Embryonic Cortical Neural Stem Cells Research Articles

Epitranscriptomic regulation of cortical neurogenesis via Mettl8-dependent mitochondrial tRNA m3C modification.

Increasing evidence implicates the critical roles of various epitranscriptomic RNA modifications in different biological processes. Methyltransferase METTL8 installs 3-methylcytosine (m3C) modification of mitochondrial tRNAs invitro; however, its role in intact biological systems is unknown. Here, we show that Mettl8 is localized in mitochondria and installs m3C specifically on mitochondrial tRNAThr/Ser(UCN) in mouse embryonic cortical neural stem cells. At molecular and cellular levels, Mettl8 deletion in cortical neural stem cells leads to reduced mitochondrial protein translation and attenuated respiration activity. At the functional level, conditional Mettl8 deletion in mice results in impaired embryonic cortical neural stem cell maintenance invivo, which can be rescued by pharmacologically enhancing mitochondrial functions. Similarly, METTL8 promotes mitochondrial protein expression and neural stem cell maintenance in human forebrain cortical organoids. Together, our study reveals a conserved epitranscriptomic mechanism of Mettl8 and mitochondrial tRNA m3C modification in maintaining embryonic cortical neural stem cells in mice and humans.

LncRNA RUS shapes the gene expression program towards neurogenesis.

The evolution of brain complexity correlates with an increased expression of long, noncoding (lnc) RNAs in neural tissues. Although prominent examples illustrate the potential of lncRNAs to scaffold and target epigenetic regulators to chromatin loci, only few cases have been described to function during brain development. We present a first functional characterization of the lncRNA LINC01322, which we term RUS for "RNA upstream of Slitrk3." The RUS gene is well conserved in mammals by sequence and synteny next to the neurodevelopmental gene Slitrk3. RUS is exclusively expressed in neural cells and its expression increases during neuronal differentiation of mouse embryonic cortical neural stem cells. Depletion of RUS locks neuronal precursors in an intermediate state towards neuronal differentiation resulting in arrested cell cycle and increased apoptosis. RUS associates with chromatin in the vicinity of genes involved in neurogenesis, most of which change their expression upon RUS depletion. The identification of a range of epigenetic regulators as specific RUS interactors suggests that the lncRNA may mediate gene activation and repression in a highly context-dependent manner.

Effects of Panax notoginseng saponins on proliferation and differentiation of rat embryonic cortical neural stem cells

BackgroundWe aimed to study the effect of Panax notoginseng saponins (PNS) on the proliferation, differentiation, self-renewal, and expressions of basic fibroblast growth factor (bFGF) and brain-derived neurotrophic factor (BDNF) in rat embryonic neural stem cells (NSCs). MethodsCortical stem cells were isolated from rat embryos on Embryonic Day 17 (E17) and identified by nestin expression. Subsequently, primary culture, subculturing, and single cell cloning were performed on the cells. After the first cell passage (P1), the cells were resuspended and divided into a control group and a treatment group. Control cells were cultured in serum-free basal culture medium with B27 and dulbecco's modified eagle medium (DMEM)/F12. The same medium supplemented with PNS (100 μg/mL) was used to culture cells in the treatment group. Both groups were incubated at 37°C in a 5% CO2 incubator. Immunocytochemistry was performed 4 days after incubation. ResultsPrimary, P1, and P2 cells in the treatment group formed neurospheres, as did single cell clones of the P1 cells in this group. After being cultured for 4 days, the number of nestin-, proliferating cell nuclear antigen (PCNA)-, Tuj-1-, neurofilament (NF)-, vimentin-, glial fibrillary acidic protein (GFAP)-, bFGF-, and BDNF-positive cells significantly increased in the treatment group in comparison to the control group. All positively stained cells could form clear clusters. ConclusionPNS can promote rat embryonic cortical NSC survival, self-renewal, proliferation, and differentiation through neurotrophic factors by autocrine or paracrine signaling.

