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Kazuaki Yoshikawa - One of the best experts on this subject based on the ideXlab platform.
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K (2014) Antagonistic Interplay between Necdin and
2016Co-Authors: Ryohei Minamide, Koichi Hasegawa, Kazushiro Fujiwara, Kazuaki YoshikawaAbstract:Neural precursor cells (NPCs) in the neocortex exhibit a high proliferation capacity during early embryonic development and give rise to cortical projection neurons after maturation. Necdin, a mammal-specific MAGE (melanoma antigen) family protein that possesses anti-mitotic and pro-survival activities, is expressed abundantly in postmitotic neurons and moderately in tissue-specific stem cells or progenitors. Necdin interacts with E2F transcription factors and suppresses E2F1-dependent transcriptional activation of the cyclin-dependent kinase Cdk1 gene. Here we show that Necdin serves as a suppressor of NPC proliferation in the embryonic neocortex. Necdin is moderately expressed in the ventricular zone of mouse embryonic neocortex, in which proliferative cell populations are significantly increased in Necdin-null mice. In the neocortex of Necdin-null embryos, expression of Cdk1 and Sox2, a stem cell marker, is significantly increased, whereas expression of p16, a cyclin-dependent kinase inhibitor, is markedly diminished. Cdk1 and p16 expression levels are also significantly increased and decreased, respectively, in primary NPCs prepared from Necdin-null embryos. Intriguingly, Necdin interacts directly with Bmi1, a Polycomb group protein that suppresses p16 expression and promotes NPC proliferation. In HEK293A cells transfected with luciferase reporter constructs, Necdin relieves Bmi1-dependent repression of p16 promoter activity, whereas Bmi1 counteracts Necdin-mediated repression of E2F1-dependent Cdk1 promoter activity. In lentivirus-infected primary NPCs, Necdin overexpression increases p16 expression, suppresses Cdk1 expression, and inhibits NP
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Promotion of mitochondrial biogenesis by Necdin protects neurons against mitochondrial insults.
Nature communications, 2016Co-Authors: Koichi Hasegawa, Kazushiro Fujiwara, Hideki Mochizuki, Toru Yasuda, Chinatsu Shiraishi, Serge Przedborski, Kazuaki YoshikawaAbstract:Neurons rely heavily on mitochondria for their function and survival. Mitochondrial dysfunction contributes to the pathogenesis of neurodegenerative diseases such as Parkinson's disease. PGC-1α is a master regulator of mitochondrial biogenesis and function. Here we identify Necdin as a potent PGC-1α stabilizer that promotes mitochondrial biogenesis via PGC-1α in mammalian neurons. Expression of genes encoding mitochondria-specific proteins decreases significantly in Necdin-null cortical neurons, where mitochondrial function and expression of the PGC-1α protein are reduced. Necdin strongly stabilizes PGC-1α by inhibiting its ubiquitin-dependent degradation. Forced expression of Necdin enhances mitochondrial function in primary cortical neurons and human SH-SY5Y neuroblastoma cells to prevent mitochondrial respiratory chain inhibitor-induced degeneration. Moreover, overexpression of Necdin in the substantia nigra in vivo of adult mice protects dopaminergic neurons against degeneration in experimental Parkinson's disease. These data reveal that Necdin promotes mitochondrial biogenesis through stabilization of endogenous PGC-1α to exert neuroprotection against mitochondrial insults.
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Necdin controls EGFR signaling linked to astrocyte differentiation in primary cortical progenitor cells
Cellular signalling, 2015Co-Authors: Izumi Fujimoto, Koichi Hasegawa, Kazushiro Fujiwara, Masashi Yamada, Kazuaki YoshikawaAbstract:Cellular signaling mediated by the EGF receptor (EGFR) plays a key role in controlling proliferation and differentiation of cortical progenitor cells (CPCs). However, regulatory mechanisms of EGFR signaling in CPCs remain largely unknown. Here we demonstrate that Necdin, a MAGE (melanoma antigen) family protein, interacts with EGFR in primary CPCs and represses its downstream signaling linked to astrocyte differentiation. EGFR was autophosphorylated and interacted with Necdin in EGF-stimulated CPCs. Necdin bound to autophosphorylated EGFR via its tyrosine kinase domain. EGF-induced phosphorylation of ERK was enhanced in Necdin-null CPCs, where the interaction between EGFR and the adaptor protein Grb2 was strengthened, suggesting that endogenous Necdin suppresses the EGFR/ERK signaling pathway in CPCs. In Necdin-null CPCs, astrocyte differentiation induced by the gliogenic cytokine cardiotrophin-1 was significantly accelerated in the presence of EGF, and inhibition of EGFR/ERK signaling abolished the acceleration. Furthermore, Necdin strongly suppressed astrocyte differentiation induced by overexpression of EGFR or its ligand binding-defective mutant equivalent to a glioblastoma-associated EGFR variant. These results suggest that Necdin acts as an intrinsic suppressor of the EGFR/ERK signaling pathway in EGF-responsive CPCs to restrain astroglial development in a cell-autonomous manner.
