Ablation of NF1 function in neurons induces abnormal development of cerebral cortex and reactive gliosis in the brain.
Zhu Y,Romero M I,Ghosh P,Ye Z,Charnay P,Rushing E J,Marth J D,Parada L F
Genes & development
Neurofibromatosis type 1 (NF1) is a prevalent genetic disorder that affects growth properties of neural-crest-derived cell populations. In addition, approximately one-half of NF1 patients exhibit learning disabilities. To characterize NF1 function both in vitro and in vivo, we circumvent the embryonic lethality of NF1 null mouse embryos by generating a conditional mutation in the NF1 gene using Cre/loxP technology. Introduction of a Synapsin I promoter driven Cre transgenic mouse strain into the conditional NF1 background has ablated NF1 function in most differentiated neuronal populations. These mice have abnormal development of the cerebral cortex, which suggests that NF1 has an indispensable role in this aspect of CNS development. Furthermore, although they are tumor free, these mice display extensive astrogliosis in the absence of conspicuous neurodegeneration or microgliosis. These results indicate that NF1-deficient neurons are capable of inducing reactive astrogliosis via a non-cell autonomous mechanism.
10.1101/gad.862101
Synapsin I Cre transgene expression in male mice produces germline recombination in progeny.
Rempe D,Vangeison G,Hamilton J,Li Y,Jepson M,Federoff H J
Genesis (New York, N.Y. : 2000)
The cre/LoxP system can produce conditional loss of gene function in specific cell types such as neurons. A transgenic mouse line, utilized by multiple studies, used the Synapsin I promoter to drive expression of cre (SynCre) to achieve neuronal-specific cre expression. Herein we describe that cre expression can also be observed in SynCre mice within the testes after being bred into a floxed transgenic mouse line. Cre transcript was expressed in testes resulting in recombination of the floxed substrate in testes. In the majority of cases, progeny of male SynCre mice inherited a germline recombined floxed allele, while this was never observed in progeny from female mice carrying the SynCre allele. This observation should alert investigators to a potential confound using these mice and enables male germ cell "deletor" strategies.
10.1002/gene.20183
Different Responses to a High-Fat Diet in IL-6 Conditional Knockout Mice Driven by Constitutive GFAP-Cre and Synapsin 1-Cre Expression.
Fernández-Gayol Olaya,Sanchis Paula,Aguilar Kevin,Navarro-Sempere Alicia,Comes Gemma,Molinero Amalia,Giralt Mercedes,Hidalgo Juan
Neuroendocrinology
BACKGROUND/AIMS:Interleukin-6 (IL-6) is a major cytokine controlling body weight and metabolism, at least in part through actions in the central nervous system (CNS) from local sources. METHODS:We herewith report results obtained in conditional IL-6 KO mice for brain cells (Il6ΔGfap and Il6ΔSyn). RESULTS:The reporter RiboTag mouse line demonstrated specific astrocytic expression of GFAP-dependent Cre in the hypothalamus but not in other brain areas, whereas that of synapsin 1-dependent Cre was specific for neurons. Feeding a high-fat diet (HFD) or a control diet showed that Il6ΔGfap and Il6ΔSyn mice were more prone and resistant, respectively, to HFD-induced obesity. Energy intake was not altered in HFD experiments, but it was reduced in Il6ΔSyn male mice following a 24-h fast. HFD increased circulating insulin, leptin, and cholesterol levels, decreased triglycerides, and caused impaired responses to the insulin and glucose tolerance tests. In Il6ΔGfap mice, the only significant difference observed was an increase in insulin levels of females, whereas in Il6ΔSyn mice the effects of HFD were decreased. Hypothalamic Agrp expression was significantly decreased by HFD, further decreased in Il6ΔGfap, and increased in Il6ΔSyn female mice. Hypothalamic Il-6 mRNA levels were not decreased in Il6ΔSyn mice and even increased in Il6ΔGfapmale mice. Microarray analysis of hypothalamic RNA showed that female Il6ΔGfap mice had increased interferon-related pathways and affected processes in behavior, modulation of chemical synaptic transmission, learning, and memory. CONCLUSION:The present results demonstrate that brain production of IL-6 regulates body weight in the context of caloric excess and that the cellular source is critical.