Tanshinone II A, a multiple target neuroprotectant, promotes caveolae-dependent neuronal differentiation

Neuron loss is one fundamental features of neurodegenerative diseases. Stimulating endogenous neurogenesis, especially neuronal differentiation, might potentially provide therapeutic effects to these diseases. In this study, tanshinone II A (TIIA), a multiple target neuroprotectant, was demonstrated to promote dose-dependent neuronal differentiation in three cell models of immortalized C17.2 neuronal stem cells, rat embryonic cortical neural stem cells (NSCs) and rat PC12 pheochromocytoma cells. In particular, TIIA exerted promising effects on NSCs even at the dose of 3nM. In PC12 cells, TIIA activated mitogen-activated protein kinase 42/44 (MAPK42/44) and its downstream transcription factor, cAMP response element-binding protein (CREB). In addition, TIIA up-regulated the expressions of brain derived neurotrophic factor (BDNF) and nerve growth factor (NGF). The MEK inhibitor and the antagonist to the receptors of NGF and BDNF could partially attenuate the differentiation effects, indicating that MAPK42/44 mediated BDNF and NGF signals were involved in TIIA's differentiation effects. Caveolin-1 (CAV-1), the major functional protein of membrane caveolae, plays critical roles in the endocytosis of exogenous materials. CAV1, which was activated by TIIA, might help TIIA transport across cell membrane to initiate its differentiation effects. It was proven by the evidences that suppressing the function of caveolin inhibited the differentiation effects of TIIA. Therefore, we concluded that TIIA promoted neuronal differentiation partially through MAPK42/44 mediated BDNF and NGF signals in a caveolae-dependent manner.

SoxB, cell cycle and neurogenesis

SoxB, cell cycle and neurogenesis

Flexibility of Neural Stem Cells

Embryonic cortical neural stem cells are self-renewing progenitors that can differentiate into neurons and glia. We generated neurospheres from the developing cerebral cortex using a mouse genetic model that allows for lineage selection and found that the self-renewing neural stem cells are restricted to Sox2 expressing cells. Under normal conditions, embryonic cortical neurospheres are heterogeneous with regard to Sox2 expression and contain astrocytes, neural stem cells, and neural progenitor cells sufficiently plastic to give rise to neural crest cells when transplanted into the hindbrain of E1.5 chick and E8 mouse embryos. However, when neurospheres are maintained under lineage selection, such that all cells express Sox2, neural stem cells maintain their Pax6+ cortical radial glia identity and exhibit a more restricted fate in vitro and after transplantation. These data demonstrate that Sox2 preserves the cortical identity and regulates the plasticity of self-renewing Pax6+ radial glia cells.

TAp73 Acts via the bHLH Hey2 to Promote Long-Term Maintenance of Neural Precursors

Increasing evidence suggests that deficits in adult stem cell maintenance cause aberrant tissue repair and premature aging [1]. While the mechanisms regulating stem cell longevity are largely unknown, recent studies have implicated p53 and its family member p63. Both proteins regulate organismal aging [2-4] as well as survival and self-renewal of tissue stem cells [5-9]. Intriguingly, haploinsufficiency for a third family member, p73, causes age-related neurodegeneration [10]. While this phenotype is at least partially due to loss of the ΔNp73 isoform, a potent neuronal prosurvival protein [11-16], a recent study showed that mice lacking the other p73 isoform, TAp73, have perturbations in the hippocampal dentate gyrus [17], a major neurogenic site in the adult brain. These findings, and the link between the p53 family, stem cells, and aging, suggest that TAp73 might play a previously unanticipated role in maintenance of neural stem cells. Here, we have tested this hypothesis and show that TAp73 ensures normal adult neurogenesis by promoting the long-term maintenance of neural stem cells. Moreover, we show that TAp73 does this by transcriptionally regulating the bHLH Hey2, which itself promotes neural precursor maintenance by preventing premature differentiation.