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Necdin Promotes Ubiquitin-Dependent Degradation of PIAS1 SUMO E3 Ligase
PloS one, 2014Co-Authors: Ibrahim Gur, Koichi Hasegawa, Kazushiro Fujiwara, Kazuaki YoshikawaAbstract:Necdin, a pleiotropic protein that promotes differentiation and survival of mammalian neurons, is a member of MAGE (melanoma antigen) family proteins that share a highly conserved MAGE homology domain. Several MAGE proteins interact with ubiquitin E3 ligases and modulate their activities. However, it remains unknown whether MAGE family proteins interact with SUMO (small ubiquitin-like modifier) E3 ligases such as PIAS (protein inhibitor of activated STAT) family, Nsmce2/Mms21 and Cbx4/Pc2. In the present study, we examined whether Necdin interacts with these SUMO E3 ligases. Co-immunoprecipitation analysis revealed that Necdin, MAGED1, MAGEF1 and MAGEL2 bound to PIAS1 but not to Nsmce2 or Cbx4. These SUMO E3 ligases bound to MAGEA1 but failed to interact with Necdin-like 2/MAGEG1. Necdin bound to PIAS1 central domains that are highly conserved among PIAS family proteins and suppressed PIAS1-dependent sumoylation of the substrates STAT1 and PML (promyelocytic leukemia protein). Remarkably, Necdin promoted degradation of PIAS1 via the ubiquitin-proteasome pathway. In transfected HEK293A cells, amino- and carboxyl-terminally truncated mutants of PIAS1 bound to Necdin but failed to undergo Necdin-dependent ubiquitination. Both PIAS1 and Necdin were associated with the nuclear matrix, where the PIAS1 terminal deletion mutants failed to localize, implying that the nuclear matrix is indispensable for Necdin-dependent ubiquitination of PIAS1. Our data suggest that Necdin suppresses PIAS1 both by inhibiting SUMO E3 ligase activity and by promoting ubiquitin-dependent degradation.
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Antagonistic interplay between Necdin and Bmi1 controls proliferation of neural precursor cells in the embryonic mouse neocortex.
PloS one, 2014Co-Authors: Ryohei Minamide, Koichi Hasegawa, Kazushiro Fujiwara, Kazuaki YoshikawaAbstract:Neural precursor cells (NPCs) in the neocortex exhibit a high proliferation capacity during early embryonic development and give rise to cortical projection neurons after maturation. Necdin, a mammal-specific MAGE (melanoma antigen) family protein that possesses anti-mitotic and pro-survival activities, is expressed abundantly in postmitotic neurons and moderately in tissue-specific stem cells or progenitors. Necdin interacts with E2F transcription factors and suppresses E2F1-dependent transcriptional activation of the cyclin-dependent kinase Cdk1 gene. Here we show that Necdin serves as a suppressor of NPC proliferation in the embryonic neocortex. Necdin is moderately expressed in the ventricular zone of mouse embryonic neocortex, in which proliferative cell populations are significantly increased in Necdin-null mice. In the neocortex of Necdin-null embryos, expression of Cdk1 and Sox2, a stem cell marker, is significantly increased, whereas expression of p16, a cyclin-dependent kinase inhibitor, is markedly diminished. Cdk1 and p16 expression levels are also significantly increased and decreased, respectively, in primary NPCs prepared from Necdin-null embryos. Intriguingly, Necdin interacts directly with Bmi1, a Polycomb group protein that suppresses p16 expression and promotes NPC proliferation. In HEK293A cells transfected with luciferase reporter constructs, Necdin relieves Bmi1-dependent repression of p16 promoter activity, whereas Bmi1 counteracts Necdin-mediated repression of E2F1-dependent Cdk1 promoter activity. In lentivirus-infected primary NPCs, Necdin overexpression increases p16 expression, suppresses Cdk1 expression, and inhibits NPC proliferation, whereas Bmi1 overexpression suppresses p16 expression, increases Cdk1 expression, and promotes NPC proliferation. Our data suggest that embryonic NPC proliferation in the neocortex is regulated by the antagonistic interplay between Necdin and Bmi1.
Rachel Wevrick - One of the best experts on this subject based on the ideXlab platform.
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The Necdin interactome: evaluating the effects of amino acid substitutions and cell stress using proximity-dependent biotinylation (BioID) and mass spectrometry
Human Genetics, 2020Co-Authors: Matthea R. Sanderson, Katherine E. Badior, Richard P. Fahlman, Rachel WevrickAbstract:Prader–Willi syndrome (PWS) is a neurodevelopmental disorder caused by the loss of function of a set of imprinted genes on chromosome 15q11–15q13. One of these genes, NDN , encodes Necdin, a protein that is important for neuronal differentiation and survival. Loss of Ndn in mice causes defects in the formation and function of the nervous system. Necdin is a member of the melanoma-associated antigen gene (MAGE) protein family. The functions of MAGE proteins depend highly on their interactions with other proteins, and in particular MAGE proteins interact with E3 ubiquitin ligases and deubiquitinases to form MAGE-RING E3 ligase-deubiquitinase complexes. Here, we used proximity-dependent biotin identification (BioID) and mass spectrometry (MS) to determine the network of protein–protein interactions (interactome) of the Necdin protein. This process yielded novel as well as known Necdin-proximate proteins that cluster into a protein network. Next, we used BioID-MS to define the interactomes of Necdin proteins carrying coding variants. Variant Necdin proteins had interactomes that were distinct from wildtype Necdin. BioID-MS is not only a useful tool to identify protein–protein interactions, but also to analyze the effects of variants of unknown significance on the interactomes of proteins involved in genetic disease.
