Regulatory sequences driving expression of the sea urchin Otp homeobox gene in oral ectoderm cells

Several homeobox genes are expressed in the sea urchin embryo but their roles in development have yet to be elucidated. Of particular interest are homologues of homeobox genes that in mouse and Drosophila are involved in patterning the developing central nervous system (CNS). Here, we report the cloning of an orthopedia (Otp)-related gene from Paracentrotus lividus, PlOtp. Otp is a single copy zygotic gene that presents a unique and highly restricted expression pattern. Transcripts were first detected at the mid-gastrula stage in two pairs of oral ectoderm cells located in a ventrolateral position, overlying primary mesenchyme cell (PMC) clusters. Increases in both transcript abundance and the number of Otp-expressing cells were observed at prism and pluteus stages. Otp transcripts are symmetrically distributed in a few ectodermal cells of the oral field. Labelled cells were observed close to sites of active skeletal rod growth (tips of the budding oral and anal arms), and at the juxtaposition of stomodeum and foregut. Chemicals known to perturb PMC patterning along animal-vegetal and oral-aboral axes altered the pattern of Otp expression. Vegetalization by LiCl caused a shift in Otp-expressing cells toward the animal pole, adjacent to shifted PMC aggregates. Nickel treatment induced expression of the Otp gene in an increased number of ectodermal cells, which adopted a radialized pattern. Finally, ectopic expression of Otp mRNA affected patterning along the oral-aboral axis and caused skeletal abnormalities that resembled those exhibited by nickel-treated embryos. From these results, we conclude that the Otp homeodomain gene is involved in short-range cell signalling within the oral ectoderm for patterning the endoskeleton of the larva through epithelial-mesenchymal interactions.

Download Full-text

Hedgehog signaling is required for pituitary gland development

Development ◽

10.1242/dev.128.3.377 ◽

2001 ◽

Vol 128 (3) ◽

pp. 377-386 ◽

Cited By ~ 8

Author(s):

M. Treier ◽

S. O'Connell ◽

A. Gleiberman ◽

J. Price ◽

D.P. Szeto ◽

...

Keyword(s):

Pituitary Gland ◽

Hedgehog Signaling ◽

Homeobox Gene ◽

Cell Types ◽

Oral Epithelium ◽

Regulatory Sequences ◽

Rathke’S Pouch ◽

Rathke's Pouch ◽

Oral Ectoderm ◽

Gland Development

Pituitary gland development serves as an excellent model system in which to study the emergence of distinct cell types from a common primordium in mammalian organogenesis. We have investigated the role of the morphogen Sonic hedgehog (SHH) in outgrowth and differentiation of the pituitary gland using loss- and gain-of-function studies in transgenic mice. Shh is expressed throughout the ventral diencephalon and the oral ectoderm, but its expression is subsequently absent from the nascent Rathke's pouch as soon as it becomes morphologically visible, creating a Shh boundary within the oral epithelium. We used oral ectoderm/Rathke's pouch-specific 5′ regulatory sequences (Pitx1(HS)) from the bicoid related pituitary homeobox gene (Pitx1) to target overexpression of the Hedgehog inhibitor Hip (Huntingtin interacting protein) to block Hedgehog signaling, finding that SHH is required for proliferation of the pituitary gland. In addition, we provide evidence that Hedgehog signaling, acting at the Shh boundary within the oral ectoderm, may exert a role in differentiation of ventral cell types (gonadotropes and thyrotropes) by inducing Bmp2 expression in Rathke's pouch, which subsequently regulates expression of ventral transcription factors, particularly Gata2. Furthermore, our data suggest that Hedgehog signaling, together with FGF8/10 signaling, synergizes to regulate expression of the LIM homeobox gene Lhx3, which has been proved to be essential for initial pituitary gland formation. Thus, SHH appears to exert effects on both proliferation and cell-type determination in pituitary gland development.

Download Full-text

Autonomous and non-autonomous differentiation of ectoderm in different sea urchin species

Development ◽

10.1242/dev.121.5.1497 ◽

1995 ◽

Vol 121 (5) ◽

pp. 1497-1505 ◽

Cited By ~ 7

Author(s):

A.H. Wikramanayake ◽

B.P. Brandhorst ◽

W.H. Klein

Keyword(s):

