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Cited by 1,325Neurogenesis and neuroplasticity mechanismsDevelopmental Biology and Gene RegulationSingle-cell and spatial transcriptomics

Transcriptional landscape of the prenatal human brain

Open access
Miller, Jeremy A., Ding, Song-Lin, Sunkin, Susan M., Smith, Kimberly A., Ng, Lydia, Szafer, Aaron, Ebbert, Amanda, Riley, Zackery L. +72 more·2014·Nature
DOI 10.1038/nature13185PMC PMC4105188
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Abstract

The anatomical and functional architecture of the human brain is mainly determined by prenatal transcriptional processes. We describe an anatomically comprehensive atlas of the mid-gestational human brain, including de novo reference atlases, in situ hybridization, ultra-high-resolution magnetic resonance imaging (MRI) and microarray analysis on highly discrete laser-microdissected brain regions. In developing cerebral cortex, transcriptional differences are found between different proliferative and post-mitotic layers, wherein laminar signatures reflect cellular composition and developmental processes. Cytoarchitectural differences between human and mouse have molecular correlates, including species differences in gene expression in subplate, although surprisingly we find minimal differences between the inner and outer subventricular zones even though the outer zone is expanded in humans. Both germinal and post-mitotic cortical layers exhibit fronto-temporal gradients, with particular enrichment in the frontal lobe. Finally, many neurodevelopmental disorder and human-evolution-related genes show patterned expression, potentially underlying unique features of human cortical formation. These data provide a rich, freely-accessible resource for understanding human brain development.

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Crossref · 10.1038/nature13185

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Claims the paper makes

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  • Findingown data

    Laminar profiling identified many SP-enriched genes in human.

    “In mouse and other species18 this layer is molecularly distinct, and our laminar profiling also identified many SP-enriched genes in human (Fig. 2).”
    Quote in the paperSubstantial claim
  • Findingown data

    At 21pcw, about 2000 layer-enriched genes with R>0.5 were identified in both brains.

    “To identify laminar signatures at 21pcw, we correlated each gene with a binary vector (1 in tested layer versus 0 elsewhere), which identified ~2000 layer-enriched genes with R>0.5 in both brains (Suppl. Table 3).”
    Quote in the paperSubstantial claim
  • Findingown data

    WGCNA on 526 neocortical samples identified 42 modules of co-expressed genes.

    “To identify principal features of the developing cortical transcriptome, we performed weighted gene co-expression network analysis (WGCNA)36 on all 526 neocortical samples, and identified 42 modules of co-expressed genes (Fig. 3a; Suppl. Tables 5–6; Suppl. Methods).”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [36] Zhang, B. & Horvath, S. A general framework for weighted gene co-expression network analysis. Stat. Appl. Genet. Mol. Biol. 4, 17 (2005) source

      Not checked in the free run.

  • Findingown data

    CALB2 is enriched in MZ and SG.

    “For example, the Cajal-Retzius cell marker CALB230 showed enrichment in MZ and SG as expected.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [30] Vogt Weisenhorn, D. M., Prieto, E. W. & Celio, M. R. Localization of calretinin in cells of layer I (Cajal-Retzius cells) of the developing cortex of the rat. Brain Res. Dev. Brain Res. 82, 293–297 (1994) source

      Not checked in the free run.

  • Findingown data

    Most C31 genes were assigned to a module showing common enrichment in both GEs and VZ.

    “Most genes were assigned to a module showing common enrichment in both the GEs and VZ (Extended Data Fig. 6).”
    Quote in the paperSubstantial claim
  • May combine two claims. 44
  • Classified as probably not a substantial claim. 25

  • May combine two claims.

    Findingown data

    Each layer included genes with high laminar specificity, although SZi profiles tended to overlap with neighboring SZo and VZ.

    “Each layer included genes with high laminar specificity (Fig. 2d), although the SZi profiles tended to overlap with the neighboring SZo and VZ.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    At 15-16 pcw, 39 genes were enriched in SZi and only eight were specifically enriched in SZo.

    “Remarkably, few genes met both criteria at 15–16 pcw: 39 genes were enriched in SZi (including several in module G3), while only eight were specifically enriched in SZo.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingcitation

    About 95% of RefSeq genes are expressed in developing neocortex, compared with 84% in adult using these arrays.

    “Approximately 95% of RefSeq genes are expressed in developing neocortex, compared with 84% identified using these arrays in adult23.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [23] Hawrylycz, M. J. et al. An anatomically comprehensive atlas of the adult human brain transcriptome. Nature 489, 391–399 (2012) source

      Not checked in the free run.

  • May combine two claims.

    Findingown data

    CPi is enriched for genes involved in forming connections, whereas CPo is primarily enriched for metabolism rather than mature neuronal function.

    “The next oldest neurons in CPi are additionally enriched for genes involved in forming connections, whereas the youngest neurons in CPo are primarily enriched for terms related to metabolism rather than mature neuronal function.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Different cortical layers show robust and unique molecular signatures, and samples group by layer using hierarchical clustering at both timepoints.

