Expression of the oscillating gene her1 is directly regulated by hairy/enhancer of split, T‐box, and suppressor of hairless proteins in the zebrafish segmentation clock

Somites are segmental units of the mesoderm in vertebrate embryos that give rise to the axial skeleton, muscle, and dermis. Somitogenesis occurs in a periodic manner and is governed by a segmentation clock that causes cells to undergo repeated oscillations of gene expression. Here, we present a detailed analysis of cis‐regulatory elements that control oscillating expression of the zebrafish her1 gene in the anterior presomitic mesoderm. We identify binding sites for Her proteins and demonstrate that they are necessary for transcriptional repression. This result confirms that direct negative autoregulation of her gene expression constitutes part of the oscillator mechanism. We also characterize binding sites for fused somites/Tbx24 and Suppressor of Hairless proteins and show that they are required for activation of her1 expression. These data provide the foundation for a precise description of the regulatory grammar that defines oscillating gene expression in the zebrafish segmentation clock. Developmental Dynamics 238:2745–2759, 2009. © 2009 Wiley‐Liss, Inc.

[1]  O. Pourquié,et al.  Notch signalling is required for cyclic expression of the hairy-like gene HES1 in the presomitic mesoderm. , 2000, Development.

[2]  Piotr Sliz,et al.  Cooperative assembly of higher-order Notch complexes functions as a switch to induce transcription , 2007, Proceedings of the National Academy of Sciences.

[3]  Julian Lewis,et al.  Setting the Tempo in Development: An Investigation of the Zebrafish Somite Clock Mechanism , 2007, PLoS biology.

[4]  A. Oates,et al.  Coordination of symmetric cyclic gene expression during somitogenesis by Suppressor of Hairless involves regulation of retinoic acid catabolism. , 2007, Developmental biology.

[5]  A. Oates,et al.  Completing the set of h/E(spl) cyclic genes in zebrafish: her12 and her15 reveal novel modes of expression and contribute to the segmentation clock. , 2007, Developmental biology.

[6]  C. Nüsslein-Volhard,et al.  Zebrafish segmentation and pair-rule patterning. , 1998, Developmental genetics.

[7]  Olivier Pourquié,et al.  FGF Signaling Controls Somite Boundary Position and Regulates Segmentation Clock Control of Spatiotemporal Hox Gene Activation , 2001, Cell.

[8]  Nigel A. Brown,et al.  Waves of mouse Lunatic fringe expression, in four-hour cycles at two-hour intervals, precede somite boundary formation , 1998, Current Biology.

[9]  David Ish-Horowicz,et al.  Notch signalling and the synchronization of the somite segmentation clock , 2000, Nature.

[10]  Y. Bessho,et al.  Dynamic expression and essential functions of Hes7 in somite segmentation. , 2001, Genes & development.

[11]  A. Kispert,et al.  An interacting network of T-box genes directs gene expression and fate in the zebrafish mesoderm , 2003, Proceedings of the National Academy of Sciences of the United States of America.

[12]  Diethard Tautz,et al.  The role of Suppressor of Hairless in Notch mediated signalling during zebrafish somitogenesis , 2003, Mechanisms of Development.

[13]  D. Tautz,et al.  her11 is involved in the somitogenesis clock in zebrafish , 2004, Development Genes and Evolution.

[14]  Stefan Hans,et al.  Anterior and posterior waves of cyclic her1 gene expression are differentially regulated in the presomitic mesoderm of zebrafish , 2003, Development.

[15]  A. Kuroiwa,et al.  Fgf/MAPK signalling is a crucial positional cue in somite boundary formation. , 2001, Development.

[16]  Ryoichiro Kageyama,et al.  Periodic repression by the bHLH factor Hes7 is an essential mechanism for the somite segmentation clock. , 2003, Genes & development.

[17]  Makoto Furutani-Seiki,et al.  Tbx24, encoding a T-box protein, is mutated in the zebrafish somite-segmentation mutant fused somites , 2002, Nature Genetics.

[18]  M. Eisen,et al.  Identification of direct T-box target genes in the developing zebrafish mesoderm , 2009, Development.

