Neural Progenitors Adopt Specific Identities by Directly Repressing All Alternative Progenitor Transcriptional Programs

Summary In the vertebrate neural tube, a morphogen-induced transcriptional network produces multiple molecularly distinct progenitor domains, each generating different neuronal subtypes. Using an in vitro differentiation system, we defined gene expression signatures of distinct progenitor populations and identified direct gene-regulatory inputs corresponding to locations of specific transcription factor binding. Combined with targeted perturbations of the network, this revealed a mechanism in which a progenitor identity is installed by active repression of the entire transcriptional programs of other neural progenitor fates. In the ventral neural tube, sonic hedgehog (Shh) signaling, together with broadly expressed transcriptional activators, concurrently activates the gene expression programs of several domains. The specific outcome is selected by repressive input provided by Shh-induced transcription factors that act as the key nodes in the network, enabling progenitors to adopt a single definitive identity from several initially permitted options. Together, the data suggest design principles relevant to many developing tissues.

[1]  R. Sandberg,et al.  SoxB1-driven transcriptional network underlies neural-specific interpretation of morphogen signals , 2013, Proceedings of the National Academy of Sciences.

[2]  A. Mansour,et al.  Nato3 plays an integral role in dorsoventral patterning of the spinal cord by segregating floor plate/p3 fates via Nkx2.2 suppression and Foxa2 maintenance , 2014, Development.

[3]  H. Wichterle,et al.  A Requirement for Retinoic Acid-Mediated Transcriptional Activation in Ventral Neural Patterning and Motor Neuron Specification , 2003, Neuron.

[4]  Andrew P. McMahon,et al.  Genomic characterization of Gli-activator targets in sonic hedgehog-mediated neural patterning , 2007, Development.

[5]  Wolfgang Huber,et al.  Enhancer loops appear stable during development and are associated with paused polymerase , 2014, Nature.

[6]  Lior Pachter,et al.  Sequence Analysis , 2020, Definitions.

[7]  Austin G Smith,et al.  Conversion of embryonic stem cells into neuroectodermal precursors in adherent monoculture , 2003, Nature Biotechnology.

[8]  James Briscoe,et al.  Distinct Sonic Hedgehog signaling dynamics specify floor plate and ventral neuronal progenitors in the vertebrate neural tube. , 2010, Genes & development.

[9]  S. Pfaff,et al.  Transcriptional networks regulating neuronal identity in the developing spinal cord , 2001, Nature Neuroscience.

[10]  Ryoichiro Kageyama,et al.  Oscillatory Control of Factors Determining Multipotency and Fate in Mouse Neural Progenitors , 2013, Science.

[11]  James Briscoe,et al.  A theoretical framework for the regulation of Shh morphogen-controlled gene expression , 2014, Development.

[12]  Amy Tsurumi,et al.  STAT Is an Essential Activator of the Zygotic Genome in the Early Drosophila Embryo , 2011, PLoS genetics.

[13]  P. Wittkopp,et al.  Cis-regulatory elements: molecular mechanisms and evolutionary processes underlying divergence , 2011, Nature Reviews Genetics.

[14]  E. Davidson.  Emerging properties of animal gene regulatory networks , 2010, Nature.

[15]  R. Sandberg,et al.  Sequentially acting Sox transcription factors in neural lineage development. , 2011, Genes & development.

[16]  Thomas M. Jessell,et al.  Groucho-Mediated Transcriptional Repression Establishes Progenitor Cell Pattern and Neuronal Fate in the Ventral Neural Tube , 2001, Cell.

[17]  David J. Anderson,et al.  The bHLH Transcription Factor Olig2 Promotes Oligodendrocyte Differentiation in Collaboration with Nkx2.2 , 2001, Neuron.

[18]  Manu,et al.  Characterization of the Drosophila segment determination morphome. , 2008, Developmental biology.

[19]  David A. Orlando,et al.  Master Transcription Factors Determine Cell-Type-Specific Responses to TGF-β Signaling , 2011, Cell.

[20]  A. McMahon,et al.  A direct requirement for Hedgehog signaling for normal specification of all ventral progenitor domains in the presumptive mammalian spinal cord. , 2002, Genes & development.