    “Different layers show robust and unique molecular signatures, and samples group by layer using hierarchical clustering (using differential genes from ANOVA, p<10−29) at both timepoints (Fig. 2b).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Module C38 contains genes enriched in germinal layers and decreases with age.

    “Module C38 contains genes enriched in germinal layers, and also decreased expression with age (Fig. 3b, upper left).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingcitation

    TBR2 and PAX6 are enriched in germinal layers as in mouse, but PAX6 is also highly expressed in human SZ whereas in mouse it is restricted to VZ.

    “The cortical progenitor markers TBR2 and PAX6 are enriched in germinal layers as in mouse, although PAX6 in mouse is restricted to VZ whereas it is also highly expressed in human SZ31.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [31] Englund, C. et al. Pax6, Tbr2, and Tbr1 are expressed sequentially by radial glia, intermediate progenitor cells, and postmitotic neurons in developing neocortex. J. Neurosci. 25, 247–251 (2005) source

      Not checked in the free run.

  • May combine two claims.

    Findingown data

    NPY is enriched in SP at 21pcw but not at 15-16pcw, as shown by microarray and ISH.

    “For example, NPY is enriched in SP at 21pcw (but not 15–16pcw) as shown both by microarray and ISH (Fig. 4a).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    Module C31 is enriched in SG and VZ and contains many interneuron-associated genes.

    “Finally, we identified a module (C31) with particular enrichment in SG and VZ (Fig. 3b, upper right), containing many interneuron-associated genes (Fig. 3c).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    A consensus co-expression network using VZ, SZi, and SZo samples from 15/16 pcw brains identified eight co-expression modules with selective enrichment in either SZ or VZ, highly conserved between brains and highly distinct.

    “We found eight co-expression modules with selective enrichment in either the SZ or the VZ (Fig. 3d,e), highly conserved between brains (p<10−50), and highly distinct.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    MDS separates samples by layer, with germinal zones distinct from postmitotic layers, and dimension two roughly reflects rostrocaudal position.

    “Samples also cluster with multidimensional scaling (MDS), where dimension one separates samples by layer, with germinal zones (VZ, SZi, SZo) distinct from layers containing primarily postmitotic cells, and dimension two roughly reflects rostrocaudal position (Fig. 2c).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Most gene modules corresponded to layers and/or changes with age, while areal patterning appeared to be a more subtle transcriptional feature.

    “Here, most gene clusters (“modules”) corresponded to layers and/or changes with age, (Fig. 3a-b; Extended Data Fig. 4) while areal patterning appeared to be a more subtle transcriptional feature.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Findingcitation

    These results are consistent with a previous study of laminar enrichment in 13-16pcw human prenatal cortex, which also found few genes specifically enriched in SZo or SZi.

    “These results are consistent with a previous study of laminar enrichment in 13–16pcw human prenatal cortex, which also found few genes specifically enriched in SZo (55) or SZi (61)2.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [2] Fietz, S. A. et al. Transcriptomes of germinal zones of human and mouse fetal neocortex suggest a role of extracellular matrix in progenitor self-renewal. Proc. Natl Acad. Sci. USA 109, 11836–11841 (2012) source

      Not checked in the free run.

  • May combine two claims.

    Findingown data

    In human, ZIC1 and CALB2 expression do not overlap in MZ, unlike mouse where Zic1 is expressed by virtually all Cajal-Retzius neurons.

    “However, while mouse Zic1 is expressed by virtually all Cajal-Retzius neurons34, our results indicate that this is not true in human as ZIC1 and CALB2 expression do not overlap in MZ (Figs. 1b and 2e).”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [34] Inoue, T., Ogawa, M., Mikoshiba, K. & Aruga, J. Zic deficiency in the cortical marginal zone and meninges results in cortical lamination defects resembling those in type II lissencephaly. J. Neurosci. 28, 4712–4725 (2008) source

      Not checked in the free run.

  • May combine two claims.

    Findingown data

    Module C22 is enriched in newly generated postmitotic neurons of the CP and increases with age.

    “Conversely, module C22 is enriched in newly generated postmitotic neurons of the CP, and increases with age (Fig. 3b, lower left).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingcitation

    Modules G2 and G3 are enriched in SZi, and G3 is enriched for genes marking Svet1+ IPs in E14 mouse SZ.

    “Modules G2 and G3 are enriched in the SZi (Fig. 3d, white bars in bottom two rows), and G3 is enriched for genes marking Svet1+ IPs in E14 mouse SZ43, including ELAVL4, LRP8, NEUROD1, NRN1, SLC17A6, SSTR2, and TP53INP1.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [43] Kawaguchi, A. et al. Single-cell gene profiling defines differential progenitor subclasses in mammalian neurogenesis. Development 135, 3113–3124 (2008) source

      Not checked in the free run.

  • May combine two claims.

    Findingown data

    Module C16 is enriched in SP and shows hallmarks of mature neuronal function.

    “For example, module C16 is enriched in SP (Fig. 3b, lower right), and shows hallmarks of mature neuronal function.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    Trh and Nxph4 show enriched expression in mouse but not human SP.