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The Prader-Willi syndrome proteins MAGEL2 and Necdin regulate leptin receptor cell surface abundance through ubiquitination pathways.
Human molecular genetics, 2017Co-Authors: Tishani Methsala Wijesuriya, Matthea R. Sanderson, Leentje De Ceuninck, Delphine Masschaele, Karin Vanessa Carias, Jan Tavernier, Rachel WevrickAbstract:In Prader-Willi syndrome (PWS), obesity is caused by the disruption of appetite-controlling pathways in the brain. Two PWS candidate genes encode MAGEL2 and Necdin, related melanoma antigen proteins that assemble into ubiquitination complexes. Mice lacking Magel2 are obese and lack leptin sensitivity in hypothalamic pro-opiomelanocortin neurons, suggesting dysregulation of leptin receptor (LepR) activity. Hypothalamus from Magel2-null mice had less LepR and altered levels of ubiquitin pathway proteins that regulate LepR processing (Rnf41, Usp8, and Stam1). MAGEL2 increased the cell surface abundance of LepR and decreased their degradation. LepR interacts with Necdin, which interacts with MAGEL2, which complexes with RNF41 and USP8. Mutations in the MAGE homology domain of MAGEL2 suppress RNF41 stabilization and prevent the MAGEL2-mediated increase of cell surface LepR. Thus, MAGEL2 and Necdin together control LepR sorting and degradation through a dynamic ubiquitin-dependent pathway. Loss of MAGEL2 and Necdin may uncouple LepR from ubiquitination pathways, providing a cellular mechanism for obesity in PWS.
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Necdin, a p53 target gene, regulates the quiescence and response to genotoxic stress of hematopoietic stem/progenitor cells
Blood, 2012Co-Authors: Takashi Asai, Yan Liu, Silvana Di Giandomenico, Anthony Deblasio, Silvia Menendez, Boris Reva, Narae Bae, Delphine Ndiaye-lobry, Yevgeniy Antipin, Rachel WevrickAbstract:We recently defined a critical role for p53 in regulating the quiescence of adult hematopoietic stem cells (HSCs) and identified Necdin as a candidate p53 target gene. Necdin is a growth-suppressing protein and the gene encoding it is one of several that are deleted in patients with Prader-Willi syndrome. To define the intrinsic role of Necdin in adult hematopoiesis, in the present study, we transplanted Necdin-null fetal liver cells into lethally irradiated recipients. We show that Necdin-null adult HSCs are less quiescent and more proliferative than normal HSCs, demonstrating the similar role of Necdin and p53 in promoting HSC quiescence during steady-state conditions. However, wild-type recipients repopulated with Necdin-null hematopoietic stem/progenitor cells show enhanced sensitivity to irradiation and chemotherapy, with increased p53-dependent apoptosis, myelosuppression, and mortality. Necdin controls the HSC response to genotoxic stress via both cell-cycle–dependent and cell-cycle–independent mechanisms, with the latter occurring in a Gas2L3-dependent manner. We conclude that Necdin functions as a molecular switch in adult hematopoiesis, acting in a p53-like manner to promote HSC quiescence in the steady state, but suppressing p53-dependent apoptosis in response to genotoxic stress.
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Loss of the Prader-Willi obesity syndrome protein Necdin promotes adipogenesis.
Gene, 2012Co-Authors: Jason R. Bush, Rachel WevrickAbstract:We investigated the role of Necdin during adipogenic differentiation. Necdin is one of several genes inactivated in children with Prader-Willi syndrome, who are predisposed to increased adiposity at the expense of lean mass. Necdin promotes neuronal and muscle differentiation and survival through interactions with a variety of proteins, including cell surface receptors, modifiers of protein stability, and transcription factors. In pre-adipocytes, Necdin over-expression inhibits adipogenesis, while reducing Necdin levels enhances adipogenic differentiation in tissue culture cells. We now directly demonstrate a role for Necdin in inhibiting adipogenesis using cells derived from Necdin deficient mice.
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Necdin Regulates Hematopoietic Stem Cell Quiescence and Sensitivity to Genotoxic Stress.
