Delivering progranulin to neuronal lysosomes protects against excitotoxicity

Davis, Skylar E.; Roth, Jonathan R.; Aljabi, Qays; Hakim, Ahmad R.; Savell, Katherine E.; Day, Jeremy J.; Arrant, Andrew E.

doi:10.1016/j.jbc.2021.100993

Cited by 17 publications

(13 citation statements)

References 98 publications

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“…hiPSC-derived neurons were seeded at a density of 400,000 cells/well of a Matrigel coated 48-well plate. After 24 h, cells were washed once with DPBS, treated with complete media containing either 10 µg/ml DQ Red BSA or vehicle (PBS) and incubated for 6 h or 24 h [ 16 , 72 , 90 ] at 37 °C in a 5% CO 2 incubator as described in [ 54 ]. At the end of 6 or 24 h, cells were washed with PBS, fixed with 4% PFA and immunocytochemistry was performed as described in “ Methods ”.…”

Section: Methodsmentioning

confidence: 99%

The Alzheimer’s gene SORL1 is a regulator of endosomal traffic and recycling in human neurons

Mishra

Knupp

Szabo

et al. 2022

Cell. Mol. Life Sci.

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Background Loss of the Sortilin-related receptor 1 (SORL1) gene seems to act as a causal event for Alzheimer’s disease (AD). Recent studies have established that loss of SORL1, as well as mutations in autosomal dominant AD genes APP and PSEN1/2, pathogenically converge by swelling early endosomes, AD’s cytopathological hallmark. Acting together with the retromer trafficking complex, SORL1 has been shown to regulate the recycling of the amyloid precursor protein (APP) out of the endosome, contributing to endosomal swelling and to APP misprocessing. We hypothesized that SORL1 plays a broader role in neuronal endosomal recycling and used human induced pluripotent stem cell-derived neurons (hiPSC-Ns) to test this hypothesis. We examined endosomal recycling of three transmembrane proteins linked to AD pathophysiology: APP, the BDNF receptor Tropomyosin-related kinase B (TRKB), and the glutamate receptor subunit AMPA1 (GLUA1). Methods We used isogenic hiPSCs engineered to have SORL1 depleted or to have enhanced SORL1 expression. We differentiated neurons from these cell lines and mapped the trafficking of APP, TRKB and GLUA1 within the endosomal network using confocal microscopy. We also performed cell surface recycling and lysosomal degradation assays to assess the functionality of the endosomal network in both SORL1-depleted and -overexpressing neurons. The functional impact of GLUA1 recycling was determined by measuring synaptic activity. Finally, we analyzed alterations in gene expression in SORL1-depleted neurons using RNA sequencing. Results We find that as with APP, endosomal trafficking of GLUA1 and TRKB is impaired by loss of SORL1. We show that trafficking of all three cargoes to late endosomes and lysosomes is affected by manipulating SORL1 expression. We also show that depletion of SORL1 significantly impacts the endosomal recycling pathway for APP and GLUA1 at the level of the recycling endosome and trafficking to the cell surface. This has a functional effect on neuronal activity as shown by multi-electrode array (MEA). Conversely, increased SORL1 expression enhances endosomal recycling for APP and GLUA1. Our unbiased transcriptomic data further support SORL1’s role in endosomal recycling. We observe altered expression networks that regulate cell surface trafficking and neurotrophic signaling in SORL1-depleted neurons. Conclusion Collectively, and together with other recent observations, these findings suggest that one role for SORL1 is to contribute to endosomal degradation and recycling pathways in neurons, a conclusion that has both pathogenic and therapeutic implications for Alzheimer’s disease.

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Section: Methodsmentioning

confidence: 99%

The Alzheimer’s gene SORL1 is a regulator of endosomal traffic and recycling in human neurons

Mishra

Knupp

Szabo

et al. 2022

Cell. Mol. Life Sci.

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“…The suppression of ERS also mitigates epileptic behaviors of the seizure model ( Yokoi et al, 2015 ; Zhu et al, 2017 ) and AMPA-induced cognitive impairment in rats ( Bhardwaj et al, 2021 ). In recent years, another essential Ca 2+ storage-related organelle, the lysosome, has gradually been emphasized with excitotoxicity ( Vucicevic et al, 2020 ; Davis et al, 2021 ). Furthermore, enhanced cytoprotective autophagy can mitigate glutamate-induced excitotoxicity.…”

Section: Discussionmentioning

confidence: 99%

Restoration of Sarco/Endoplasmic Reticulum Ca2+-ATPase Activity Functions as a Pivotal Therapeutic Target of Anti-Glutamate-Induced Excitotoxicity to Attenuate Endoplasmic Reticulum Ca2+ Depletion

Zhang

Pang

et al. 2022

Front. Pharmacol.