    “Conversely, Trh and Nxph4 show enriched expression in mouse but not human SP (Fig. 4d; Extended Data Fig. 8b).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    FOXP2 is included in module C16 and shows enrichment in parietal and temporal lobes including presumptive language areas.

    “Interestingly, FOXP2, implicated in the specialization of language areas54, is included in the module C16, and shows enrichment in parietal and temporal lobes including presumptive language areas (Extended Data Fig. 10).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [54] Enard, W. et al. Molecular evolution of FOXP2, a gene involved in speech and language. Nature 418, 869–872 (2002) source

      Not checked in the free run.

  • May combine two claims.

    Findingown data

    In developing cerebral cortex, transcriptional differences exist between proliferative and postmitotic layers, and laminar signatures reflect cellular composition and developmental processes.

    “In developing cerebral cortex, transcriptional differences are found between different proliferative and postmitotic layers, wherein laminar signatures reflect cellular composition and developmental processes.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Both germinal and postmitotic cortical layers exhibit fronto-temporal gradients, with particular enrichment in frontal lobe.

    “Both germinal and postmitotic cortical layers exhibit fronto-temporal gradients, with particular enrichment in frontal lobe.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation

    SP generation is protracted in primates, and its thickness is particularly expanded in human.

    “SP generation is protracted in primates8, and its thickness particularly expanded in human8.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [8] Smart, I., Dehay, C., Giroud, P., Berland, M. & Kennedy, H. Unique morphological features of the proliferative zones and postmitotic compartments of the neural epithelium giving rise to striate and extrastriate cortex in the monkey. Cereb.Cortex 12, 37–53 (2002) source

      Not checked in the free run.

  • May combine two claims.

    Interpretationown data

    Both VZ-enriched modules G7 and G8 contained cell cycle genes and many astrocyte markers, suggesting they may represent different RG populations.

    “Both VZ-enriched modules (G7, G8) contained cell cycle genes and many astrocyte markers, suggesting that these modules may represent different RG populations.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    DLX1 and DLX2 are essential for interneuron migration and survival.

    “DLX1 and DLX2, homeobox transcription factors essential for interneuron migration and survival10, were central (“hub”) genes therein.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [10] Anderson, S. A., Eisenstat, D. D., Shi, L. & Rubenstein, J. L. Interneuron migration from basal forebrain to neocortex: dependence on Dlx genes. Science 278, 474–476 (1997) source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    Several recent studies provide strong evidence that primate interneurons are generated extracortically in the ganglionic eminences, as with rodents.

    “However, several recent studies have shown strong evidence that, as with rodents, primate interneurons are generated extracortically in the GEs11,14.”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [11] Ma, T. et al. Subcortical origins of human and monkey neocortical interneurons. Nature Neurosci. 16, 1588–1597 (2013) source

      Not checked in the free run.

    • cites [14] Hansen, D. V. et al. Non-epithelial stem cells and cortical interneuron production in the human ganglionic eminences. Nature Neurosci. 16, 1576–1587 (2013) source

      Not checked in the free run.

  • May combine two claims.

    Interpretationcitation

    SZo-enriched categories primarily relate to cell division and contain many astrocytic markers likely expressed in outer radial glia.

    “SZo-enriched categories primarily related to cell division and contained many astrocytic markers likely expressed in outer radial glia (ORG)7.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [7] Hansen, D. V., Lui, J., Parker, P. & Kriegstein, A. Neurogenic radial glia in the outer subventricular zone of human neocortex. Nature 464, 554–561 (2010) source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    In mouse and other species, SP is molecularly distinct.

    “In mouse and other species18 this layer is molecularly distinct, and our laminar profiling also identified many SP-enriched genes in human (Fig. 2).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [18] Wang, W. Z. et al. Comparative aspects of subplate zone studied with gene expression in sauropsids and mammals. Cereb. Cortex 21, 2187–2203 (2011) source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation

    Germinal layers contain various cortical progenitors including RG in VZ, IP in SZi, and ORG in SZo, and these RG may be diverse.

    “Germinal layers contain various cortical progenitors including RG in the VZ, intermediate progenitors (IP) in the SZi, and ORG in the SZo (reviewed in41), and these RG may be quite diverse42.”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [41] Gaspard, N. & Vanderhaeghen, P. Laminar fate specification in the cerebral cortex. F1000 Biol. Rep. 3, 6 (2011) source

      Not checked in the free run.

    • cites [42] Betizeau, M. et al. Precursor diversity and complexity of lineage relationships in the outer subventricular zone of the primate. Neuron 80, 442–457 (2013) source

      Not checked in the free run.

  • May combine two claims.

    Interpretationown data

    Modules G4-G6 are enriched in SZo but are enriched for neuron-associated categories likely expressed in postmitotic cells during radial migration.

    “Modules G4-G6 are enriched in the SZo (Fig. 3d, gray bars); however, these modules are enriched for neuron-associated categories likely expressed in postmitotic cells during their radial migration.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data

    ZIC1 expression is restricted to the pia mater overlying cortex, indicating SG samples captured pial cells in addition to granule cells.