[19]  Ryoichiro Kageyama,et al.  Real-time imaging of the somite segmentation clock: Revelation of unstable oscillators in the individual presomitic mesoderm cells , 2006, Proceedings of the National Academy of Sciences of the United States of America.

[20]  S. Cole,et al.  Oscillatory lunatic fringe activity is crucial for segmentation of the anterior but not posterior skeleton , 2008, Development.

[21]  Andrew C Oates,et al.  Hairy/E(spl)-related (Her) genes are central components of the segmentation oscillator and display redundancy with the Delta/Notch signaling pathway in the formation of anterior segmental boundaries in the zebrafish , 2002 .

[22]  Olivier Pourquié,et al.  fgf8 mRNA decay establishes a gradient that couples axial elongation to patterning in the vertebrate embryo , 2004, Nature.

[23]  V. Papaioannou,et al.  T-box genes in vertebrate development. , 2005, Annual review of genetics.

[24]  Olivier Cinquin,et al.  Repressor Dimerization in the Zebrafish Somitogenesis Clock , 2007, PLoS Comput. Biol..

[25]  R. Beddington,et al.  Axial skeletal defects caused by mutation in the spondylocostal dysplasia/pudgy gene Dll3 are associated with disruption of the segmentation clock within the presomitic mesoderm. , 2002, Development.

[26]  J. Smith,et al.  Determinants of T box protein specificity. , 2001, Development.

[27]  T. Vogt,et al.  Clock regulatory elements control cyclic expression of Lunatic fringe during somitogenesis. , 2002, Developmental cell.

[28]  M. Taketo,et al.  Wnt3a/β-catenin signaling controls posterior body development by coordinating mesoderm formation and segmentation , 2007, Development.

[29]  Jun Kanno,et al.  Mouse Nkd1, a Wnt antagonist, exhibits oscillatory gene expression in the PSM under the control of Notch signaling , 2004, Mechanisms of Development.

[30]  C. Voolstra,et al.  Comparative analysis of somitogenesis related genes of the hairy/Enhancer of split class in Fugu and zebrafish , 2002, BMC Genomics.

[31]  Paul Flicek,et al.  A gene regulatory network directed by zebrafish No tail accounts for its roles in mesoderm formation , 2009, Proceedings of the National Academy of Sciences.

[32]  A. Fischer,et al.  Delta–Notch—and then? Protein interactions and proposed modes of repression by Hes and Hey bHLH factors , 2007, Nucleic acids research.

[33]  E. Lai,et al.  Discrete enhancer elements mediate selective responsiveness of enhancer of split complex genes to common transcriptional activators. , 1999, Developmental biology.

[34]  R. Kageyama,et al.  Structure, chromosomal locus, and promoter analysis of the gene encoding the mouse helix-loop-helix factor HES-1. Negative autoregulation through the multiple N box elements. , 1994, The Journal of biological chemistry.

[35]  S. Bray,et al.  Target Specificities of DrosophilaEnhancer of split Basic Helix-Loop-Helix Proteins , 1999, Molecular and Cellular Biology.

[36]  H. Farin,et al.  Transcriptional Repression by the T-box Proteins Tbx18 and Tbx15 Depends on Groucho Corepressors*♦ , 2007, Journal of Biological Chemistry.

[37]  C. Nüsslein-Volhard,et al.  Control of her1 expression during zebrafish somitogenesis by a delta-dependent oscillator and an independent wave-front activity. , 2000, Genes & development.

[38]  Scott A. Holley,et al.  Zebrafish Whole Mount High-Resolution Double Fluorescent In Situ Hybridization , 2009, Journal of visualized experiments : JoVE.

[39]  C. Kimmel,et al.  Stages of embryonic development of the zebrafish , 1995, Developmental dynamics : an official publication of the American Association of Anatomists.

[40]  Christian Wehrle,et al.  Wnt3a plays a major role in the segmentation clock controlling somitogenesis. , 2003, Developmental cell.

[41]  Ryoichiro Kageyama,et al.  The initiation and propagation of Hes7 oscillation are cooperatively regulated by Fgf and notch signaling in the somite segmentation clock. , 2007, Developmental cell.

