DEPDC5 and NPRL3 modulate cell size, filopodial outgrowth, and localization of mTOR in neural progenitor cells and neurons
Mutations in DEPDC5 and NPRL3 subunits of GATOR1, a modulator of mechanistic target of rapamycin (mTOR), are linked to malformations of cortical development (MCD). Brain specimens from these individuals reveal abnormal cortical lamination, altered cell morphology, and hyperphosphorylation of ribosom...
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doaj-d96a4bc055504f58a5b144f60abff0d52021-03-22T12:46:19ZengElsevierNeurobiology of Disease1095-953X2018-06-01114184193DEPDC5 and NPRL3 modulate cell size, filopodial outgrowth, and localization of mTOR in neural progenitor cells and neuronsPhilip H. Iffland, II0Marianna Baybis1Allan E. Barnes2Richard J. Leventer3Paul J. Lockhart4Peter B. Crino5Shriners Hospitals Pediatric Research Center, Lewis Katz School of Medicine at Temple University, Philadelphia, PA, United States; Department of Neurology, University of Maryland School of Medicine, Baltimore, MD, United StatesShriners Hospitals Pediatric Research Center, Lewis Katz School of Medicine at Temple University, Philadelphia, PA, United States; Department of Neurology, University of Maryland School of Medicine, Baltimore, MD, United StatesDepartment of Neurology, University of Maryland School of Medicine, Baltimore, MD, United StatesDepartment of Neurology, Royal Children's Hospital and Neuroscience Research Group, Murdoch Children's Research Institute and University of Melbourne, Department of Pediatrics, Melbourne, AustraliaBruce Lefroy Center for Genetic Health Research, Murdoch Children's Research Institute, and University of Melbourne, Department of Pediatrics, Melbourne, AustraliaShriners Hospitals Pediatric Research Center, Lewis Katz School of Medicine at Temple University, Philadelphia, PA, United States; Department of Neurology, University of Maryland School of Medicine, Baltimore, MD, United States; Corresponding author at: Department of Neurology, 110 S. Paca St., Baltimore, MD 21201, United States.Mutations in DEPDC5 and NPRL3 subunits of GATOR1, a modulator of mechanistic target of rapamycin (mTOR), are linked to malformations of cortical development (MCD). Brain specimens from these individuals reveal abnormal cortical lamination, altered cell morphology, and hyperphosphorylation of ribosomal S6 protein (PS6), a marker for mTOR activation. While numerous studies have examined GATOR1 subunit function in non-neuronal cell lines, few have directly assessed loss of GATOR1 subunit function in neuronal cell types. We hypothesized that DEPDC5 or NPRL3 shRNA-mediated knockdown (DEPDC5/NPRL3 KD) leads to inappropriate functional activation of mTOR and mTOR-dependent alterations in neuronal morphology.Neuronal size was determined in human specimens harboring DEPDC5 or NPRL3 mutations resected for epilepsy treatment. DEPDC5/NPRL3 KD effects on cell size, filopodial extension, subcellular mTOR complex 1 (mTORC1) localization, and mTORC1 activation during nutrient deprivation were assayed in mouse neuroblastoma cells (N2aC) and mouse subventricular zone derived neural progenitor cells (mNPCs). mTORC1-dependent effects of DEPDC5/NPRL3 KD were determined using the mTOR inhibitor rapamycin. Changes in mTOR subcellular localization and mTORC1 pathway activation following DEPDC5/NPRL3 KD were determined by examining the proximity of mTOR to the lysosomal surface during amino acid starvation.Neurons exhibiting PS6 immunoreactivity (Ser 235/236) in human specimens were 1.5× larger than neurons in post-mortem control samples. DEPDC5/NPRL3 KD caused mTORC1, but not mTORC2, hyperactivation, soma enlargement, and increased filopodia in N2aC and mNPCs compared with wildtype cells. DEPDC5/NPRL3 KD led to inappropriate mTOR localization at the lysosome along with constitutive mTOR activation following amino acid deprivation. DEPDC5/NPRL3 KD effects on morphology and functional mTOR activation were reversed by rapamycin. mTOR-dependent effects of DEPDC5/NPRL3 KD on morphology and subcellular localization of mTOR in neurons suggests that loss-of-function in GATOR1 subunits may play a role in MCD formation during fetal brain development.http://www.sciencedirect.com/science/article/pii/S0969996118300457GATOR1Focal cortical dysplasiamTORopathyEpilepsyNeural progenitor cellsBrain development |