    “Finally, expression of ZIC1, associated with Dandy-Walker congenital brain malformation33, was restricted to the pia mater overlying the cortex, therefore indicating that SG samples captured pial cells in addition to granule cells.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [33] Grinberg, I. et al. Heterozygous deletion of the linked genes ZIC1 and ZIC4 is involved in Dandy-Walker malformation. Nature Genet. 36, 1053–1055 (2004) source

      Not checked in the free run.

  • May combine two claims.

    Interpretationreasoning

    Module C38 has a large signature of glia and cell division, suggesting these genes reflect decreasing progenitor cell division.

    “This module has a large signature of glia and cell division, suggesting that these genes reflect decreasing progenitor cell division.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation

    SP is a largely transient zone beneath CP that plays an important role in establishment of thalamocortical connectivity.

    “The SP is a largely transient zone beneath the CP that plays an important role in establishment of thalamocortical connectivity (see35 for review).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [35] Kostović, I. & Judas, M. The development of the subplate and thalamocortical connections in the human foetal brain. Acta Paediatr. 99, 1119–1127 (2010) source

      Not checked in the free run.

  • May combine two claims.

    Interpretationown data

    This human frontal asymmetry is most apparent in outer postmitotic layers MZ and CPo, and in SZo, suggesting these genes may play a part in expansion and reorganization of human PFC.

    “This human frontal asymmetry is most apparent in the outer postmitotic layers MZ and CPo (Fig. 5i), and in SZo, which generates most superficial CP neurons in primate7,8, suggesting that these genes may play a part in the expansion and reorganization of human PFC8.”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites [7] Hansen, D. V., Lui, J., Parker, P. & Kriegstein, A. Neurogenic radial glia in the outer subventricular zone of human neocortex. Nature 464, 554–561 (2010) source

      Not checked in the free run.

    • cites [8] Smart, I., Dehay, C., Giroud, P., Berland, M. & Kennedy, H. Unique morphological features of the proliferative zones and postmitotic compartments of the neural epithelium giving rise to striate and extrastriate cortex in the monkey. Cereb.Cortex 12, 37–53 (2002) source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    More detailed analysis of layers of fetal mouse neocortex found more than 2500 genes differentially expressed between VZ, SZ, IZ, and CP.

    “Expression varies between cell populations, and more detailed analysis of layers of fetal mouse neocortex found >2500 genes differentially expressed between ventricular zone (VZ), SZ, intermediate zone (IZ), and cortical plate (CP)16.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [16] Ayoub, A. E. et al. Transcriptional programs in transient embryonic zones of the cerebral cortex defined by high-resolution mRNA sequencing. Proc. Natl Acad. Sci. USA 108, 14950–14955 (2011) source

      Not checked in the free run.

  • Classified as probably not a substantial claim.

    Premisecitation

    In mouse, nearly all cortical GABAergic interneurons originate from the striatal ganglionic eminences of the ventral telencephalon.

    “In mouse, nearly all originate from the striatal ganglionic eminences (GEs) of the ventral telencephalon10; however, the origin of human cortical interneurons remains controversial7,11–14.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [10] Anderson, S. A., Eisenstat, D. D., Shi, L. & Rubenstein, J. L. Interneuron migration from basal forebrain to neocortex: dependence on Dlx genes. Science 278, 474–476 (1997) source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation

    CNSs are typically located by genes important for developmental regulation, and many show significant enhancer activity in brain.

    “CNSs are typically located by genes important for developmental regulation, and many show significant enhancer activity in brain5.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [5] Lambert, N. et al. Genes expressed in specific areas of the human fetal cerebral cortex display distinct patterns of evolution. PLoS ONE 6, e17753 (2011) source

      Not checked in the free run.

  • May combine two claims.

    Interpretationcitation

    Trh is also not expressed in rat SP, suggesting this pattern is specific to mouse.

    “Interestingly, Trh is also not expressed in the rat SP18, suggesting this pattern is specific to mouse.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [18] Wang, W. Z. et al. Comparative aspects of subplate zone studied with gene expression in sauropsids and mammals. Cereb. Cortex 21, 2187–2203 (2011) source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    Species differences in distinct fetal transient zones, including SZ, SP, CP, and SG, have been described.

    “Species differences in distinct fetal transient zones, including the SZ2, SP17,18, CP6, and SG9, have also been described.”
    Quote in the paperNot a substantial claim0 of 3 sources checked
    • cites [2] Fietz, S. A. et al. Transcriptomes of germinal zones of human and mouse fetal neocortex suggest a role of extracellular matrix in progenitor self-renewal. Proc. Natl Acad. Sci. USA 109, 11836–11841 (2012) source

      Not checked in the free run.

    • cites [17] Montiel, J. et al. Hypothesis on the dual origin of the mammalian subplate. Front. Neuroanat. 5, 25 (2011) source

      Not checked in the free run.

    • cites [18] Wang, W. Z. et al. Comparative aspects of subplate zone studied with gene expression in sauropsids and mammals. Cereb. Cortex 21, 2187–2203 (2011) source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    Profiling of mid-gestational human brain identified many genes differentially expressed between major regions, including genes associated with human-accelerated conserved noncoding sequences.