Blood, 2009Co-Authors: Takashi Asai, Rachel Wevrick, Yan Liu, Silvana Di Giandomenico, Anthony Deblasio, Silvia Menendez, Jack Antipin, Boris Reva, Stephen D. NimerAbstract:Abstract Abstract 379 Necdin, a member of MAGE (melanoma antigen) family proteins, is a growth suppressing protein that was first identified in post mitotic neurons. The gene encoding Necdin is one of several deleted in individuals with Prader-Willi syndrome, a neurobehavioural disorder associated with an increased risk of myeloid leukemia. It is reported that Necdin interacts with p53 and represses p53-mediated apoptosis in neurons, but its role in hematopoiesis is largely unknown. Recently, we defined a critical role of p53 in regulating hematopoietic stem cell quiescence, and identified Necdin as a target gene of p53, that is highly expressed in LT-HSCs (Liu Y et al., Cell Stem Cell, 2009). To define the role of Necdin in hematopoiesis, we have analyzed the hematopoietic compartment of Necdin-null mice. As Necdin-null mice die perinatally, we first investigated fetal hematopoiesis and found no alteration in the frequency of fetal liver HSCs, defined as Lin-Sca1+Mac1+CD48-CD150+ within the fetal liver cells. Although Necdin-null fetal liver HSCs increase serial replating capability in methylcellulose and maintain stemness in long-term stromal based cultures better than wild type HSCs, Necdin-null fetal liver HSCs repopulate lethally irradiated recipient mice similar to wild type HSCs, in primary, secondary, and tertiary serial bone marrow transplantation assays. In addition, Necdin-null HSCs show almost comparable repopulating ability as wild type HSCs, after secondary competitive bone marrow transplantation assays. These imply that Necdin is dispensable for HSC self renewal. On the other hand, BM-derived Necdin-null HSCs show decreased quiescence 4 months after transplantation, and increased proliferation as indicated by in vivo BrdU incorporation assays. Furthermore, recipient mice repopulated with Necdin-null HSCs show enhanced sensitivity both to weekly 5-FU administration and to total body irradiation, as manifested by increased mortality. This suggests that the decreased quiescence of Necdin-null HSCs leads to their depletion under conditions of genotoxic stress. Gene expression profiling studies have identified several deregulated signaling pathways in the Necdin-null HSCs. Expression of several p53 target genes is altered in irradiated Necdin-null HSCs, which may account for their enhanced radiosensitivity. We are now investigating these Necdin target genes to clarify how Necdin functions to critically regulate HSC quiescence. We are also determining whether targeting Necdin could be a therapeutic approach to eliminate quiescent leukemia stem cells, using a murine CML model. Disclosures: No relevant conflicts of interest to declare.
Françoise Muscatelli - One of the best experts on this subject based on the ideXlab platform.
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JCB: ARTICLE Necdin mediates skeletal muscle regeneration by promoting myoblast survival and differentiation
2013Co-Authors: Daniela Deponti, Françoise Muscatelli, Stephanie François, Silvia Baesso, Clara Sciorati, Anna Innocenzi, Vania Broccoli, Raffaella Meneveri, Emilio Clementi, Giulio CossuAbstract:Regeneration of muscle fibers that are lost during pathological muscle degeneration or after injuries is sustained by the production of new myofi bers. An important cell type involved in muscle regeneration is the satellite cell. Necdin is a protein expressed in satellite cell–derived myogenic precursors during perinatal growth. However, its function in myogenesis is not known. We compare transgenic mice that overexpress Necdin in skeletal muscle with both wild-type and Necdin null mice
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Functional consequences of Necdin nucleocytoplasmic localization.
PloS one, 2012Co-Authors: Anat Lavi-itzkovitz, Françoise Muscatelli, Marianna Tcherpakov, Zehava Levy, Shalev Itzkovitz, Mike FainzilberAbstract:Necdin, a MAGE family protein expressed primarily in the nervous system, has been shown to interact with both nuclear and cytoplasmic proteins, but the mechanism of its nucleocytoplasmic transport are unknown. We carried out a large-scale interaction screen using Necdin as a bait in the yeast RRS system, and found a wide range of potential interactors with different subcellular localizations, including over 60 new candidates for direct binding to Necdin. Integration of these interactions into a comprehensive network revealed a number of coherent interaction modules, including a cytoplasmic module connecting to Necdin through huntingtin-associated protein 1 (Hap1), dynactin and hip-1 protein interactor (Hippi); a nuclear P53 and Creb-binding-protein (Crebbp) module, connecting through Crebbp and WW domain-containing transcription regulator protein 1 (Wwtr1); and a nucleocytoplasmic transport module, connecting through transportins 1 and 2. We validated the Necdin-transportin1 interaction and characterized a sequence motif in Necdin that modulates karyopherin interaction. Surprisingly, a D234P Necdin mutant showed enhanced binding to both transportin1 and importin β1. Finally, exclusion of Necdin from the nucleus triggered extensive cell death. These data suggest that Necdin has multiple roles within protein complexes in different subcellular compartments, and indicate that it can utilize multiple karyopherin-dependent pathways to modulate its localization.
