prader-Willi syndrome leads to decreased secretory granule and neuropeptide production

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Abstract

Prader-Willi syndrome (PWS) is a developmental disorder caused by loss of maternally imprinted genes on 15q11-q13, including melanoma antigen gene family member L2 (MAGEL2). The clinical phenotypes of PWS suggest impaired hypothalamic neuroendocrine function; however, the exact cellular defects are unknown. Here, we report deficits in secretory granule (SG) abundance and bioactive neuropeptide production upon loss of MAGEL2 in humans and mice. Unbiased proteomic analysis of Magel2pΔ/m+ mice revealed a reduction in components of SG in the hypothalamus that was confirmed in 2 PWS patient–derived neuronal cell models. Mechanistically, we show that proper endosomal trafficking by the MAGEL2-regulated WASH complex is required to prevent aberrant lysosomal degradation of SG proteins and reduction of mature SG abundance. Importantly, loss of MAGEL2 in mice, NGN2-induced neurons, and human patients led to reduced neuropeptide production. Thus, MAGEL2 plays an important role in hypothalamic neuroendocrine function, and cellular defects in this pathway may contribute to PWS disease etiology. Moreover, these findings suggest unanticipated approaches for therapeutic intervention.

Authors

Helen Chen, A. Kaitlyn Victor, Jonathon Klein, Klementina Fon Tacer, Derek J.C. Tai, Celine de Esch, Alexander Nuttle, Jamshid Temirov, Lisa C. Burnett, Michael Rosenbaum, Yiying Zhang, Li Ding, James J. Moresco, Jolene K. Diedrich, John R. Yates III, Heather S. Tillman, Rudolph L. Leibel, Michael E. Talkowski, Daniel D. Billadeau, Lawrence T. Reiter, Patrick Ryan Potts

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Figure 4

DPSC-derived neurons from PWS patients have impaired endosome-mediated protein trafficking and decreased SG protein abundance.

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DPSC-derived neurons from PWS patients have impaired endosome-mediated p...
(A) Representative images of immunofluorescence staining of M6PR in DPSC-derived neurons. Proper M6PR trafficking shows clustered juxtanuclear localization. (B) Proportion of cells with impaired M6PR trafficking is significantly increased in PWS and SYS DPSC-derived neurons. Each data point represents 1 individual, plotted as mean ± SD; more than 75 cells/data point, analyzed by 1-way ANOVA. (C) Representative images of immunofluorescence staining of integrin α5 in DPSC-derived neurons. Yellow arrows indicate increased intracellular (endosomal) pools. (D) Quantification of Western blot analysis showed reduced expression of integrin α5, αV, and β1 in PWS deletion DPSC-derived neurons. Each target protein is first normalized to GAPDH, then normalized to averaged control. Each data point represents 1 individual (n = 6), plotted as mean ± SD and analyzed by unpaired, 2-tailed t test. (E) Representative images of immunofluorescence staining of phalloidin on VPS35-marked endosomes in DPSC-derived neurons. (F) Fluorescence intensity of F-actin on VPS35-marked endosomes is significantly decreased in PWS and SYS DPSC-derived neurons. Each data point represents 1 individual, plotted as mean ± SD; more than 75 cells/data point, analyzed by 1-way ANOVA. (G) Fluorescence intensity of ArpC5 on VPS35-marked endosomes is significantly decreased in PWS and SYS DPSC-derived neurons. Each data point represents 1 individual, plotted as mean ± SD; more than 75 cells/data point, analyzed by 1-way ANOVA. (H) Western blot analysis of M6PR, α5, αV, and β5 between control and PWS iN. GAPDH served as loading control. (I) Quantification of Western blot analysis showed reduced expression of PCSK1, PCSK2, and CHGB in PWS deletion and SYS DPSC-derived neurons. Each target protein is first normalized to GAPDH, then normalized to averaged control. Each data point represents 1 individual, plotted as mean ± SD and analyzed by unpaired, 2-tailed t test. *P < 0.05 and **P < 0.01. Scale bars: 10 μm.
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