[21]  H. Arnold,et al.  A homeodomain feedback circuit underlies step-function interpretation of a Shh morphogen gradient during ventral neural patterning , 2010, Development.

[22]  Helen M. Byrne,et al.  Boolean Modelling Reveals New Regulatory Connections between Transcription Factors Orchestrating the Development of the Ventral Spinal Cord , 2014, PloS one.

[23]  Danyang Yu,et al.  Impacts of the ubiquitous factor Zelda on Bicoid-dependent DNA binding and transcription in Drosophila , 2014, Genes & development.

[24]  W. Sung,et al.  Chromatin connectivity maps reveal dynamic promoter–enhancer long-range associations , 2013, Nature.

[25]  David J. Anderson,et al.  The bHLH Transcription Factors OLIG2 and OLIG1 Couple Neuronal and Glial Subtype Specification , 2002, Cell.

[26]  N. Friedman,et al.  Densely Interconnected Transcriptional Circuits Control Cell States in Human Hematopoiesis , 2011, Cell.

[27]  Q. Deng,et al.  Identification of Intrinsic Determinants of Midbrain Dopamine Neurons , 2006, Cell.

[28]  A. Gavalas,et al.  Hoxb1 Controls Cell Fate Specification and Proliferative Capacity of Neural Stem and Progenitor Cells , 2008, Stem cells.

[29]  C. Chiang,et al.  Specification of ventral neuron types is mediated by an antagonistic interaction between Shh and Gli3 , 2000, Nature Neuroscience.

[30]  S. Shvartsman,et al.  Temporal dynamics, spatial range, and transcriptional interpretation of the Dorsal morphogen gradient. , 2012, Current opinion in genetics & development.

[31]  George Q. Daley,et al.  Lineage Regulators Direct BMP and Wnt Pathways to Cell-Specific Programs during Differentiation and Regeneration , 2011, Cell.

[32]  M. Levine.  Transcriptional Enhancers in Animal Development and Evolution , 2010, Current Biology.

[33]  T. Jessell,et al.  Pax6 Controls Progenitor Cell Identity and Neuronal Fate in Response to Graded Shh Signaling , 1997, Cell.

[34]  M. Nakafuku,et al.  Combinatorial Roles of Olig2 and Neurogenin2 in the Coordinated Induction of Pan-Neuronal and Subtype-Specific Properties of Motoneurons , 2001, Neuron.

[35]  James Briscoe,et al.  Gene Regulatory Logic for Reading the Sonic Hedgehog Signaling Gradient in the Vertebrate Neural Tube , 2012, Cell.

[36]  Scott Barolo,et al.  Shadow enhancers: Frequently asked questions about distributed cis‐regulatory information and enhancer redundancy , 2012, BioEssays : news and reviews in molecular, cellular and developmental biology.

[37]  C. Glass,et al.  Simple combinations of lineage-determining transcription factors prime cis-regulatory elements required for macrophage and B cell identities. , 2010, Molecular cell.

[38]  Paul Theodor Pyl,et al.  HTSeq—a Python framework to work with high-throughput sequencing data , 2014, bioRxiv.

[39]  J. Dekker,et al.  The long-range interaction landscape of gene promoters , 2012, Nature.

[40]  W. Huber,et al.  which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. MAnorm: a robust model for quantitative comparison of ChIP-Seq data sets , 2011 .

[41]  A. McMahon,et al.  Growth and pattern of the mammalian neural tube are governed by partially overlapping feedback activities of the hedgehog antagonists patched 1 and Hhip1 , 2004, Development.

[42]  M. Levine,et al.  Shadow Enhancers as a Source of Evolutionary Novelty , 2008, Science.

[43]  J. Ericson,et al.  Mechanistic differences in the transcriptional interpretation of local and long-range Shh morphogen signaling. , 2012, Developmental cell.

[44]  T. Jessell.  Neuronal specification in the spinal cord: inductive signals and transcriptional codes , 2000, Nature Reviews Genetics.

[45]  M. Bulyk,et al.  A direct fate exclusion mechanism by Sonic hedgehog-regulated transcriptional repressors , 2015, Development.

[46]  Cole Trapnell,et al.  Ultrafast and memory-efficient alignment of short DNA sequences to the human genome , 2009, Genome Biology.