Sea Urchin ◽

Strongylocentrotus Purpuratus ◽

Protein A ◽

Cellular Interactions ◽

V Protein ◽

Lytechinus Pictus ◽

Sea Urchin Embryo ◽

Oral Ectoderm ◽

Temporal And Spatial

During early embryogenesis, the highly regulative sea urchin embryo relies extensively on cell-cell interactions for cellular specification. Here, the role of cellular interactions in the temporal and spatial expression of markers for oral and aboral ectoderm in Strongylocentrotus purpuratus and Lytechinus pictus was investigated. When pairs of mesomeres or animal caps, which are fated to give rise to ectoderm, were isolated and cultured they developed into ciliated embryoids that were morphologically polarized. In animal explants from S. purpuratus, the aboral ectoderm-specific Spec1 gene was activated at the same time as in control embryos and at relatively high levels. The Spec1 protein was restricted to the squamous epithelial cells in the embryoids suggesting that an oral-aboral axis formed and aboral ectoderm differentiation occurred correctly. However, the Ecto V protein, a marker for oral ectoderm differentiation, was detected throughout the embryoid and no stomodeum or ciliary band formed. These results indicated that animal explants from S. purpuratus were autonomous in their ability to form an oral-aboral axis and to differentiate aboral ectoderm, but other aspects of ectoderm differentiation require interaction with vegetal blastomeres. In contrast to S. purpuratus, aboral ectoderm-specific genes were not expressed in animal explants from L. pictus even though the resulting embryoids were morphologically very similar to those of S. purpuratus. Recombination of the explants with vegetal blastomeres or exposure to the vegetalizing agent LiCl restored activity of aboral ectoderm-specific genes, suggesting the requirement of a vegetal induction for differentiation of aboral ectoderm cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Download Full-text

Encoding regulatory state boundaries in the pregastrular oral ectoderm of the sea urchin embryo

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1323105111 ◽

2014 ◽

Vol 111 (10) ◽

pp. E906-E913 ◽

Cited By ~ 35

Author(s):

Enhu Li ◽

Miao Cui ◽

Isabelle S. Peter ◽

Eric H. Davidson

Keyword(s):

Spatial Pattern ◽

Sea Urchin ◽

Regulatory Network ◽

Regulatory State ◽

Apical Plate ◽

Sea Urchin Embryo ◽

Oral Ectoderm ◽

Positive Regulators ◽

Gene Regulatory ◽

Per Se

By gastrulation the ectodermal territories of the sea urchin embryo have developed an unexpectedly complex spatial pattern of sharply bounded regulatory states, organized orthogonally with respect to the animal/vegetal and oral/aboral axes of the embryo. Although much is known of the gene regulatory network (GRN) linkages that generate these regulatory states, the principles by which the boundaries between them are positioned and maintained have remained undiscovered. Here we determine the encoded genomic logic responsible for the boundaries of the oral aspect of the embryo that separate endoderm from ectoderm and ectoderm from neurogenic apical plate and that delineate the several further subdivisions into which the oral ectoderm per se is partitioned. Comprehensive regulatory state maps, including all spatially expressed oral ectoderm regulatory genes, were established. The circuitry at each boundary deploys specific repressors of regulatory states across the boundary, identified in this work, plus activation by broadly expressed positive regulators. These network linkages are integrated with previously established interactions on the oral/aboral axis to generate a GRN model encompassing the 2D organization of the regulatory state pattern in the pregastrular oral ectoderm of the embryo.

Download Full-text

A hyaline layer protein that becomes localized to the oral ectoderm and foregut of sea urchin embryos

Developmental Biology ◽

10.1016/0012-1606(90)90056-o ◽

1990 ◽

Vol 140 (1) ◽

pp. 93-104 ◽

Cited By ~ 50

Author(s):

James A. Coffman ◽

David R. McClay

Keyword(s):

Sea Urchin ◽

Sea Urchin Embryos ◽

Oral Ectoderm

Download Full-text

Lim1-related homeobox gene (HpLim1) expressed in sea urchin embryo

Zygote ◽

10.1017/s0967199400130394 ◽

1999 ◽

Vol 8 (S1) ◽

pp. S71-S72

Author(s):

Keiko Mitsunaga-Nakatsubo ◽

Takahiko Kawasaki ◽

Koichi Takeda ◽

Koji Akasaka ◽

Hiraku Shimada

Keyword(s):