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functional consequences of Necdin nucleocytoplasmic localization
PLOS ONE, 2012Co-Authors: Anat Laviitzkovitz, Françoise Muscatelli, Marianna Tcherpakov, Zehava Levy, Shalev Itzkovitz, Mike FainzilberAbstract:Background Necdin, a MAGE family protein expressed primarily in the nervous system, has been shown to interact with both nuclear and cytoplasmic proteins, but the mechanism of its nucleocytoplasmic transport are unknown. Methodology/Principal Findings We carried out a large-scale interaction screen using Necdin as a bait in the yeast RRS system, and found a wide range of potential interactors with different subcellular localizations, including over 60 new candidates for direct binding to Necdin. Integration of these interactions into a comprehensive network revealed a number of coherent interaction modules, including a cytoplasmic module connecting to Necdin through huntingtin-associated protein 1 (Hap1), dynactin and hip-1 protein interactor (Hippi); a nuclear P53 and Creb-binding-protein (Crebbp) module, connecting through Crebbp and WW domain-containing transcription regulator protein 1 (Wwtr1); and a nucleocytoplasmic transport module, connecting through transportins 1 and 2. We validated the Necdin-transportin1 interaction and characterized a sequence motif in Necdin that modulates karyopherin interaction. Surprisingly, a D234P Necdin mutant showed enhanced binding to both transportin1 and importin β1. Finally, exclusion of Necdin from the nucleus triggered extensive cell death. Conclusions/Significance These data suggest that Necdin has multiple roles within protein complexes in different subcellular compartments, and indicate that it can utilize multiple karyopherin-dependent pathways to modulate its localization.
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Module decomposition of the Necdin network.
2012Co-Authors: Anat Lavi-itzkovitz, Françoise Muscatelli, Marianna Tcherpakov, Zehava Levy, Shalev Itzkovitz, Mike FainzilberAbstract:Modules were detected using the betweenness-centrality clustering algorithm [21]. Only modules containing more than 8 nodes are displayed. Blue edges denote published interactions, red are interactions detected in the present screen. Note that Necdin-Eid1 and Necdin-Nucleobindin1 are connected with 2 color edges. Modules - A. Necdin; B. p75; C. Grin-Ywhab; D. p53-Crebbp; E. Transportin; F. Huntingtin; G. MAGE-D1; H. APP; I. Clock. Necdin (Ndn) is shown for each module, together with interactions with the module members.
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Necdin Protects Embryonic Motoneurons from Programmed Cell Death
PloS one, 2011Co-Authors: Julianne Aebischer, David Andrieu, Rachel Sturny, Anne Rieusset, Fabienne Schaller, Sandrine Geib, Cédric Raoul, Françoise MuscatelliAbstract:Necdin belongs to the type II Melanoma Associated Antigen Gene Expression gene family and is located in the Prader-Willi Syndrome (PWS) critical region. Necdin-deficient mice develop symptoms of PWS, including a sensory and motor deficit. However, the mechanisms underlying the motor deficit remain elusive. Here, we show that the genetic ablation of Necdin, whose expression is restricted to post-mitotic neurons in the spinal cord during development, leads to a loss of 31% of specified motoneurons. The increased neuronal loss occurs during the period of naturally-occurring cell death and is not confined to specific pools of motoneurons. To better understand the role of Necdin during the period of programmed cell death of motoneurons we used embryonic spinal cord explants and primary motoneuron cultures from Necdin-deficient mice. Interestingly, while Necdin-deficient motoneurons present the same survival response to neurotrophic factors, we demonstrate that deletion of Necdin leads to an increased susceptibility of motoneurons to neurotrophic factor deprivation. We show that by neutralizing TNFα this increased susceptibility of Necdin-deficient motoneurons to trophic factor deprivation can be reduced to the normal level. We propose that Necdin is implicated through the TNF-receptor 1 pathway in the developmental death of motoneurons.
Kazushiro Fujiwara - One of the best experts on this subject based on the ideXlab platform.
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K (2014) Antagonistic Interplay between Necdin and
2016Co-Authors: Ryohei Minamide, Koichi Hasegawa, Kazushiro Fujiwara, Kazuaki YoshikawaAbstract:Neural precursor cells (NPCs) in the neocortex exhibit a high proliferation capacity during early embryonic development and give rise to cortical projection neurons after maturation. Necdin, a mammal-specific MAGE (melanoma antigen) family protein that possesses anti-mitotic and pro-survival activities, is expressed abundantly in postmitotic neurons and moderately in tissue-specific stem cells or progenitors. Necdin interacts with E2F transcription factors and suppresses E2F1-dependent transcriptional activation of the cyclin-dependent kinase Cdk1 gene. Here we show that Necdin serves as a suppressor of NPC proliferation in the embryonic neocortex. Necdin is moderately expressed in the ventricular zone of mouse embryonic neocortex, in which proliferative cell populations are significantly increased in Necdin-null mice. In the neocortex of Necdin-null embryos, expression of Cdk1 and Sox2, a stem cell marker, is significantly increased, whereas expression of p16, a cyclin-dependent kinase inhibitor, is markedly diminished. Cdk1 and p16 expression levels are also significantly increased and decreased, respectively, in primary NPCs prepared from Necdin-null embryos. Intriguingly, Necdin interacts directly with Bmi1, a Polycomb group protein that suppresses p16 expression and promotes NPC proliferation. In HEK293A cells transfected with luciferase reporter constructs, Necdin relieves Bmi1-dependent repression of p16 promoter activity, whereas Bmi1 counteracts Necdin-mediated repression of E2F1-dependent Cdk1 promoter activity. In lentivirus-infected primary NPCs, Necdin overexpression increases p16 expression, suppresses Cdk1 expression, and inhibits NP