collection |
DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Philip H. Iffland, II Marianna Baybis Allan E. Barnes Richard J. Leventer Paul J. Lockhart Peter B. Crino |
spellingShingle |
Philip H. Iffland, II Marianna Baybis Allan E. Barnes Richard J. Leventer Paul J. Lockhart Peter B. Crino DEPDC5 and NPRL3 modulate cell size, filopodial outgrowth, and localization of mTOR in neural progenitor cells and neurons Neurobiology of Disease GATOR1 Focal cortical dysplasia mTORopathy Epilepsy Neural progenitor cells Brain development |
author_facet |
Philip H. Iffland, II Marianna Baybis Allan E. Barnes Richard J. Leventer Paul J. Lockhart Peter B. Crino |
author_sort |
Philip H. Iffland, II |
title |
DEPDC5 and NPRL3 modulate cell size, filopodial outgrowth, and localization of mTOR in neural progenitor cells and neurons |
title_short |
DEPDC5 and NPRL3 modulate cell size, filopodial outgrowth, and localization of mTOR in neural progenitor cells and neurons |
title_full |
DEPDC5 and NPRL3 modulate cell size, filopodial outgrowth, and localization of mTOR in neural progenitor cells and neurons |
title_fullStr |
DEPDC5 and NPRL3 modulate cell size, filopodial outgrowth, and localization of mTOR in neural progenitor cells and neurons |
title_full_unstemmed |
DEPDC5 and NPRL3 modulate cell size, filopodial outgrowth, and localization of mTOR in neural progenitor cells and neurons |
title_sort |
depdc5 and nprl3 modulate cell size, filopodial outgrowth, and localization of mtor in neural progenitor cells and neurons |
publisher |
Elsevier |
series |
Neurobiology of Disease |
issn |
1095-953X |
publishDate |
2018-06-01 |
description |
Mutations in DEPDC5 and NPRL3 subunits of GATOR1, a modulator of mechanistic target of rapamycin (mTOR), are linked to malformations of cortical development (MCD). Brain specimens from these individuals reveal abnormal cortical lamination, altered cell morphology, and hyperphosphorylation of ribosomal S6 protein (PS6), a marker for mTOR activation. While numerous studies have examined GATOR1 subunit function in non-neuronal cell lines, few have directly assessed loss of GATOR1 subunit function in neuronal cell types. We hypothesized that DEPDC5 or NPRL3 shRNA-mediated knockdown (DEPDC5/NPRL3 KD) leads to inappropriate functional activation of mTOR and mTOR-dependent alterations in neuronal morphology.Neuronal size was determined in human specimens harboring DEPDC5 or NPRL3 mutations resected for epilepsy treatment. DEPDC5/NPRL3 KD effects on cell size, filopodial extension, subcellular mTOR complex 1 (mTORC1) localization, and mTORC1 activation during nutrient deprivation were assayed in mouse neuroblastoma cells (N2aC) and mouse subventricular zone derived neural progenitor cells (mNPCs). mTORC1-dependent effects of DEPDC5/NPRL3 KD were determined using the mTOR inhibitor rapamycin. Changes in mTOR subcellular localization and mTORC1 pathway activation following DEPDC5/NPRL3 KD were determined by examining the proximity of mTOR to the lysosomal surface during amino acid starvation.Neurons exhibiting PS6 immunoreactivity (Ser 235/236) in human specimens were 1.5× larger than neurons in post-mortem control samples. DEPDC5/NPRL3 KD caused mTORC1, but not mTORC2, hyperactivation, soma enlargement, and increased filopodia in N2aC and mNPCs compared with wildtype cells. DEPDC5/NPRL3 KD led to inappropriate mTOR localization at the lysosome along with constitutive mTOR activation following amino acid deprivation. DEPDC5/NPRL3 KD effects on morphology and functional mTOR activation were reversed by rapamycin. mTOR-dependent effects of DEPDC5/NPRL3 KD on morphology and subcellular localization of mTOR in neurons suggests that loss-of-function in GATOR1 subunits may play a role in MCD formation during fetal brain development. |
topic |
GATOR1 Focal cortical dysplasia mTORopathy Epilepsy Neural progenitor cells Brain development |
url |
http://www.sciencedirect.com/science/article/pii/S0969996118300457 |
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