The cerebral cortex underwent rapid expansion and increased complexity during recent hominid evolution. Gene duplications constitute a major evolutionary force, but their impact on human brain ...development remains unclear. Using tailored RNA sequencing (RNA-seq), we profiled the spatial and temporal expression of hominid-specific duplicated (HS) genes in the human fetal cortex and identified a repertoire of 35 HS genes displaying robust and dynamic patterns during cortical neurogenesis. Among them NOTCH2NL, human-specific paralogs of the NOTCH2 receptor, stood out for their ability to promote cortical progenitor maintenance. NOTCH2NL promote the clonal expansion of human cortical progenitors, ultimately leading to higher neuronal output. At the molecular level, NOTCH2NL function by activating the Notch pathway through inhibition of cis Delta/Notch interactions. Our study uncovers a large repertoire of recently evolved genes active during human corticogenesis and reveals how human-specific NOTCH paralogs may have contributed to the expansion of the human cortex.
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•Identification of >35 HS protein-coding genes expressed during human corticogenesis•NOTCH2NL human-specific paralogs of NOTCH2 expressed in human cortical progenitors•NOTCH2NL genes expand human cortical progenitors and their neuronal output•NOTCH2NL promotes Notch signaling through cis-inhibition of Delta/Notch interactions
Human-specific NOTCH2NL expands cortical progenitors and neuronal output and thus may have contributed to the expansion of the human cortex.
The cerebral cortex develops through the coordinated generation of dozens of neuronal subtypes, but the mechanisms involved remain unclear. Here we show that mouse embryonic stem cells, cultured ...without any morphogen but in the presence of a sonic hedgehog inhibitor, recapitulate in vitro the major milestones of cortical development, leading to the sequential generation of a diverse repertoire of neurons that display most salient features of genuine cortical pyramidal neurons. When grafted into the cerebral cortex, these neurons develop patterns of axonal projections corresponding to a wide range of cortical layers, but also to highly specific cortical areas, in particular visual and limbic areas, thereby demonstrating that the identity of a cortical area can be specified without any influence from the brain. The discovery of intrinsic corticogenesis sheds new light on the mechanisms of neuronal specification, and opens new avenues for the modelling and treatment of brain diseases.
The primary cilium is a central signaling component during embryonic development. Here we focus on CROCCP2, a hominid-specific gene duplicate from ciliary rootlet coiled coil (CROCC), also known as ...rootletin, that encodes the major component of the ciliary rootlet. We find that CROCCP2 is highly expressed in the human fetal brain and not in other primate species. CROCCP2 gain of function in the mouse embryonic cortex and human cortical cells and organoids results in decreased ciliogenesis and increased cortical progenitor amplification, particularly basal progenitors. CROCCP2 decreases ciliary dynamics by inhibition of the IFT20 ciliary trafficking protein, which then impacts neurogenesis through increased mTOR signaling. Loss of function of CROCCP2 in human cortical cells and organoids leads to increased ciliogenesis, decreased mTOR signaling, and impaired basal progenitor amplification. These data identify CROCCP2 as a human-specific modifier of cortical neurogenesis that acts through modulation of ciliary dynamics and mTOR signaling.
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•CROCCP2: a hominid-specific gene duplicate uniquely expressed in human fetal cortex•CROCCP2 overexpression in the mouse embryo leads to cortical progenitor amplification•CROCCP2 is required for basal progenitor amplification in human cortical organoids•CROCCP2 acts by decreasing ciliogenesis and enhancing the mTOR pathway
van Heurck et al. identify CROCCP2, a hominid-specific gene duplicate, as a human-specific modifier of neurogenesis in the developing cerebral cortex. They find that CROCCP2 is necessary and sufficient to enhance human cortical progenitor amplification and acts by decreasing primary cilia dynamics and enhancing the mTOR pathway.