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Glutamate-induced excitotoxicity is a pathological basis of many acute/chronic neurodegenerative diseases. Sarco/endoplasmic reticulum Ca2+-ATPase (SERCA2b) is a membrane-embedded P-type ATPase pump that manages the translocation of calcium ions (Ca2+) from cytosol into the lumen of the endoplasmic reticulum (ER) calcium stores. It participates in a wide range of biological functions in the central nervous system (CNS). However, the role of SERCA2b in glutamate-induced excitotoxicity and its mechanism must be elucidated. Herein, we demonstrate that SERCA2b mutants exacerbate the excitotoxicity of hypo-glutamate stimulation on HT22 cells. In this study, SERCA2b mutants accelerated Ca2+ depletion through loss-of-function (reduced pumping capacity) or gain-of-function (acquired leakage), resulting in ER stress. In addition, the occurrence of ER Ca2+ depletion increased mitochondria-associated membrane formation, which led to mitochondrial Ca2+ overload and dysfunction. Moreover, the enhancement of SERCA2b pumping capacity or inhibition of Ca2+ leakage attenuated Ca2+ depletion and impeded excitotoxicity in response to hypo-glutamate stimulation. In conclusion, SERCA2b mutants exacerbate ER Ca2+-depletion-mediated excitotoxicity in glutamate-sensitive HT22 cells. The mechanism of disruption is mainly related to the heterogeneity of SERCA2b mutation sites. Stabilization of SRECA2b function is a critical therapeutic approach against glutamate-induced excitotoxicity. These data will expand understanding of organelle regulatory networks and facilitate the discovery and creation of drugs against excitatory/inhibitory imbalance in the CNS.

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“…hiPSC-derived neurons were seeded at a density of 400,000 cells/well of a Matrigel coated 48-well plate. After 24 h, cells were washed once with DPBS, treated with complete media containing either 10 µg/ml DQ Red BSA or vehicle (PBS) and incubated for 24 h at 37 °C in a 5% CO2 incubator as described in( 4) and (5). At the end of 24 h, cells were washed with PBS, fixed with 4% PFA and immunocytochemistry was performed as described below in Supplemental Experimental Procedures.…”

Section: Author Contributionsmentioning

confidence: 99%

Pharmacologic Stabilization of Retromer Rescues Endosomal Pathology Induced by Defects in the Alzheimer’s geneSORL1

Mishra

Knupp

Kinoshita

et al. 2022

Preprint

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Defects in the Sortilin-related receptor 1 gene, SORL1, are highly pathogenic in Alzheimer's disease (AD), on par with other causal AD genes. SORL1 is a key regulator of endosomal trafficking in neurons, and loss of SORL1 function disrupts trafficking pathways that are vital for neuronal health. SORL1 has been found to interact with retromer, a protein complex that is a 'master conductor' of endosomal trafficking. Here we rely on retromer pharmacological chaperones that stabilize retromer in vitro, and thus enhance retromer function in cells. to test the hypothesis that they will correct AD-associated neuronal pathologies. We used an isogenic series of human induced pluripotent stem cell (hiPSCs) lines either deficient in SORL1 expression (SORL1 KO or SORL1+/-) or harboring one copy of a missense variant linked to increased risk for AD (SORL1Var). We treated neurons derived from these hiPSCs with the established retromer chaperone, TPT-260, and tested whether they corrected indicators of AD's defining endosomal, amyloid, and Tau pathologies. We observed that TPT-260 treatment was able to correct all these pathological indicators; however, the degree of rescue of some of the phenotypes was dependent on whether the neurons harbor at least one functional copy of SORL1 or whether they are fully deficient. More than just validating that SORL1-retromer acts as a functional trafficking unit in neurons, our work sheds new light on how SORL1-retromer is linked to AD's cardinal pathologies and illuminates how this pathway can be therapeutically harnessed.

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Delivering progranulin to neuronal lysosomes protects against excitotoxicity

Cited by 17 publications

References 98 publications

The Alzheimer’s gene SORL1 is a regulator of endosomal traffic and recycling in human neurons

The Alzheimer’s gene SORL1 is a regulator of endosomal traffic and recycling in human neurons

Restoration of Sarco/Endoplasmic Reticulum Ca2+-ATPase Activity Functions as a Pivotal Therapeutic Target of Anti-Glutamate-Induced Excitotoxicity to Attenuate Endoplasmic Reticulum Ca2+ Depletion

Pharmacologic Stabilization of Retromer Rescues Endosomal Pathology Induced by Defects in the Alzheimer’s geneSORL1

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