10.1159/000496845
The master negative regulator REST/NRSF controls adult neurogenesis by restraining the neurogenic program in quiescent stem cells.
Gao Zhengliang,Ure Kerstin,Ding Peiguo,Nashaat Mostafa,Yuan Laura,Ma Jing,Hammer Robert E,Hsieh Jenny
The Journal of neuroscience : the official journal of the Society for Neuroscience
Transcriptional regulation is a critical mechanism in the birth, specification, and differentiation of granule neurons in the adult hippocampus. One of the first negative-acting transcriptional regulators implicated in vertebrate development is repressor element 1-silencing transcription/neuron-restrictive silencer factor (REST/NRSF)--thought to regulate hundreds of neuron-specific genes--yet its function in the adult brain remains elusive. Here we report that REST/NRSF is required to maintain the adult neural stem cell (NSC) pool and orchestrate stage-specific differentiation. REST/NRSF recruits CoREST and mSin3A corepressors to stem cell chromatin for the regulation of pro-neuronal target genes to prevent precocious neuronal differentiation in cultured adult NSCs. Moreover, mice lacking REST/NRSF specifically in NSCs display a transient increase in adult neurogenesis that leads to a loss in the neurogenic capacity of NSCs and eventually diminished granule neurons. Our work identifies REST/NRSF as a master negative regulator of adult NSC differentiation and offers a potential molecular target for neuroregenerative approaches.
10.1523/JNEUROSCI.1604-11.2011
NRSF/REST neuronal deficient mice are more vulnerable to the neurotoxin MPTP.
Yu Mei,Suo Haiyun,Liu Ming,Cai Lei,Liu Jie,Huang Yufang,Xu Jing,Wang Yancong,Zhu Cuiqing,Fei Jian,Huang Fang
Neurobiology of aging
Parkinson's disease (PD) is characterized by progressing loss of dopaminergic neurons in the midbrain. Abnormal gene expression plays a critical role in its pathogenesis. Neuron-restrictive silencer factor (NRSF)/neuronal repressor element-1 silencing transcription factor (REST), a member of the zinc finger transcription factors, inhibits the expression of neuron-specific genes in nonneuronal cells, and regulates neurogenesis. Our previous work showed that 1-methyl-4-phenyl-pyridinium ion triggers dynamic changes of messenger RNA and protein expression of NRSF in human dopaminergic SH-SY5Y cells, and alteration of NRSF expression exacerbates 1-methyl-4-phenyl-pyridinium ion-induced cell death. The purpose of this study was to explore the in vivo role of NRSF in the progress of PD by using NRSF/REST neuron-specific conditional knockout mice (cKO). 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) was adopted to generate PD models in the cKO mice and wild type littermates. At 1, 3, 7, 14, 21, and 28 days after MPTP injection, behavioral tests were performed, and cKO mice displayed some impairments in locomotor activities. Also, the reduction of tyrosine hydroxylase protein in the striatum and the loss of dopaminergic neurons in the substantia nigra were more severe in the cKO mice. Meanwhile, the cKO mice exhibited a more dramatic depletion of striatal dopamine, accompanied by an increase in glial fibrillary acidic protein (GFAP) expression and sustained interleukin-1β transcription. These results suggested that NRSF/REST neuronal cKO mice are more vulnerable to the dopaminergic neurotoxin MPTP. Disturbance of the homeostasis of NRSF and its target genes, gliogenesis, and inflammation may contribute to the higher MPTP sensitivity in NRSF/REST neuronal cKO mice.
10.1016/j.neurobiolaging.2012.06.002
MS-275 inhibits aroclor 1254-induced SH-SY5Y neuronal cell toxicity by preventing the formation of the HDAC3/REST complex on the synapsin-1 promoter.