    “Profiling of mid-gestational human brain3,5 identified many genes differentially expressed between major regions, including genes associated with human-accelerated conserved noncoding sequences (haCNSs)15.”
    Quote in the paperNot a substantial claim0 of 3 sources checked
    • cites [3] Johnson, M. B. et al. Functional and evolutionary insights into human brain development through global transcriptome analysis. Neuron 62, 494–509 (2009) source

      Not checked in the free run.

    • cites [5] Lambert, N. et al. Genes expressed in specific areas of the human fetal cerebral cortex display distinct patterns of evolution. PLoS ONE 6, e17753 (2011) source

      Not checked in the free run.

    • cites [15] Prabhakar, S., Noonan, J., Pääbo, S. & Rubin, E. Accelerated evolution of conserved noncoding sequences in humans. Science 314, 786 (2006) source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    The subpial granular zone is greatly expanded in human and is a transient compartment at the pial surface composed primarily of tangentially migrating neurons.

    “The transient subplate zone (SP) is greatly expanded in human8, as is the subpial granular zone (SG), a transient compartment at the pial surface composed primarily of tangentially migrating neurons9.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [9] Brun, A. The subpial granular layer of the foetal cerebral cortex in man. Its ontogeny and significance in congenital cortical malformations. Acta Pathologica et Microbiologica Scandinavica 179 (suppl.). 3–98 (1965)

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Genes near haCNSs are particularly likely to show differential expression between regions of developing human neocortex, suggesting transcriptional regulation by haCNSs may be important in human-specific neurodevelopment.

    “Genes near CNSs with significantly accelerated rates of substitution in the human lineage (haCNSs)15 are particularly likely to show differential expression between regions of developing human neocortex3,5, suggesting transcriptional regulation by haCNSs may be important in human-specific neurodevelopment.”
    Quote in the paperSubstantial claim0 of 3 sources checked
    • cites [15] Prabhakar, S., Noonan, J., Pääbo, S. & Rubin, E. Accelerated evolution of conserved noncoding sequences in humans. Science 314, 786 (2006) source

      Not checked in the free run.

    • cites [3] Johnson, M. B. et al. Functional and evolutionary insights into human brain development through global transcriptome analysis. Neuron 62, 494–509 (2009) source

      Not checked in the free run.

    • cites [5] Lambert, N. et al. Genes expressed in specific areas of the human fetal cerebral cortex display distinct patterns of evolution. PLoS ONE 6, e17753 (2011) source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation

    A secondary progenitor zone, the subventricular zone, is present in all mammals but is split into an outer and inner region in primates.

    “A secondary progenitor zone, the subventricular zone (SZ) is present in all mammals, but is split into an outer and inner region in primates7.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [7] Hansen, D. V., Lui, J., Parker, P. & Kriegstein, A. Neurogenic radial glia in the outer subventricular zone of human neocortex. Nature 464, 554–561 (2010) source

      Not checked in the free run.

  • May combine two claims.

    Interpretationown data

    CBLN2 shows rostral enrichment in human CPo and in mouse, with an abrupt expression cutoff, implicating CBLN2 in early rostral patterning.

    “Conversely, CBLN2 shows rostral enrichment in human CPo (Fig. 5f) and in mouse (Fig. 5g), with an abrupt expression cutoff, implicating CBLN2 in early rostral patterning.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data

    This result cannot be explained by over-representation of neural adhesion genes in IZ, suggesting IZ is of particular importance in areal cortical identity during human development.

    “This result, which cannot be explained by over-representation of neural adhesion genes in IZ, suggests that IZ is of particular importance in areal cortical identity during human development53.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [53] Allendoerfer, K. L. & Shatz, C. J. The subplate, a transient neocortical structure: its role in the development of connections between thalamus and cortex. Annu. Rev. Neurosci. 17, 185–218 (1994) source

      Not checked in the free run.

  • May combine two claims.

    Interpretationreasoning

    Gradient genes in VZ likely reflect intrinsic areal specification because VZ does not receive thalamic innervation.

    “Gradient genes in VZ likely reflect intrinsic areal specification as VZ does not receive thalamic innervation.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Interpretationown data

    Cytoarchitectural differences between human and mouse have molecular correlates, including species differences in gene expression in subplate.

    “Cytoarchitectural differences between human and mouse have molecular correlates, including species differences in gene expression in subplate, although surprisingly we find minimal differences between the inner and human-expanded outer subventricular zones.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    FGFR3 mutations cause defects in human temporal cortex.

    “For example, FGFR3, which is known to cause defects in human temporal cortex when mutated50, shows significant caudal enrichment in all germinal layers in both species (Fig. 5d–e).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [50] Hevner, R. F. The cerebral cortex malformation in thanatophoric dysplasia: neuropathology and pathogenesis. Acta Neuropathol. 110, 208–221 (2005) source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation

    HCRTR2 is thought to regulate sleep-wakefulness and is highly expressed in mouse hypothalamus.

