Differential vulnerability of adult neurogenic niches to dosage of the neurodevelopmental-disorder linked gene Foxg1.

The transcription factor FOXG1 serves pleiotropic functions in brain development ranging from the regulation of precursor proliferation to the control of cortical circuit formation. Loss-of-function mutations and duplications of FOXG1 are associated with neurodevelopmental disorders in humans illustrating the importance of FOXG1 dosage for brain development. Aberrant FOXG1 dosage has been found to disrupt the balanced activity of glutamatergic and GABAergic neurons, but the underlying mechanisms are not fully understood. We report that FOXG1 is expressed in the main adult neurogenic niches in mice, i.e. the hippocampal dentate gyrus and the subependymal zone/olfactory bulb system, where neurogenesis of glutamatergic and GABAergic neurons persists into adulthood. These niches displayed differential vulnerability to increased FOXG1 dosage: high FOXG1 levels severely compromised survival and glutamatergic dentate granule neuron fate acquisition in the hippocampal neurogenic niche, but left neurogenesis of GABAergic neurons in the subependymal zone/olfactory bulb system unaffected. Comparative transcriptomic analyses revealed a significantly higher expression of the apoptosis-linked nuclear receptor Nr4a1 in FOXG1-overexpressing hippocampal neural precursors. Strikingly, pharmacological interference with NR4A1 function rescued FOXG1-dependent death of hippocampal progenitors. Our results reveal differential vulnerability of neuronal subtypes to increased FOXG1 dosage and suggest that activity of a FOXG1/NR4A1 axis contributes to such subtype-specific response.

Dentate gyrus astrocytes exhibit layer-specific molecular, morphological and physiological features.

Neuronal heterogeneity has been established as a pillar of higher central nervous system function, but glial heterogeneity and its implications for neural circuit function are poorly understood. Here we show that the adult mouse dentate gyrus (DG) of the hippocampus is populated by molecularly distinct astrocyte subtypes that are associated with distinct DG layers. Astrocytes localized to different DG compartments also exhibit subtype-specific morphologies. Physiologically, astrocytes in upper DG layers form large syncytia, while those in lower DG compartments form smaller networks. Astrocyte subtypes differentially express glutamate transporters, which is associated with different amplitudes of glutamate transporter-mediated currents. Key molecular and morphological features of astrocyte diversity in the mice DG are conserved in humans. This adds another layer of complexity to our understanding of brain network composition and function, which will be crucial for further studies on astrocytes in health and disease.

Astrogenesis in the murine dentate gyrus is a life-long and dynamic process.

Astrocytes are highly abundant in the mammalian brain, and their functions are of vital importance for all aspects of development, adaption, and aging of the central nervous system (CNS). Mounting evidence indicates the important contributions of astrocytes to a wide range of neuropathies. Still, our understanding of astrocyte development significantly lags behind that of other CNS cells. We here combine immunohistochemical approaches with genetic fate-mapping, behavioural paradigms, single-cell transcriptomics, and in vivo two-photon imaging, to comprehensively assess the generation and the proliferation of astrocytes in the dentate gyrus (DG) across the life span of a mouse. Astrogenesis in the DG is initiated by radial glia-like neural stem cells giving rise to locally dividing astrocytes that enlarge the astrocyte compartment in an outside-in-pattern. Also in the adult DG, the vast majority of astrogenesis is mediated through the proliferation of local astrocytes. Interestingly, locally dividing astrocytes were able to adapt their proliferation to environmental and behavioral stimuli revealing an unexpected plasticity. Our study establishes astrocytes as enduring plastic elements in DG circuits, implicating a vital contribution of astrocyte dynamics to hippocampal plasticity.

scRNA sequencing uncovers a TCF4-dependent transcription factor network regulating commissure development in mouse.

Transcription factor 4 (TCF4) is a crucial regulator of neurodevelopment and has been linked to the pathogenesis of autism, intellectual disability and schizophrenia. As a class I bHLH transcription factor (TF), it is assumed that TCF4 exerts its neurodevelopmental functions through dimerization with proneural class II bHLH TFs. Here, we aim to identify TF partners of TCF4 in the control of interhemispheric connectivity formation. Using a new bioinformatic strategy integrating TF expression levels and regulon activities from single cell RNA-sequencing data, we find evidence that TCF4 interacts with non-bHLH TFs and modulates their transcriptional activity in Satb2+ intercortical projection neurons. Notably, this network comprises regulators linked to the pathogenesis of neurodevelopmental disorders, e.g. FOXG1, SOX11 and BRG1. In support of the functional interaction of TCF4 with non-bHLH TFs, we find that TCF4 and SOX11 biochemically interact and cooperatively control commissure formation in vivo, and regulate the transcription of genes implicated in this process. In addition to identifying new candidate interactors of TCF4 in neurodevelopment, this study illustrates how scRNA-Seq data can be leveraged to predict TF networks in neurodevelopmental processes.