Expression of the Parkinson's Disease-Related Protein DJ-1 during Neural Stem Cell Proliferation

Neural stem cells (NSCs) arise through neurogenesis, and comprise all newly-formed neurons in the central and peripheral nervous systems. DJ-1 is associated with autosomal recessive familial Parkinson's disease (PD). The relationship between DJ-1 and NSC proliferation may shed light on the underlying pathogenesis of, and potential treatments for, PD. To investigate the relationship between DJ-1 and NSCs, embryonic cortical NSCs were isolated and cultured from E14 fetal rats. Immunocytochemistry, flow cytometry, and immunohistochemistry were applied to evaluate DJ-1 expression in proliferating NSCs. We found that DJ-1 was co-expressed with nestin, a marker of progenitors, during NSC proliferation from days 1-7. The present results suggest that DJ-1 is co-expressed with nestin in NSCs during proliferation.

Distinct Temporal Requirements for the Homeobox Gene Gsx2 in Specifying Striatal and Olfactory Bulb Neuronal Fates

The homeobox gene Gsx2 (formerly Gsh2) is known to be required for striatal and olfactory bulb neurogenesis; however, its specific role in the specification of these two neuronal subtypes remains unclear. To address this, we have employed a temporally regulated gain-of-function approach in transgenic mice and found that misexpression of Gsx2 at early stages of telencephalic neurogenesis favors the specification of striatal projection neuron identity over that of olfactory bulb interneurons. In contrast, delayed activation of the Gsx2 transgene until later stages exclusively promotes olfactory bulb interneuron identity. In a complementary approach, we have conditionally inactivated Gsx2 in a temporally progressive manner. Unlike germline Gsx2 mutants, which exhibit severe alterations in both striatal and olfactory bulb neurogenesis at birth, the conditional mutants exhibited defects restricted to olfactory bulb interneurons. These results demonstrate that Gsx2 specifies striatal projection neuron and olfactory bulb interneuron identity at distinct time points during telencephalic neurogenesis.

Embryonic cortical neural stem cells migrate ventrally and persist as postnatal striatal stem cells

Embryonic cortical neural stem cells apparently have a transient existence, as they do not persist in the adult cortex. We sought to determine the fate of embryonic cortical stem cells by following Emx1IREScre; LacZ/EGFP double-transgenic murine cells from midgestation into adulthood. Lineage tracing in combination with direct cell labeling and time-lapse video microscopy demonstrated that Emx1-lineage embryonic cortical stem cells migrate ventrally into the striatal germinal zone (GZ) perinatally and intermingle with striatal stem cells. Upon integration into the striatal GZ, cortical stem cells down-regulate Emx1 and up-regulate Dlx2, which is a homeobox gene characteristic of the developing striatum and striatal neural stem cells. This demonstrates the existence of a novel dorsal-to-ventral migration of neural stem cells in the perinatal forebrain.

A specific role for the TFIID subunit TAF4 and RanBPM in neural progenitor differentiation

TAF4 is crucial for the activity of many transcription factors, including CREB, RAR and CSL/RBP-Jκ, but the role for TAF4 in neural development is unknown. Embryonic cortical neural stem cells (NSC) showed strong expression of TAF4 that decreased during neuronal but not glial differentiation. In a protein–protein interaction screen, we identified the intracellular signaling factor RanBPM as a co-factor of TAF4. RanBPM co-localized with TAF4 in a subset of mitotic progenitors in vivo and endogenous TAF4 and RanBPM could be co-immunoprecipitated from NSC extracts. Interestingly, co-transfections of TAF4 and RanBPM led to a significant increase in the number of primary neurite processes but no increase in total neurite length, whereas RanBPM and a TAF4 isoform lacking the RanBPM-interacting domain exerted no significant effect. Our results demonstrate that temporally high expression levels of two factors considered to be relatively general in function can influence very specific events in neuronal differentiation.