    “Several genes showed enrichment in developing human but not mouse SP, including the hypocretin (orexin) receptor HCRTR2 (Fig. 4c; Extended Data Fig. 8a), which is thought to regulate sleep-wakefulness and is highly expressed in mouse hypothalamus45.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [45] Gerashchenko, D. et al. Hypocretin-2-saporin lesions of the lateral hypothalamus produce narcoleptic-like sleep behavior in the rat. J. Neurosci. 21, 7273–7283 (2001) source

      Not checked in the free run.

  • May combine two claims.

    Interpretationown data

    More rostrally- than caudally-enriched genes were found in nearly every cortical layer, whereas two studies of prenatal mouse brain identified more or comparable caudally-enriched genes, indicating potential species differences related to areal patterning.

    “Interestingly, we find more rostrally- than caudally-enriched genes in nearly every cortical layer (Fig. 5h), whereas two studies of gradient expression in prenatal mouse brain identified more49 or comparable51 caudally-enriched genes, indicating potential species differences related to areal patterning.”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites [49] Sansom, S. N. & Livesey, F. Gradients in the brain: the control of the development of form and function in the cerebral cortex. Cold Spring Harb. Perspect. Biol. 1, a002519 (2009) source

      Not checked in the free run.

    • cites [51] Pinto, L. et al. AP2γ regulates basal progenitor fate in a region- and layer-specific manner in the developing cortex. Nature Neurosci. 12, 1229–1237 (2009) source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Cortical patterning is likely a result of intrinsic signaling, controlled in part by graded expression of transcription factors during early cortical development, followed by extrinsic signaling from thalamic afferents after the start of corticogenesis.

    “Cortical patterning is likely a result of intrinsic signaling, controlled in part by graded expression of transcription factors during early cortical development, followed by extrinsic signaling from thalamic afferents after the start of corticogenesis46–49.”
    Quote in the paperNot a substantial claim0 of 4 sources checked
    • cites [46] Mallamaci, A. & Stoykova, A. Gene networks controlling early cerebral cortex arealization. Eur. J. Neurosci. 23, 847–856 (2006) source

      Not checked in the free run.

    • cites [47] O’Leary, D. D. Do cortical areas emerge from a protocortex? Trends Neurosci. 12, 400–406 (1989) source

      Not checked in the free run.

    • cites [48] Rakic, P. Specification of cerebral cortical areas. Science 241, 170–176 (1988) source

      Not checked in the free run.

    • cites [49] Sansom, S. N. & Livesey, F. Gradients in the brain: the control of the development of form and function in the cerebral cortex. Cold Spring Harb. Perspect. Biol. 1, a002519 (2009) source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    FOXP2 is implicated in the specialization of language areas.

    “Interestingly, FOXP2, implicated in the specialization of language areas54, is included in the module C16, and shows enrichment in parietal and temporal lobes including presumptive language areas (Extended Data Fig. 10).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [54] Enard, W. et al. Molecular evolution of FOXP2, a gene involved in speech and language. Nature 418, 869–872 (2002) source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    The human neocortex has undergone massive evolutionary expansion, particularly in superficial layers, likely due to differences in rates of progenitor pool expansion during neurogenesis compared to other species.

    “For example, the human neocortex has undergone massive evolutionary expansion, particularly in superficial layers, likely due to differences in rates of progenitor pool expansion during neurogenesis compared to other species6.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [6] Kriegstein, A., Noctor, S. & Martínez-Cerdeño, V. Patterns of neural stem and progenitor cell division may underlie evolutionary cortical expansion. Nature Rev. Neurosci. 7, 883–890 (2006) source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Interpretationcitation

    The asymmetry could alternatively reflect temporal differences, as peak generation of excitatory neurons in visual cortex is delayed relative to frontal cortex.

    “Alternatively, this asymmetry could reflect temporal differences, as peak generation of excitatory neurons in visual cortex is delayed relative to frontal cortex52.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [52] Rakic, P. Neurogenesis in adult primates. Prog. Brain Res. 138, 3–14 (2002) source

      Not checked in the free run.

  • May combine two claims.

    Interpretationown data

    The present atlas offers transcriptome data from prenatal human specimens that are usually hard to obtain for research, extending earlier molecular atlases of mouse and adult human brain.

    “The atlas of the mid-gestational human brain described here, part of the BrainSpan Atlas of the Developing Human Brain, builds on digital molecular brain atlasing efforts in mouse20,22,24 and adult human23 by providing transcriptome resources on prenatal specimens typically inaccessible for research.”
    Quote in the paperSubstantial claim0 of 4 sources checked
    • cites [20] Paxinos, G. & Franklin, K. Paxinos and Franklin’s the Mouse Brain in Stereotaxic Coordinates 4th edn (Academic Press, 2012)

      Not checked in the free run.

    • cites [22] Dong, H. W. Allen Reference Atlas: a Digital Color Brain Atlas of the C57BL/6J Male Mouse (Wiley, 2008)

      Not checked in the free run.

    • cites [24] Lein, E. S. et al. Genome-wide atlas of gene expression in the adult mouse brain. Nature 445, 168–176 (2007) source

      Not checked in the free run.