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Necdin Controls Proliferation of White Adipocyte Progenitor Cells
2016Co-Authors: Kazushiro Fujiwara, Koichi Hasegawa, Tsuyoshi Ohkumo, Hiroyuki Miyoshi, Yu-hua TsengAbstract:White adipose tissues are composed mainly of white fat cells (adipocytes), which play a key role in energy storage and metabolism. White adipocytes are terminally differentiated postmitotic cells and arise from their progenitor cells (preadipocytes) or mesenchymal stem cells residing in white adipose tissues. Thus, white adipocyte number is most likely controlled by the rate of preadipocyte proliferation, which may contribute to the etiology of obesity. However, little is known about the molecular mechanisms that regulate preadipocyte proliferation during adipose tissue development. Necdin, which is expressed predominantly in postmitotic neurons, is a pleiotropic protein that possesses anti-mitotic and pro-survival activities. Here we show that Necdin functions as an intrinsic regulator of white preadipocyte proliferation in developing adipose tissues. Necdin is expressed in early preadipocytes or mesenchymal stem cells residing in the stromal compartment of white adipose tissues in juvenile mice. Lentivirus-mediated knockdown of endogenous Necdin expression in vivo in adipose tissues markedly increases fat mass in juvenile mice fed a high-fat diet until adulthood. Furthermore, Necdin-null mutant mice exhibit a greater expansion of adipose tissues due to adipocyte hyperplasia than wild-type mice when fed the high-fat diet during the juvenile and adult periods. Adipose stromal-vascular cells prepared from Necdin-null mice differentiate in vitro into a significantly larger number of adipocytes in response to adipogenic inducers than those from wild-type mice. These results suggest that Necdin prevents excessive preadipocyte proliferation induced b
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Promotion of mitochondrial biogenesis by Necdin protects neurons against mitochondrial insults.
Nature communications, 2016Co-Authors: Koichi Hasegawa, Kazushiro Fujiwara, Hideki Mochizuki, Toru Yasuda, Chinatsu Shiraishi, Serge Przedborski, Kazuaki YoshikawaAbstract:Neurons rely heavily on mitochondria for their function and survival. Mitochondrial dysfunction contributes to the pathogenesis of neurodegenerative diseases such as Parkinson's disease. PGC-1α is a master regulator of mitochondrial biogenesis and function. Here we identify Necdin as a potent PGC-1α stabilizer that promotes mitochondrial biogenesis via PGC-1α in mammalian neurons. Expression of genes encoding mitochondria-specific proteins decreases significantly in Necdin-null cortical neurons, where mitochondrial function and expression of the PGC-1α protein are reduced. Necdin strongly stabilizes PGC-1α by inhibiting its ubiquitin-dependent degradation. Forced expression of Necdin enhances mitochondrial function in primary cortical neurons and human SH-SY5Y neuroblastoma cells to prevent mitochondrial respiratory chain inhibitor-induced degeneration. Moreover, overexpression of Necdin in the substantia nigra in vivo of adult mice protects dopaminergic neurons against degeneration in experimental Parkinson's disease. These data reveal that Necdin promotes mitochondrial biogenesis through stabilization of endogenous PGC-1α to exert neuroprotection against mitochondrial insults.
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Necdin controls EGFR signaling linked to astrocyte differentiation in primary cortical progenitor cells
Cellular signalling, 2015Co-Authors: Izumi Fujimoto, Koichi Hasegawa, Kazushiro Fujiwara, Masashi Yamada, Kazuaki YoshikawaAbstract:Cellular signaling mediated by the EGF receptor (EGFR) plays a key role in controlling proliferation and differentiation of cortical progenitor cells (CPCs). However, regulatory mechanisms of EGFR signaling in CPCs remain largely unknown. Here we demonstrate that Necdin, a MAGE (melanoma antigen) family protein, interacts with EGFR in primary CPCs and represses its downstream signaling linked to astrocyte differentiation. EGFR was autophosphorylated and interacted with Necdin in EGF-stimulated CPCs. Necdin bound to autophosphorylated EGFR via its tyrosine kinase domain. EGF-induced phosphorylation of ERK was enhanced in Necdin-null CPCs, where the interaction between EGFR and the adaptor protein Grb2 was strengthened, suggesting that endogenous Necdin suppresses the EGFR/ERK signaling pathway in CPCs. In Necdin-null CPCs, astrocyte differentiation induced by the gliogenic cytokine cardiotrophin-1 was significantly accelerated in the presence of EGF, and inhibition of EGFR/ERK signaling abolished the acceleration. Furthermore, Necdin strongly suppressed astrocyte differentiation induced by overexpression of EGFR or its ligand binding-defective mutant equivalent to a glioblastoma-associated EGFR variant. These results suggest that Necdin acts as an intrinsic suppressor of the EGFR/ERK signaling pathway in EGF-responsive CPCs to restrain astroglial development in a cell-autonomous manner.