[47]  T. Jessell,et al.  Different Levels of Repressor Activity Assign Redundant and Specific Roles to Nkx6 Genes in Motor Neuron and Interneuron Specification , 2001, Neuron.

[48]  N. Bobola,et al.  Six2 functions redundantly immediately downstream of Hoxa2 , 2008, Development.

[49]  James Briscoe,et al.  Coordination of progenitor specification and growth in mouse and chick spinal cord , 2014, Science.

[50]  Gregory T. Reeves,et al.  Dorsal-ventral gene expression in the Drosophila embryo reflects the dynamics and precision of the dorsal nuclear gradient. , 2012, Developmental cell.

[51]  William Stafford Noble,et al.  Quantifying similarity between motifs , 2007, Genome Biology.

[52]  Clifford A. Meyer,et al.  Model-based Analysis of ChIP-Seq (MACS) , 2008, Genome Biology.

[53]  Raymond Turner,et al.  Specification , 2011, Minds and Machines.

[54]  J. Rubenstein,et al.  Ventral neural patterning by Nkx homeobox genes: Nkx6.1 controls somatic motor neuron and ventral interneuron fates. , 2000, Genes & development.

[55]  William A. Alaynick,et al.  SnapShot: Spinal Cord Development , 2011, Cell.

[56]  T. Jessell,et al.  A Homeodomain Protein Code Specifies Progenitor Cell Identity and Neuronal Fate in the Ventral Neural Tube , 2000, Cell.

[57]  T. Jessell,et al.  A hedgehog-insensitive form of patched provides evidence for direct long-range morphogen activity of sonic hedgehog in the neural tube. , 2001, Molecular cell.

[58]  Samuel L. Pfaff,et al.  Analysis of embryonic motoneuron gene regulation: derepression of general activators function in concert with enhancer factors , 2004, Development.

[59]  J. Mcwhir,et al.  Gene targeting using a mouse HPRT minigene/HPRT-deficient embryonic stem cell system: Inactivation of the mouseERCC-1 gene , 1992, Somatic cell and molecular genetics.

[60]  Boris Lenhard,et al.  A global genomic transcriptional code associated with CNS-expressed genes. , 2006, Experimental cell research.

[61]  C. Rushlow,et al.  Response to the BMP gradient requires highly combinatorial inputs from multiple patterning systems in the Drosophila embryo , 2012, Development.

[62]  Alexander van Oudenaarden,et al.  Neural-specific Sox2 input and differential Gli-binding affinity provide context and positional information in Shh-directed neural patterning. , 2012, Genes & development.

[63]  Thomas Zichner,et al.  Shadow Enhancers Are Pervasive Features of Developmental Regulatory Networks , 2016, Current Biology.

[64]  Jens Böse,et al.  Dorsal-ventral patterning of the spinal cord requires Gli3 transcriptional repressor activity. , 2002, Genes & development.

[65]  T. Jessell,et al.  Coordinate Regulation of Motor Neuron Subtype Identity and Pan-Neuronal Properties by the bHLH Repressor Olig2 , 2001, Neuron.

[66]  S. Small,et al.  A System of Repressor Gradients Spatially Organizes the Boundaries of Bicoid-Dependent Target Genes , 2012, Cell.

[67]  James Briscoe,et al.  Pattern formation in the vertebrate neural tube: a sonic hedgehog morphogen-regulated transcriptional network , 2008, Development.

[68]  J. Rubenstein,et al.  Homeobox gene Nkx2.2 and specification of neuronal identity by graded Sonic hedgehog signalling , 1999, Nature.

[69]  David H. Sharp,et al.  Canalization of Gene Expression in the Drosophila Blastoderm by Gap Gene Cross Regulation , 2009, PLoS biology.

[70]  James Briscoe,et al.  Integration of Signals along Orthogonal Axes of the Vertebrate Neural Tube Controls Progenitor Competence and Increases Cell Diversity , 2014, PLoS biology.

[71]  K. White,et al.  Su(H)-mediated repression positions gene boundaries along the dorsal-ventral axis of Drosophila embryos. , 2014, Developmental cell.

[72]  James Briscoe,et al.  Interpretation of the sonic hedgehog morphogen gradient by a temporal adaptation mechanism , 2007, Nature.