    • cites [23] Hawrylycz, M. J. et al. An anatomically comprehensive atlas of the adult human brain transcriptome. Nature 489, 391–399 (2012) source

      Not checked in the free run.

    • Rests on an assumption: Prenatal human brain specimens are typically inaccessible for research.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductnone given

    To address batch effects, a common RNA pool control was amplified and hybridized in each batch.

    “To control for batch effects, common RNA pool control samples were amplified and hybridized in each batch.”
    Quote in the paperNot a substantial claim

  • Classified as probably not a substantial claim.

    Study design and conductnone given

    Except where noted, all p-values were Bonferroni corrected for multiple comparisons.

    “Unless otherwise noted, all p-values are Bonferroni corrected for multiple comparisons.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductown data

    Outliers of biological origin were found by comparing samples from related structures with hierarchical clustering and inter-array correlation measures.

    “Biological outliers were identified by comparing samples from related structures using hierarchical clustering and inter-array correlation measures.”
    Quote in the paperNot a substantial claim

  • Classified as probably not a substantial claim.

    Study design and conductreasoning

    The consensus co-expression network should identify only relationships common to both brains and exclude specimen-specific features or age-related changes.

    “This network will only identify intergenic relationships common to both brains, and should therefore produce robust co-expression relationships that exclude specimen-specific features or changes with age.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Study design and conductown data

    In total, 1,225 samples met sample quality control, with 1,202 experimental and 23 control samples.

    “A total of 1,225 samples passed sample quality control (QC), including 1,202 experimental samples and 23 control samples.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    TGIF1 and SIX3 are associated with abnormal cortical development in holoprosencephaly.

    “Similarly, two genes associated with abnormal cortical development in holoprosencephaly, TGIF1 and SIX327, are also enriched in cortical germinal zones.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [27] Dubourg, C. et al. Molecular screening of SHH, ZIC2, SIX3, and TGIF genes in patients with features of holoprosencephaly spectrum: mutation review and genotype–phenotype correlations. Hum. Mutat. 24, 43–51 (2004) source

      Not checked in the free run.

  • Classified as probably not a substantial claim. May combine two claims.

    Study design and conductnone given

    All microarray data underwent quality control and ERCC spike-in assessments, and samples that failed were excluded from analysis.

    “All microarray data was subjected to QC and ERCC spike-in assessments, and any failing samples were omitted from the analysis.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductown data

    Across the 1,202 experimental samples that passed quality control, the mean RNA Integrity Number was 6.3.

    “The average RNA Integrity Number (RIN) of all 1,202 passed experimental samples was 6.3.”
    Quote in the paperNot a substantial claim
62 of 62

  • The paper relies on this without support.

    The anatomical and functional architecture of the human brain is largely determined by prenatal transcriptional processes.

    “The anatomical and functional architecture of the human brain is largely determined by prenatal transcriptional processes.”

  • The paper relies on this without support.

    Understanding the emergence of cortical specialization for language can only be studied in humans.

    “Finally, understanding the emergence of cortical specialization for language can only be studied in humans.”

  • The paper relies on this without support.

    Prenatal human brain specimens are typically inaccessible for research.

    “providing transcriptome resources on prenatal specimens typically inaccessible for research”
3 of 3

Claims other papers make about this one

Each is checked against this paper’s own text: does it say what the citing paper says it does?

  • ApoE expression, in both mice and humans, begins relatively late in development.

    “ApoE expression, in both mice and humans, begins relatively late in development [46, 47].”

    From ApoE, ApoE Receptors, and the Synapse in Alzheimer's Disease (2017) · cites this paper as [46]

  • The 53 genes are overexpressed in 'adolescence' and 'young adult' brain tissues in BrainSpan, consistent with accumulating evidence suggesting Alzheimer's disease may start decades before the onset of disease.

    “the 53 genes are overexpressed in ‘adolescence’ and ‘young adult’ brain tissues in BrainSpan103, a transcriptomic atlas of the developing human brain, which is consistent with accumulating evidence suggesting Alzheimer’s disease may start decades before the onset of disease104,105”

    From Genetic meta-analysis of diagnosed Alzheimer’s disease identifies new risk loci and implicates Aβ, tau, immunity and lipid processing (2019) · cites this paper as [103]

  • The Allen Brain Institute and BrainSpan have evaluated spatial and temporal transcriptomes.

    “Recent important advances include the evaluation of spatial and temporal transcriptomes by the Allen Brain Institute and BrainSpan8,11–13”

    From Systems biology and gene networks in neurodevelopmental and neurodegenerative disorders (2015) · cites this paper as [11]

  • Network analysis can identify multiple levels of molecular organization within the hierarchy of brain region, cell type, organelle and molecular pathways using only transcriptional data, and can enable integration with other information.

    “A major advantage of network analysis over DGE analysis is that it can identify multiple levels of molecular organization within the hierarchy of brain region, cell type, organelle and molecular pathways using only transcriptional data, and can thus enable integration with other information, such as known pathway annotations, protein interactions and other molecular profiling data11,12,26,27”

    From Systems biology and gene networks in neurodevelopmental and neurodegenerative disorders (2015) · cites this paper as [11]

  • When evaluating co-expression across brain regions, hubs in modules associated with specific regions, such as the cerebellum, are usually markers for predominant cell types, such as granule cells.