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Necdin Promotes Ubiquitin-Dependent Degradation of PIAS1 SUMO E3 Ligase
PloS one, 2014Co-Authors: Ibrahim Gur, Koichi Hasegawa, Kazushiro Fujiwara, Kazuaki YoshikawaAbstract:Necdin, a pleiotropic protein that promotes differentiation and survival of mammalian neurons, is a member of MAGE (melanoma antigen) family proteins that share a highly conserved MAGE homology domain. Several MAGE proteins interact with ubiquitin E3 ligases and modulate their activities. However, it remains unknown whether MAGE family proteins interact with SUMO (small ubiquitin-like modifier) E3 ligases such as PIAS (protein inhibitor of activated STAT) family, Nsmce2/Mms21 and Cbx4/Pc2. In the present study, we examined whether Necdin interacts with these SUMO E3 ligases. Co-immunoprecipitation analysis revealed that Necdin, MAGED1, MAGEF1 and MAGEL2 bound to PIAS1 but not to Nsmce2 or Cbx4. These SUMO E3 ligases bound to MAGEA1 but failed to interact with Necdin-like 2/MAGEG1. Necdin bound to PIAS1 central domains that are highly conserved among PIAS family proteins and suppressed PIAS1-dependent sumoylation of the substrates STAT1 and PML (promyelocytic leukemia protein). Remarkably, Necdin promoted degradation of PIAS1 via the ubiquitin-proteasome pathway. In transfected HEK293A cells, amino- and carboxyl-terminally truncated mutants of PIAS1 bound to Necdin but failed to undergo Necdin-dependent ubiquitination. Both PIAS1 and Necdin were associated with the nuclear matrix, where the PIAS1 terminal deletion mutants failed to localize, implying that the nuclear matrix is indispensable for Necdin-dependent ubiquitination of PIAS1. Our data suggest that Necdin suppresses PIAS1 both by inhibiting SUMO E3 ligase activity and by promoting ubiquitin-dependent degradation.
Koichi Hasegawa - One of the best experts on this subject based on the ideXlab platform.
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K (2014) Antagonistic Interplay between Necdin and
2016Co-Authors: Ryohei Minamide, Koichi Hasegawa, Kazushiro Fujiwara, Kazuaki YoshikawaAbstract:Neural precursor cells (NPCs) in the neocortex exhibit a high proliferation capacity during early embryonic development and give rise to cortical projection neurons after maturation. Necdin, a mammal-specific MAGE (melanoma antigen) family protein that possesses anti-mitotic and pro-survival activities, is expressed abundantly in postmitotic neurons and moderately in tissue-specific stem cells or progenitors. Necdin interacts with E2F transcription factors and suppresses E2F1-dependent transcriptional activation of the cyclin-dependent kinase Cdk1 gene. Here we show that Necdin serves as a suppressor of NPC proliferation in the embryonic neocortex. Necdin is moderately expressed in the ventricular zone of mouse embryonic neocortex, in which proliferative cell populations are significantly increased in Necdin-null mice. In the neocortex of Necdin-null embryos, expression of Cdk1 and Sox2, a stem cell marker, is significantly increased, whereas expression of p16, a cyclin-dependent kinase inhibitor, is markedly diminished. Cdk1 and p16 expression levels are also significantly increased and decreased, respectively, in primary NPCs prepared from Necdin-null embryos. Intriguingly, Necdin interacts directly with Bmi1, a Polycomb group protein that suppresses p16 expression and promotes NPC proliferation. In HEK293A cells transfected with luciferase reporter constructs, Necdin relieves Bmi1-dependent repression of p16 promoter activity, whereas Bmi1 counteracts Necdin-mediated repression of E2F1-dependent Cdk1 promoter activity. In lentivirus-infected primary NPCs, Necdin overexpression increases p16 expression, suppresses Cdk1 expression, and inhibits NP
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Necdin Controls Proliferation of White Adipocyte Progenitor Cells
2016Co-Authors: Kazushiro Fujiwara, Koichi Hasegawa, Tsuyoshi Ohkumo, Hiroyuki Miyoshi, Yu-hua TsengAbstract:White adipose tissues are composed mainly of white fat cells (adipocytes), which play a key role in energy storage and metabolism. White adipocytes are terminally differentiated postmitotic cells and arise from their progenitor cells (preadipocytes) or mesenchymal stem cells residing in white adipose tissues. Thus, white adipocyte number is most likely controlled by the rate of preadipocyte proliferation, which may contribute to the etiology of obesity. However, little is known about the molecular mechanisms that regulate preadipocyte proliferation during adipose tissue development. Necdin, which is expressed predominantly in postmitotic neurons, is a pleiotropic protein that possesses anti-mitotic and pro-survival activities. Here we show that Necdin functions as an intrinsic regulator of white preadipocyte proliferation in developing adipose tissues. Necdin is expressed in early preadipocytes or mesenchymal stem cells residing in the stromal compartment of white adipose tissues in juvenile mice. Lentivirus-mediated knockdown of endogenous Necdin expression in vivo in adipose tissues markedly increases fat mass in juvenile mice fed a high-fat diet until adulthood. Furthermore, Necdin-null mutant mice exhibit a greater expansion of adipose tissues due to adipocyte hyperplasia than wild-type mice when fed the high-fat diet during the juvenile and adult periods. Adipose stromal-vascular cells prepared from Necdin-null mice differentiate in vitro into a significantly larger number of adipocytes in response to adipogenic inducers than those from wild-type mice. These results suggest that Necdin prevents excessive preadipocyte proliferation induced b
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Promotion of mitochondrial biogenesis by Necdin protects neurons against mitochondrial insults.