Formisano Luigi,Guida Natascia,Laudati Giusy,Mascolo Luigi,Di Renzo Gianfranco,Canzoniero Lorella M T
The Journal of pharmacology and experimental therapeutics
Polychlorinated biphenyl (PCB) exposure has been associated with neurodegenerative diseases, such as Parkinson's disease, amyotrophic lateral sclerosis, and dementia. Neuronal death elicited by the PCB mixture Aroclor 1254 (A1254) has been attributed to an increase in RE-1-silencing transcription factor (REST), which, in turn, correlates with a decrease in the synapsin-1 promoter gene. Although histone deacetylase (HDAC) inhibitors are known to be neuroprotective in several neurologic disorders, the core mechanisms governing this effect are not yet understood. Here, to examine how HDAC class I [N-(2-aminophenyl)-4-[N-(pyridin-3-yl-methoxycarbonyl)aminomethyl]-benzamide (MS-275)] and HDAC class II [3-[5-(3-(3-fluorophenyl)-3-oxopropen-1-yl)-1-methyl-1H-pyrrol-2-yl]-N-hydroxy-2-propenamide (MC-1568)] inhibitors prevent A1254-induced neuronal cell death, we exposed SH-SY5Y neuroblastoma cells to A1254. Exposure to A1254 (30.6 μM) for 24 and 48 hours resulted in a time-dependent cell death. Indeed, after 48 hours, MS-275, but not MC-1568, reverted A1254-induced cell death in a dose-dependent manner. Furthermore, A1254 significantly increased HDAC3, but not HDAC1 or HDAC2. Interestingly, REST physically interacted with HDAC3 after A1254 exposure. Chromatin immunoprecipitation assays revealed that MS-275 reverted the increased levels of HDAC3 binding and decreased acetylation of histone H3 within the synapsin-1 promoter region, thus reverting synapsin-1 mRNA reduction. Moreover, REST knockdown by small interfering RNA (siRNA) prevented HDAC3 from binding to the synapsin-1 promoter. Likewise, HDAC3 siRNA significantly reduced A1254-induced cell toxicity in SH-SY5Y cells and cortical neurons. Hence, this study demonstrates that inhibition of HDAC class I attenuates A1254-induced neuronal cell death by preventing HDAC3 binding and histone deacetylation within the synapsin-1 promoter region.
10.1124/jpet.114.219345
Expression of the repressor element-1 silencing transcription factor (REST) is influenced by insulin-like growth factor-I in differentiating human neuroblastoma cells.
Di Toro Rosanna,Baiula Monica,Spampinato Santi
The European journal of neuroscience
The repressor element-1 (RE-1) silencing transcription factor (REST) interacts with an RE-1 cis element and represses the transcription of neuron-specific genes in neuronal progenitors but is down-regulated in post-mitotic neurons. We report that REST expression is modified, in a time-dependent manner, in SH-SY5Y neuroblastoma cells exposed to insulin-like growth factor I (IGF-I), a polypeptide hormone affecting various aspects of neuronal induction and maturation. REST is increased in cells treated with IGF-I for 2 days and then declines in 5-day-treated cells concomitant with a progressive neurite extension. To investigate any role played by REST in neurodifferentiation by IGF-I, we employed an antisense oligonucleotide (AS-ODN) complementary to REST mRNA. In AS-ODN-treated cells, the effects elicited by IGF-I on cell proliferation are not influenced whereas a marked decrease of REST significantly increases neurite elongation without any gross perturbation of neurogenesis. Synapsin I and betaIII-tubulin gene promoters contain an RE-1 motif and their transcription is repressed by REST; both of them are increased in cells exposed to IGF-I for 5 days and further elevated by AS-ODN treatment. A parallel increase of growth cone-associated protein 43, a protein chosen as a neuronal marker not directly regulated by REST, is also observed. Therefore, REST is elevated during early steps of neural induction by IGF-I and could contribute to down-regulate genes not yet required by the differentiation program while it declines later for the acquisition of neural phenotypes. These results suggest a model in which differentiating neuroblastoma cells determine their extent of neurite outgrowth on the basis of REST disappearance.
10.1111/j.1460-9568.2004.03828.x
NCX1 is a new rest target gene: role in cerebral ischemia.