[42]  Jie Chen,et al.  A Complex Oscillating Network of Signaling Genes Underlies the Mouse Segmentation Clock , 2006, Science.

[43]  Scott A Holley,et al.  The genetics and embryology of zebrafish metamerism , 2007, Developmental dynamics : an official publication of the American Association of Anatomists.

[44]  S. Bray Notch signalling: a simple pathway becomes complex , 2006, Nature Reviews Molecular Cell Biology.

[45]  Julian Lewis Autoinhibition with Transcriptional Delay A Simple Mechanism for the Zebrafish Somitogenesis Oscillator , 2003, Current Biology.

[46]  D. Ish-Horowicz,et al.  Periodic Lunatic fringe expression is controlled during segmentation by a cyclic transcriptional enhancer responsive to notch signaling. , 2002, Developmental cell.

[47]  P. Tam,et al.  The control of somitogenesis in mouse embryos. , 1981, Journal of embryology and experimental morphology.

[48]  C. Kimmel,et al.  Two linked hairy/Enhancer of split-related zebrafish genes, her1 and her7, function together to refine alternating somite boundaries. , 2002, Development.

[49]  O. Pourquié,et al.  Oscillations of the snail genes in the presomitic mesoderm coordinate segmental patterning and morphogenesis in vertebrate somitogenesis. , 2006, Developmental cell.

[50]  A. Fritz,et al.  Zebrafish Mesp family genes, mesp-a and mesp-b are segmentally expressed in the presomitic mesoderm, and Mesp-b confers the anterior identity to the developing somites. , 2000, Development.

[51]  Albert Cardona,et al.  Dynamics of zebrafish somitogenesis , 2008, Developmental dynamics : an official publication of the American Association of Anatomists.

[52]  Stefan Zeiser,et al.  Number of active transcription factor binding sites is essential for the Hes7 oscillator , 2006, Theoretical Biology and Medical Modelling.

[53]  Christel Brou,et al.  Signalling downstream of activated mammalian Notch , 1995, Nature.

[54]  Winfried Wiegraebe,et al.  A β-catenin gradient links the clock and wavefront systems in mouse embryo segmentation , 2008, Nature Cell Biology.

[55]  Nian Zhang,et al.  Negative feedback loop formed by Lunatic fringe and Hes7 controls their oscillatory expression during somitogenesis , 2005, Genesis.

[56]  C. Murre,et al.  Helix-Loop-Helix Proteins: Regulators of Transcription in Eucaryotic Organisms , 2000, Molecular and Cellular Biology.

[57]  Robert Geisler,et al.  her1 and the notch pathway function within the oscillator mechanism that regulates zebrafish somitogenesis. , 2002, Development.

[58]  Olivier Pourquié,et al.  The lunatic Fringe gene is a target of the molecular clock linked to somite segmentation in avian embryos , 1998, Current Biology.

[59]  Functional importance of evolutionally conserved Tbx6 binding sites in the presomitic mesoderm-specific enhancer of Mesp2 , 2008, Development.

[60]  F Radtke,et al.  Oscillating expression of c-Hey2 in the presomitic mesoderm suggests that the segmentation clock may use combinatorial signaling through multiple interacting bHLH factors. , 2000, Developmental biology.

[61]  Robert Geisler,et al.  beamter/deltaC and the role of Notch ligands in the zebrafish somite segmentation, hindbrain neurogenesis and hypochord differentiation. , 2005, Developmental biology.

[62]  Akihiro Urasaki,et al.  Functional Dissection of the Tol2 Transposable Element Identified the Minimal cis-Sequence and a Highly Repetitive Sequence in the Subterminal Region Essential for Transposition , 2006, Genetics.

[63]  E. C. Zeeman,et al.  A clock and wavefront model for control of the number of repeated structures during animal morphogenesis. , 1976, Journal of theoretical biology.

[64]  O. Pourquié,et al.  Avian hairy Gene Expression Identifies a Molecular Clock Linked to Vertebrate Segmentation and Somitogenesis , 1997, Cell.