Nature communications, 2016Co-Authors: Koichi Hasegawa, Kazushiro Fujiwara, Hideki Mochizuki, Toru Yasuda, Chinatsu Shiraishi, Serge Przedborski, Kazuaki YoshikawaAbstract:Neurons rely heavily on mitochondria for their function and survival. Mitochondrial dysfunction contributes to the pathogenesis of neurodegenerative diseases such as Parkinson's disease. PGC-1α is a master regulator of mitochondrial biogenesis and function. Here we identify Necdin as a potent PGC-1α stabilizer that promotes mitochondrial biogenesis via PGC-1α in mammalian neurons. Expression of genes encoding mitochondria-specific proteins decreases significantly in Necdin-null cortical neurons, where mitochondrial function and expression of the PGC-1α protein are reduced. Necdin strongly stabilizes PGC-1α by inhibiting its ubiquitin-dependent degradation. Forced expression of Necdin enhances mitochondrial function in primary cortical neurons and human SH-SY5Y neuroblastoma cells to prevent mitochondrial respiratory chain inhibitor-induced degeneration. Moreover, overexpression of Necdin in the substantia nigra in vivo of adult mice protects dopaminergic neurons against degeneration in experimental Parkinson's disease. These data reveal that Necdin promotes mitochondrial biogenesis through stabilization of endogenous PGC-1α to exert neuroprotection against mitochondrial insults.
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Necdin controls EGFR signaling linked to astrocyte differentiation in primary cortical progenitor cells
Cellular signalling, 2015Co-Authors: Izumi Fujimoto, Koichi Hasegawa, Kazushiro Fujiwara, Masashi Yamada, Kazuaki YoshikawaAbstract:Cellular signaling mediated by the EGF receptor (EGFR) plays a key role in controlling proliferation and differentiation of cortical progenitor cells (CPCs). However, regulatory mechanisms of EGFR signaling in CPCs remain largely unknown. Here we demonstrate that Necdin, a MAGE (melanoma antigen) family protein, interacts with EGFR in primary CPCs and represses its downstream signaling linked to astrocyte differentiation. EGFR was autophosphorylated and interacted with Necdin in EGF-stimulated CPCs. Necdin bound to autophosphorylated EGFR via its tyrosine kinase domain. EGF-induced phosphorylation of ERK was enhanced in Necdin-null CPCs, where the interaction between EGFR and the adaptor protein Grb2 was strengthened, suggesting that endogenous Necdin suppresses the EGFR/ERK signaling pathway in CPCs. In Necdin-null CPCs, astrocyte differentiation induced by the gliogenic cytokine cardiotrophin-1 was significantly accelerated in the presence of EGF, and inhibition of EGFR/ERK signaling abolished the acceleration. Furthermore, Necdin strongly suppressed astrocyte differentiation induced by overexpression of EGFR or its ligand binding-defective mutant equivalent to a glioblastoma-associated EGFR variant. These results suggest that Necdin acts as an intrinsic suppressor of the EGFR/ERK signaling pathway in EGF-responsive CPCs to restrain astroglial development in a cell-autonomous manner.
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Necdin Promotes Ubiquitin-Dependent Degradation of PIAS1 SUMO E3 Ligase
PloS one, 2014Co-Authors: Ibrahim Gur, Koichi Hasegawa, Kazushiro Fujiwara, Kazuaki YoshikawaAbstract:Necdin, a pleiotropic protein that promotes differentiation and survival of mammalian neurons, is a member of MAGE (melanoma antigen) family proteins that share a highly conserved MAGE homology domain. Several MAGE proteins interact with ubiquitin E3 ligases and modulate their activities. However, it remains unknown whether MAGE family proteins interact with SUMO (small ubiquitin-like modifier) E3 ligases such as PIAS (protein inhibitor of activated STAT) family, Nsmce2/Mms21 and Cbx4/Pc2. In the present study, we examined whether Necdin interacts with these SUMO E3 ligases. Co-immunoprecipitation analysis revealed that Necdin, MAGED1, MAGEF1 and MAGEL2 bound to PIAS1 but not to Nsmce2 or Cbx4. These SUMO E3 ligases bound to MAGEA1 but failed to interact with Necdin-like 2/MAGEG1. Necdin bound to PIAS1 central domains that are highly conserved among PIAS family proteins and suppressed PIAS1-dependent sumoylation of the substrates STAT1 and PML (promyelocytic leukemia protein). Remarkably, Necdin promoted degradation of PIAS1 via the ubiquitin-proteasome pathway. In transfected HEK293A cells, amino- and carboxyl-terminally truncated mutants of PIAS1 bound to Necdin but failed to undergo Necdin-dependent ubiquitination. Both PIAS1 and Necdin were associated with the nuclear matrix, where the PIAS1 terminal deletion mutants failed to localize, implying that the nuclear matrix is indispensable for Necdin-dependent ubiquitination of PIAS1. Our data suggest that Necdin suppresses PIAS1 both by inhibiting SUMO E3 ligase activity and by promoting ubiquitin-dependent degradation.