The study of human cortical development has major implications for brain evolution and diseases but has remained elusive due to paucity of experimental models. Here we found that human embryonic stem ...cells (ESCs) and induced pluripotent stem cells (iPSCs), cultured without added morphogens, recapitulate corticogenesis leading to the sequential generation of functional pyramidal neurons of all six layer identities. After transplantation into mouse neonatal brain, human ESC-derived cortical neurons integrated robustly and established specific axonal projections and dendritic patterns corresponding to native cortical neurons. The differentiation and connectivity of the transplanted human cortical neurons complexified progressively over several months in vivo, culminating in the establishment of functional synapses with the host circuitry. Our data demonstrate that human cortical neurons generated in vitro from ESC/iPSC can develop complex hodological properties characteristic of the cerebral cortex in vivo, thereby offering unprecedented opportunities for the modeling of human cortex diseases and brain repair.
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► Cortical neurogenesis from human ESC/iPSC without added morphogens ► Specific axonal and dendritic patterns of grafted human ESC-derived neurons ► Functional synapses between transplanted neurons and the host cortex ► Human ESC/iPSC corticogenesis recapitulates species-specific temporality
The study of human cortical development has major implications for brain evolution and diseases. Espuny-Camacho et al. describe how human pluripotent stem cells can be converted in vitro into functional pyramidal neurons, which after transplantation integrate like native neurons into the mouse brain circuits.
The transplantation of pluripotent stem-cell-derived neurons constitutes a promising avenue for the treatment of several brain diseases. However, their potential for the repair of the cerebral cortex ...remains unclear, given its complexity and neuronal diversity. Here, we show that human visual cortical cells differentiated from embryonic stem cells can be transplanted and can integrate successfully into the lesioned mouse adult visual cortex. The transplanted human neurons expressed the appropriate repertoire of markers of six cortical layers, projected axons to specific visual cortical targets, and were synaptically active within the adult brain. Moreover, transplant maturation and integration were much less efficient following transplantation into the lesioned motor cortex, as previously observed for transplanted mouse cortical neurons. These data constitute an important milestone for the potential use of human PSC-derived cortical cells for the reassembly of cortical circuits and emphasize the importance of cortical areal identity for successful transplantation.
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•Human PSC-derived cortical neurons efficiently integrate into the adult mouse brain•PSC-derived human neurons reestablish axonal pathways in the lesioned adult cortex•Restoration of cortical pathways requires a donor and recipient area-identity match
Espuny-Camacho et al. show that transplanted ESC-derived human cortical neurons integrate functionally into the lesioned adult mouse brain. Transplanted neurons display visual cortical identity and show specific restoration of damaged cortical pathways following transplantation into the visual but not the motor cortex, suggesting the importance of areal-identity match for successful cortical repair.
During neurogenesis, neural stem/progenitor cells (NPCs) undergo an irreversible fate transition to become neurons. The Notch pathway is important for this process, and repression of Notch-dependent ...Hes genes is essential for triggering differentiation. However, Notch signaling often remains active throughout neuronal differentiation, implying a change in the transcriptional responsiveness to Notch during the neurogenic transition. We identified Bcl6, an oncogene, as encoding a proneurogenic factor that is required for proper neurogenesis of the mouse cerebral cortex. BCL6 promoted the neurogenic conversion by switching the composition of Notch-dependent transcriptional complexes at the Hes5 promoter. BCL6 triggered exclusion of the co-activator Mastermind-like 1 and recruitment of the NAD(+)-dependent deacetylase Sirt1, which was required for BCL6-dependent neurogenesis. The resulting epigenetic silencing of Hes5 led to neuronal differentiation despite active Notch signaling. Our findings suggest a role for BCL6 in neurogenesis and uncover Notch-BCL6-Sirt1 interactions that may affect other aspects of physiology and disease.
Embryonic stem cells (ESCs) constitute a tool of great potential in neurobiology, enabling the directed differentiation of specific neural cell types. We have shown recently that neurons of the ...cerebral cortex can be generated from mouse ESCs cultured in a chemically defined medium that contains no morphogen, but in the presence of the sonic hedgehog inhibitor cyclopamine. Corticogenesis from ESCs recapitulates the most important steps of cortical development, leading to the generation of multipotent cortical progenitors that sequentially produce cortical pyramidal neurons displaying distinct layer-specific identities. The protocol provides a most reductionist cellular model to tackle the complex mechanisms of cortical development and function, thereby opening new perspectives for the modeling of cortical diseases and the design of novel neurological treatments, while offering an alternative to animal use. In this protocol, we describe a method by which millions of cortical neurons can be generated in 2-3 weeks, starting from a single frozen vial of ESCs.