Enriched environment ameliorates adult hippocampal neurogenesis deficits in Tcf4 haploinsufficient mice.

BACKGROUND: Transcription factor 4 (TCF4) has been linked to human neurodevelopmental disorders such as intellectual disability, Pitt-Hopkins Syndrome (PTHS), autism, and schizophrenia. Recent work demonstrated that TCF4 participates in the control of a wide range of neurodevelopmental processes in mammalian nervous system development including neural precursor proliferation, timing of differentiation, migration, dendritogenesis and synapse formation. TCF4 is highly expressed in the adult hippocampal dentate gyrus - one of the few brain regions where neural stem / progenitor cells generate new functional neurons throughout life. RESULTS: We here investigated whether TCF4 haploinsufficiency, which in humans causes non-syndromic forms of intellectual disability and PTHS, affects adult hippocampal neurogenesis, a process that is essential for hippocampal plasticity in rodents and potentially in humans. Young adult Tcf4 heterozygote knockout mice showed a major reduction in the level of adult hippocampal neurogenesis, which was at least in part caused by lower stem/progenitor cell numbers and impaired maturation and survival of adult-generated neurons. Interestingly, housing in an enriched environment was sufficient to enhance maturation and survival of new neurons and to substantially augment neurogenesis levels in Tcf4 heterozygote knockout mice. CONCLUSION: The present findings indicate that haploinsufficiency for the intellectual disability- and PTHS-linked transcription factor TCF4 not only affects embryonic neurodevelopment but impedes neurogenesis in the hippocampus of adult mice. These findings suggest that TCF4 haploinsufficiency may have a negative impact on hippocampal function throughout adulthood by impeding hippocampal neurogenesis.

ß-catenin signaling modulates the tempo of dendritic growth of adult-born hippocampal neurons.

In adult hippocampal neurogenesis, stem/progenitor cells generate dentate granule neurons that contribute to hippocampal plasticity. The establishment of a morphologically defined dendritic arbor is central to the functional integration of adult-born neurons. We investigated the role of canonical Wnt/ß-catenin signaling in dendritogenesis of adult-born neurons. We show that canonical Wnt signaling follows a biphasic pattern, with high activity in stem/progenitor cells, attenuation in immature neurons, and reactivation during maturation, and demonstrate that this activity pattern is required for proper dendrite development. Increasing ß-catenin signaling in maturing neurons of young adult mice transiently accelerated dendritic growth, but eventually produced dendritic defects and excessive spine numbers. In middle-aged mice, in which protracted dendrite and spine development were paralleled by lower canonical Wnt signaling activity, enhancement of ß-catenin signaling restored dendritic growth and spine formation to levels observed in young adult animals. Our data indicate that precise timing and strength of ß-catenin signaling are essential for the correct functional integration of adult-born neurons and suggest Wnt/ß-catenin signaling as a pathway to ameliorate deficits in adult neurogenesis during aging.

Transcription factor Tcf4 is the preferred heterodimerization partner for Olig2 in oligodendrocytes and required for differentiation.

Development of oligodendrocytes and myelin formation in the vertebrate central nervous system is under control of several basic helix-loop-helix transcription factors such as Olig2, Ascl1, Hes5 and the Id proteins. The class I basic helix-loop-helix proteins Tcf3, Tcf4 and Tcf12 represent potential heterodimerization partners and functional modulators for all, but have not been investigated in oligodendrocytes so far. Using mouse mutants, organotypic slice and primary cell cultures we here show that Tcf4 is required in a cell-autonomous manner for proper terminal differentiation and myelination in vivo and ex vivo. Partial compensation is provided by the paralogous Tcf3, but not Tcf12. On the mechanistic level Tcf4 was identified as the preferred heterodimerization partner of the central regulator of oligodendrocyte development Olig2. Both genetic studies in the mouse as well as functional studies on enhancer regions of myelin genes confirmed the relevance of this physical interaction for oligodendrocyte differentiation. Considering that alterations in TCF4 are associated with syndromic and non-syndromic forms of intellectual disability, schizophrenia and autism in humans, our findings point to the possibility of an oligodendroglial contribution to these disorders.

Sox11 is an Activity-Regulated Gene with Dentate-Gyrus-Specific Expression Upon General Neural Activation.