Formisano Luigi,Guida Natascia,Valsecchi Valeria,Pignataro Giuseppe,Vinciguerra Antonio,Pannaccione Anna,Secondo Agnese,Boscia Francesca,Molinaro Pasquale,Sisalli Maria Jose,Sirabella Rossana,Casamassa Antonella,Canzoniero Lorella M T,Di Renzo Gianfranco,Annunziato Lucio
Neurobiology of disease
The Na(+)-Ca(2+) exchanger 1 (NCX1), a bidirectional transporter that mediates the electrogenic exchange of one calcium ion for three sodium ions across the plasma membrane, is known to be involved in brain ischemia. Since the RE1-silencing transcription factor (REST) is a key modulator of neuronal gene expression in several neurological conditions, we studied the possible involvement of REST in regulating NCX1 gene expression and activity in stroke. We found that: (1) REST binds in a sequence specific manner and represses through H4 deacetylation, ncx1 gene in neuronal cells by recruting CoREST, but not mSin3A. (2) In neurons and in SH-SY5Y cells REST silencing by siRNA and site-direct mutagenesis of REST consensus sequence on NCX1 brain promoter determined an increase in NCX1 promoter activity. (3) By contrast, REST overexpression caused a reduction in NCX1 protein expression and activity. (4) Interestingly, in rats subjected to transient middle cerebral artery occlusion (tMCAO) and in organotypic hippocampal slices or SH-SY5Y cells exposed to oxygen and glucose deprivation (OGD) plus reoxygenation (RX), the increase in REST was associated with a decrease in NCX1. However, this reduction was reverted by REST silencing. (5) REST knocking down, along with the deriving NCX1 overexpression in the deep V and VIb cortical layers caused a marked reduction in infarct volume after tMCAO. Double silencing of REST and NCX1 completely abolished neuroprotection induced by siREST administration. Collectively, these results demonstrate that REST, by regulating NCX1 expression, may represent a potential druggable target for the treatment of brain ischemia.
10.1016/j.nbd.2012.10.010
Methylmercury upregulates RE-1 silencing transcription factor (REST) in SH-SY5Y cells and mouse cerebellum.
Guida Natascia,Laudati Giusy,Anzilotti Serenella,Sirabella Rossana,Cuomo Ornella,Brancaccio Paola,Santopaolo Marianna,Galgani Mario,Montuori Paolo,Di Renzo Gianfranco,Canzoniero Lorella M T,Formisano Luigi
Neurotoxicology
Methylmercury (MeHg) is a highly neurotoxic compound that, in adequate doses, can cause damage to the brain, including developmental defects and in severe cases cell death. The RE-1-silencing transcription factor (REST) has been found to be involved in the neurotoxic effects of environmental pollutants such as polychlorinated biphenyls (PCBs). In this study, we investigated the effects of MeHg treatment on REST expression and its role in MeHg-induced neurotoxicity in neuroblastoma SH-SY5Y cells. We found that MeHg exposure caused a dose- and time- dependent apoptotic cell death, as evidenced by the appearance of apoptotic hallmarks including caspase-3 processing and annexin V uptake. Moreover, MeHg increased REST gene and gene product expression. MeHg-induced apoptotic cell death was completely abolished by REST knockdown. Interestingly, MeHg (1μM/24h) increased the expression of REST Corepressor (Co-REST) and its binding with REST whereas the other REST cofactor mammalian SIN3 homolog A transcription regulator (mSin3A) was not modified. In addition, we demonstrated that the acetylation of histone protein H4 was reduced after MeHg treatment and was critical for MeHg-induced apoptosis. Accordingly, the pan-histone deacetylase inhibitor trichostatin-A (TSA) prevented MeHg-induced histone protein H4 deacetylation, thereby reverting MeHg-induced neurotoxic effect. Male mice subcutaneously injected with 10mg/kg of MeHg for 10 days showed an increase in REST expression in the granule cell layer of the cerebellum together with a decrease in histone H4 acetylation. Collectively, we demonstrated that methylmercury exposure can cause neurotoxicity by activating REST gene expression and H4 deacetylation.