    “For example, when evaluating co-expression across brain regions, hubs in modules that are associated with specific regions, such as the cerebellum, are usually markers for predominant cell types, such as granule cells11,12,26,32.”

    From Systems biology and gene networks in neurodevelopmental and neurodegenerative disorders (2015) · cites this paper as [11]

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Authors
Miller, J. A., Ding, S.-L., Sunkin, S. M., Smith, K. A., Ng, L., Szafer, A., Ebbert, A., Riley, Z. L., Royall, J. J., Aiona, K., Arnold, J. M., Bennet, C., Bertagnolli, D., Brouner, K., Butler, S., Caldejon, S., Carey, A., Cuhaciyan, C., Dalley, R. A., Dee, N., Dolbeare, T. A., Facer, B. A. C., Feng, D., Fliss, T. P., Gee, G., Goldy, J., Gourley, L., Gregor, B. W., Gu, G., Howard, R. E., Jochim, J. M., Kuan, C. L., Lau, C., Lee, C.-K., Lee, F., Lemon, T. A., Lesnar, P., McMurray, B., Mastan, N., Mosqueda, N., Naluai-Cecchini, T., Ngo, N.-K., Nyhus, J., Oldre, A., Olson, E., Parente, J., Parker, P. D., Parry, S. E., Stevens, A., Pletikos, M., Reding, M., Roll, K., Sandman, D., Sarreal, M., Shapouri, S., Shapovalova, N. V., Shen, E. H., Sjoquist, N., Slaughterbeck, C. R., Smith, M., Sodt, A. J., Williams, D., Zöllei, L., Fischl, B., Gerstein, M. B., Geschwind, D. H., Glass, I. A., Hawrylycz, M. J., Hevner, R. F., Huang, H., Jones, A. R., Knowles, J. A., Levitt, P., Phillips, J. W., Šestan, N., Wohnoutka, P., Dang, C., Bernard, A., Hohmann, J. G., & Lein, E. S.
Affiliations
  • Allen Institute for Brain Science (US)
  • University of Washington (US)
  • Harvard University (US)
  • Yale University (US)
  • University of California, Los Angeles (US)
  • The University of Texas Southwestern Medical Center (US)
  • University of Southern California (US)
Corresponding author
Lein, Ed S.
ORCID
Venue
Nature
Publisher
Springer Science and Business Media LLC
Published
2014-04
Volume
508
Issue
7495
Pages
199-206
ISSN
  • 0028-0836 (ISSN-L)
  • 1476-4687
DOI
10.1038/nature13185
PMC
PMC4105188
OpenAlex
W2163302580
Semantic Scholar
b2924d6a2e0bbf3e450c2e19b3154a39eb19e0c2
Cited by
1,325
References
72
Citations by year
2026: 90 · 2025: 69 · 2024: 100 · 2023: 109 · 2022: 107 · 2021: 154 · 2020: 178 · 2019: 110 · 2018: 104 · 2017: 104 · 2016: 79 · 2015: 83 · 2014: 35 · 2013: 1
FWCI
34.86
Type
journal article
Language
en
Open access
Licence
www.springer.com/tdm (tdm)
Locations
  • Nature (publishedVersion)
  • PubMed (publishedVersion)
  • Europe PMC (PubMed Central) (acceptedVersion)
  • PubMed Central (acceptedVersion)
  • Digital Access to Scholarship at Harvard (DASH) (Harvard University) (submittedVersion, cc-by)
  • RePEc: Research Papers in Economics (submittedVersion)
  • eScholarship (California Digital Library) (submittedVersion)
  • Cambridge University Engineering Department Publications Database (submittedVersion)
Funders
  • National Institutes of Health
  • National Institute of Mental Health (R37MH060233, RC2MH089921, R01MH092535)
  • Eunice Kennedy Shriver National Institute of Child Health and Human Development (Z01HD008836, R00HD061485, 5R24HD0008836, R24HD000836)
Topics
  • Life Sciences › Neuroscience › Developmental Neuroscience › Neurogenesis and neuroplasticity mechanisms
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Developmental Biology and Gene Regulation
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Single-cell and spatial transcriptomics
Keywords
human brain development, transcriptional atlas, subplate, subventricular zone, cortical layers, neurodevelopmental disorders, human brain evolution, Allen Mouse Brain Connectivity Atlas, brain connectivity, laser capture microdissection, in situ hybridization
MeSH terms
Anatomy, Artistic, Animals, Atlases as Topic, Brain, Fetus, Humans, Species Specificity, Conserved Sequence, Gene Expression Regulation, Developmental, Neocortex, Mice, Gene Regulatory Networks, Transcriptome

About these checks

Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.

Claims read by deepseek-v4.1-flash under definition version 3 (extract-v3) on 2026-10-06.

Classified by jev-1.13-20260917 (gold-guideline-v0.2) on 2026-10-06.

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