Neuronal activity initiates transcriptional programs that shape long-term changes in plasticity. Although neuron subtypes differ in their plasticity response, most activity-dependent transcription factors (TFs) are broadly expressed across neuron subtypes and brain regions. Thus, how region- and neuronal subtype-specific plasticity are established on the transcriptional level remains poorly understood. We report that in young adult (i.e., 6-8 weeks old) mice, the developmental TF SOX11 is induced in neurons within 6 h either by electroconvulsive stimulation or by exploration of a novel environment. Strikingly, SOX11 induction was restricted to the dentate gyrus (DG) of the hippocampus. In the novel environment paradigm, SOX11 was observed in a subset of c-FOS expressing neurons (ca. 15%); whereas around 75% of SOX11+ DG granule neurons were c-FOS+, indicating that SOX11 was induced in an activity-dependent fashion in a subset of neurons. Environmental enrichment or virus-mediated overexpression of SOX11 enhanced the excitability of DG granule cells and downregulated the expression of different potassium channel subunits, whereas conditional Sox11/4 knock-out mice presented the opposite phenotype. We propose that Sox11 is regulated in an activity-dependent fashion, which is specific to the DG, and speculate that activity-dependent Sox11 expression may participate in the modulation of DG neuron plasticity.

Phosphorylation of the neurogenic transcription factor SOX11 on serine 133 modulates neuronal morphogenesis.

The intellectual disability gene, Sox11, encodes for a critical neurodevelopmental transcription factor with functions in precursor survival, neuronal fate determination, migration and morphogenesis. The mechanisms regulating SOX11's activity remain largely unknown. Mass spectrometric analysis uncovered that SOX11 can be post-translationally modified by phosphorylation. Here, we report that phosphorylatable serines surrounding the high-mobility group box modulate SOX11's transcriptional activity. Through Mass Spectrometry (MS), co-immunoprecipitation assays and in vitro phosphorylation assays followed by MS we verified that protein kinase A (PKA) interacts with SOX11 and phosphorylates it on S133. In vivo replacement of SoxC factors in developing adult-generated hippocampal neurons with SOX11 S133 phospho-mutants indicated that phosphorylation on S133 modulates dendrite development of adult-born dentate granule neurons, while reporter assays suggested that S133 phosphorylation fine-tunes the activation of select target genes. These data provide novel insight into the control of the critical neurodevelopmental regulator SOX11 and imply SOX11 as a mediator of PKA-regulated neuronal development.

Phosphorylation Modulates the Subcellular Localization of SOX11.

SOX11 is a key Transcription Factor (TF) in the regulation of embryonic and adult neurogenesis, whose mutation has recently been linked to an intellectual disability syndrome in humans. SOX11's transient activity during neurogenesis is critical to ensure the precise execution of the neurogenic program. Here, we report that SOX11 displays differential subcellular localizations during the course of neurogenesis. Western-Blot analysis of embryonic mouse brain lysates indicated that SOX11 is post-translationally modified by phosphorylation. Using Mass Spectrometry, we found 10 serine residues in the SOX11 protein that are putatively phosphorylated. Systematic analysis of phospho-mutant SOX11 resulted in the identification of the S30 residue, whose phosphorylation promotes nuclear over cytoplasmic localization of SOX11. Collectively, these findings uncover phosphorylation as a novel layer of regulation of the intellectual disability gene Sox11.

Analysis of the expression pattern of the schizophrenia-risk and intellectual disability gene TCF4 in the developing and adult brain suggests a role in development and plasticity of cortical and hippocampal neurons.

Background: Haploinsufficiency of the class I bHLH transcription factor TCF4 causes Pitt-Hopkins syndrome (PTHS), a severe neurodevelopmental disorder, while common variants in the TCF4 gene have been identified as susceptibility factors for schizophrenia. It remains largely unknown, which brain regions are dependent on TCF4 for their development and function. Methods: We systematically analyzed the expression pattern of TCF4 in the developing and adult mouse brain. We used immunofluorescent staining to identify candidate regions whose development and function depend on TCF4. In addition, we determined TCF4 expression in the developing rhesus monkey brain and in the developing and adult human brain through analysis of transcriptomic datasets and compared the expression pattern between species. Finally, we morphometrically and histologically analyzed selected brain structures in Tcf4-haploinsufficient mice and compared our morphometric findings to neuroanatomical findings in PTHS patients. Results: TCF4 is broadly expressed in cortical and subcortical structures in the developing and adult mouse brain. The TCF4 expression pattern was highly similar between humans, rhesus monkeys, and mice. Moreover, Tcf4 haploinsufficiency in mice replicated structural brain anomalies observed in PTHS patients. Conclusion: Our data suggests that TCF4 is involved in the development and function of multiple brain regions and indicates that its regulation is evolutionary conserved. Moreover, our data validate Tcf4-haploinsufficient mice as a model to study the neurodevelopmental basis of PTHS.