10.1016/j.neuro.2015.11.007
α-Synuclein enhances histone H3 lysine-9 dimethylation and H3K9me2-dependent transcriptional responses.
Sugeno Naoto,Jäckel Sandra,Voigt Aaron,Wassouf Zinah,Schulze-Hentrich Julia,Kahle Philipp J
Scientific reports
α-Synuclein (αS) is a protein linked to Parkinson's disease (PD) and related neurodegenerative disorders. It is mostly localized within synapses, but αS has also been suggested to play a role in the nucleus. We used transgenic Drosophila and inducible SH-SY5Y neuroblastoma cells to investigate the effects of αS on chromatin with a particular focus on histone modifications. Overexpression of αS in male flies as well as in retinoic acid pre-treated neuroblastoma cells led to an elevation of histone H3K9 methylations, mostly mono- (H3K9me1) and di- (H3K9me2). The transient increase of H3K9 methylation in αS-induced SH-SY5Y cells was preceded by mRNA induction of the euchromatic histone lysine N-methyltransferase 2 (EHMT2). EHMT2 and H3K9me2 can function within the REST complex. Chromatin immunoprecipitation (ChIP) analyses of selected candidate, REST regulated genes showed significantly increased H3K9me2 promoter occupancy of genes encoding the L1CAM cell adhesion molecule and the synaptosomal-associated protein SNAP25, whose reduced expression levels were confirmed by RT-qPCR in αS induced cells. Treatment with EHMT inhibitor UNC0638 restored the mRNA levels of L1CAM and SNAP25. Thus, αS overexpression enhances H3K9 methylations via ΕΗΜΤ2 resulting in elevated H3K9me2 at the SNAP25 promoter, possibly affecting SNARE complex assembly and hence synaptic vesicle fusion events regulated by αS.
10.1038/srep36328
Human positive coactivator 4 controls heterochromatinization and silencing of neural gene expression by interacting with REST/NRSF and CoREST.
Das Chandrima,Gadad Shrikanth S,Kundu Tapas K
Journal of molecular biology
The highly abundant, multifunctional transcriptional positive coactivator 4 (PC4) plays important roles in transcription, replication and DNA repair. Our recent work showed that PC4 is a bona fide non-histone component of chromatin. Here, we report that knockdown of PC4 dramatically alters heterochromatin organization of the genome, accompanied by increased H3K9 (histone H3 at lysine residue 9)/14 acetylation, H3K4 trimethylation and reduction in the level of H3K9 dimethylation. These posttranslational modifications of histone H3 result in overexpression of normally silenced genes (e.g., neural genes) located in heterochromatin. The results of ChIP (chromatin immunoprecipitation) and re-ChIP assays showed that overexpression of a neuronal-specific gene is accompanied by histone hyperacetylation. We further show that PC4 interacts with heterochromatin protein 1alpha, REST/NRSF (RE1-silencing transcription factor/neuron-restrictive silencer factor) and CoREST to establish the repressed state of neural genes in nonneuronal cells. Thus, PC4 plays a crucial role in maintaining a dynamic chromatin state and heterochromatin gene silencing.
10.1016/j.jmb.2009.12.058
Screening for genes that accelerate the epigenetic aging clock in humans reveals a role for the H3K36 methyltransferase NSD1.
Genome biology
BACKGROUND:Epigenetic clocks are mathematical models that predict the biological age of an individual using DNA methylation data and have emerged in the last few years as the most accurate biomarkers of the aging process. However, little is known about the molecular mechanisms that control the rate of such clocks. Here, we have examined the human epigenetic clock in patients with a variety of developmental disorders, harboring mutations in proteins of the epigenetic machinery. RESULTS:Using the Horvath epigenetic clock, we perform an unbiased screen for epigenetic age acceleration in the blood of these patients. We demonstrate that loss-of-function mutations in the H3K36 histone methyltransferase NSD1, which cause Sotos syndrome, substantially accelerate epigenetic aging. Furthermore, we show that the normal aging process and Sotos syndrome share methylation changes and the genomic context in which they occur. Finally, we found that the Horvath clock CpG sites are characterized by a higher Shannon methylation entropy when compared with the rest of the genome, which is dramatically decreased in Sotos syndrome patients. CONCLUSIONS:These results suggest that the H3K36 methylation machinery is a key component of the epigenetic maintenance system in humans, which controls the rate of epigenetic aging, and this role seems to be conserved in model organisms. Our observations provide novel insights into the mechanisms behind the epigenetic aging clock and we expect will shed light on the different processes that erode the human epigenetic landscape during aging.