During neurogenesis, progenitors switch from self-renewal to differentiation through the interplay of intrinsic and extrinsic cues, but how these are integrated remains poorly understood. Here, we ...combine whole-genome transcriptional and epigenetic analyses with in vivo functional studies to demonstrate that Bcl6, a transcriptional repressor previously reported to promote cortical neurogenesis, acts as a driver of the neurogenic transition through direct silencing of a selective repertoire of genes belonging to multiple extrinsic pathways promoting self-renewal, most strikingly the Wnt pathway. At the molecular level, Bcl6 represses its targets through Sirt1 recruitment followed by histone deacetylation. Our data identify a molecular logic by which a single cell-intrinsic factor represses multiple extrinsic pathways that favor self-renewal, thereby ensuring robustness of neuronal fate transition.
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•Bcl6 ensures robust neurogenesis by repressing major extrinsic self-renewal pathways•Bcl6 inhibits the Notch, Wnt, SHH, and FGF signaling pathways at multiple levels•Bcl6 represses transcription through Sirt1 recruitment and histone deacetylation
Bonnefont et al. show that Bcl6 promotes neurogenesis by directly repressing genes belonging to the major signaling pathways promoting cortical progenitor self-renewal. These data indicate that a single cell-intrinsic factor represses multiple extrinsic signaling pathways to ensure irreversible neurogenic commitment.
Disrupted differentiation during development can lead to oncogenesis, but the underlying mechanisms remain poorly understood. Here we identify BCL6, a transcriptional repressor and lymphoma ...oncoprotein, as a pivotal factor required for neurogenesis and tumor suppression of medulloblastoma (MB). BCL6 is necessary for and capable of preventing the development of GNP-derived MB in mice, and can block the growth of human MB cells in vitro. BCL6 neurogenic and oncosuppressor effects rely on direct transcriptional repression of Gli1 and Gli2 effectors of the SHH pathway, through recruitment of BCOR corepressor and SIRT1 deacetylase. Our findings identify the BCL6/BCOR/SIRT1 complex as a potent repressor of the SHH pathway in normal and oncogenic neural development, with direct diagnostic and/or therapeutic relevance for SHH MB.
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•BCL6 is required for neurogenesis of cerebellar granule neurons•BCL6 directly represses the Sonic Hedgehog effectors Gli1 and Gli2•BCL6/BCOR/SIRT1 complex blocks the growth of human medulloblastoma cells•BCL6 prevents the development of medulloblastoma in the mouse
Tiberi et al. show that BCL6, a lymphoma oncoprotein, is required for neurogenesis and suppression of medulloblastoma. These effects depend on direct transcriptional repression of Gli1 and Gli2, SHH pathway effectors, by BCL6 through recruiting the corepressor BCOR and the deacetylase SIRT1.
The transcription factor Eomesodermin (Eomes) is involved in early embryonic patterning, but the range of cell fates that it controls as well as its mechanisms of action remain unclear. Here we show ...that transient expression of Eomes promotes cardiovascular fate during embryonic stem cell differentiation. Eomes also rapidly induces the expression of Mesp1, a key regulator of cardiovascular differentiation, and directly binds to regulatory sequences of Mesp1. Eomes effects are strikingly modulated by Activin signalling: high levels of Activin inhibit the promotion of cardiac mesoderm by Eomes, while they enhance Eomes‐dependent endodermal specification. These results place Eomes upstream of the Mesp1‐dependent programme of cardiogenesis, and at the intersection of mesodermal and endodermal specification, depending on the levels of Activin/Nodal signalling.
The transcription factor Eomes induces Mesp1, thereby promoting cardiovascular fate during embryonic stem cell differentiation. This effect is modulated by Activin signals, which inhibit Eomes‐dependent cardiac fate and instead enhance endodermal differentiation.
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