10.1186/s13059-019-1753-9
Erythropoietin in perinatal hypoxic-ischemic encephalopathy: a systematic review and meta-analysis.
Razak Abdul,Hussain Asif
Journal of perinatal medicine
Background Erythropoietin (EPO) appears to confer neuroprotection to the injured brain. Randomized clinical trials (RCTs) have demonstrated its safety in neonates with hypoxic-ischemic encephalopathy (HIE); however, the evidence is unclear. The objective of this study was to examine the role of EPO in perinatal HIE by a systematic review and meta-analysis. Methods Database search included Embase, MEDLINE, Cumulative Index to Nursing and Allied Health Literature (CINAHL) and Cochrane Central Register of Controlled Trials (CENTRAL). RCTs reporting a death, neurodevelopmental outcomes or brain injury were included. Two authors extracted the data independently from included studies and assessed the level of evidence (LOE). Results Six RCTs (EPO=5 and darbepoetin α=1) involving 454 neonates were included. A trend toward a lower risk of death was identified in infants treated with EPO [EPO with or without hypothermia: five RCTs, 368 participants, relative risk (RR) 0.74, 95% confidence interval (CI) 0.47-1.19, LOE-low; EPO without hypothermia: four RCTs, 318 participants, RR 0.89, 95% CI 0.49-1.32, LOE-low]. EPO treatment without hypothermia compared to placebo resulted in a reduced risk of cerebral palsy (two RCTs, 230 participants, RR 0.47, 95% CI 0.27-0.80, LOE-moderate) and moderate to severe cognitive impairment (two RCTs, 226 participants, RR 0.49, 95% CI 0.28-0.85, LOE-moderate). A reduced risk of brain injury was identified in EPO treated infants (EPO with or without hypothermia, two RCTs, 148 participants, RR 0.70, 95% CI 0.53-0.92, LOE-moderate). Conclusion EPO administration in neonates with perinatal HIE reduces the risk of brain injury, cerebral palsy and cognitive impairment. The evidence is limited to suggest its role as an adjuvant to hypothermia. Larger powered trials are underway to overcome this limitation.
10.1515/jpm-2018-0360
TET3 is recruited by REST for context-specific hydroxymethylation and induction of gene expression.
Perera Arshan,Eisen David,Wagner Mirko,Laube Silvia K,Künzel Andrea F,Koch Susanne,Steinbacher Jessica,Schulze Elisabeth,Splith Victoria,Mittermeier Nana,Müller Markus,Biel Martin,Carell Thomas,Michalakis Stylianos
Cell reports
Ten-eleven translocation hydroxylases (TET1-3) oxidize 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC). In neurons, increased 5hmC levels within gene bodies correlate positively with gene expression. The mechanisms controlling TET activity and 5hmC levels are poorly understood. In particular, it is not known how the neuronal TET3 isoform lacking a DNA-binding domain is targeted to the DNA. To identify factors binding to TET3, we screened for proteins that co-precipitate with TET3 from mouse retina and identified the transcriptional repressor REST as a highly enriched TET3-specific interactor. REST was able to enhance TET3 hydroxylase activity after co-expression and overexpression of TET3-activated transcription of REST target genes. Moreover, we found that TET3 also interacts with NSD3 and two other H3K36 methyltransferases and is able to induce H3K36 trimethylation. We propose a mechanism for transcriptional activation in neurons that involves REST-guided targeting of TET3 to the DNA for directed 5hmC generation and NSD3-mediated H3K36 trimethylation.
10.1016/j.celrep.2015.03.020