Sea Urchin ◽

Expression Patterns ◽

Homeobox Gene ◽

Mesoderm Induction ◽

Over Expression ◽

Lim Domains ◽

Sea Urchin Embryo ◽

Cell Type Specific ◽

Islet 1

A characteristic cysteine-rich motif, LIM domain, was first detected in three different transcription factors: lin-11, Islet-1 and mec-3. A feature shared by these genes is the presence of two LIM domains linked to a DNA-binding homeodomain (Sánchez-García et al., 1994). LIM homeodomain (LHX) proteins have been reported to be implicated in a variety of developmental processes (Dawid et al., 1998).Expression patterns of LHX genes have been analysed in a wide variety of organisms and reported to be cell-type specific (Dawid et al., 1998). In vertebrates, they are expressed in organiser equivalent regions at the gastrula stage, suggesting their involvement in mesoderm induction (Taira et al., 1992; Barnes et al., 1994; Toyama et al., 1995). Hrlim, an ascidian Lim3, zygotically expresses in the endoderm lineage before gastrulation, suggesting that it is involved in the endoderm determination (Wada et al., 1995).Here, cDNA cloning of the Lim1-related homeobox gene (HpLim1) of the sea urchin, Hemicentrotus pulcherrimus, is described together with the spatially as well as temporally regulated expression of HpLim1 during sea urchin development. A possible role of HpLiml in sea urchin development is also discussed based on its spatial pattern of expression and on the result of an over-expression study.

Download Full-text

Lim1 related homeobox gene (HpLim1) expressed in sea urchin embryos

Development Growth & Differentiation ◽

10.1046/j.1440-169x.1999.413432.x ◽

1999 ◽

Vol 41 (3) ◽

pp. 273-282 ◽

Cited By ~ 7

Author(s):

Takahiko Kawasaki ◽

Keiko Mitsunaga-Nakatsubo ◽

Koichi Takeda ◽

Koji Akasaka ◽

Hiraku Shimada

Keyword(s):

Sea Urchin ◽

Homeobox Gene ◽

Sea Urchin Embryos

Download Full-text

Inhibition of TLX3 and NKX2-5 in t(5;14)(q35;q32) T-ALL after Blocking Remote 3′-BCL11B Enhancer Sequences with Matching DNA Oligos Reveals Coregulation by PU.1 and HMGA1.

Blood ◽

10.1182/blood.v108.11.2212.2212 ◽

2006 ◽

Vol 108 (11) ◽

pp. 2212-2212

Author(s):

Stefan Nagel ◽

Michaela Scherr ◽

Alexander Kel ◽

Klaus Hornischer ◽

Gregory E. Crawford ◽

...

Keyword(s):

T Cell ◽

Nuclear Matrix ◽

Gene Activation ◽

Regulatory Region ◽

Homeobox Genes ◽

Homeobox Gene ◽

Genomic Analysis ◽

Regulatory Sequences ◽

Promoter Regions ◽

Gene Dysregulation

Abstract In T-cell acute lymphoblastic leukaemia (T-ALL) alternative t(5;14)(q35;q32.2) forms effect leukemic dysregulation of either TLX3 or NKX2-5 homeobox genes at 5q35 by juxtaposition with 3′-BCL11B at 14q32.2. Putative regulatory sequences underlying ectopic homeobox gene activation in t(5;14), and their mode of action have remained poorly understood mainly because breakpoints at 14q32.2 are widely scattered over the ~1 Mbp genomic desert region. We pooled cytogenetic data from t(5;14) cell lines together with published clinical data to refine the BCL11B downstream breakpoint cluster region (bcr). Ectopic homeobox gene dysregulation was investigated by DNA-i(nhibitory-treatments) with 26-mer double-stranded DNA oligo(nucleotide)s directed against putative enhancers using NKX2-5 expression as endpoints. Enhancer targets were provisionally identified from orphan T-cell DNase-I hypersensive sites (DNaseI-HS) located between 3′-BCL11B and VRK1. NKX2-5 downregulation in t(5;14) PEER cells was almost entirely restricted to DNAi targeting enhancers within the distal bcr and was dose- and sequence-dependent. Interestingly, enhancers near 3′-BCL11B regulated that gene only. These data imply that enhancer-promoter distances and/or locations are important for long-range gene regulation. Chromatin immunoprecipation assays showed that the four most effectual NKX2-5 ectopic enhancers were hyperacetylated. These enhancers clustered ~1 Mbp downstream of BCL11B, within a region displaying multiple regulatory stigmata, including a TCRA-enhancer motif, and abyssal sequence-conservation (“5-Way Regulatory Potential”). Paradoxically, although TLX3/NKX2-5 promoter/exon-1 regions were hypo-acetylated, their expression decreased after TSA treatment, implying extrinsic regulation by factor(s) subject to acetylation-control. PU.1 is known to get transcriptionally repressed by TSA and potentially binds TLX3/NKX2-5 upstream promoter regions. Knockdown of PU.1 effected downregulation of both homeobox genes. Moreover, genomic analysis showed preferential enrichment near validated ectopic enhancers of binding sites for the PU.1-cofactor HMGA1, knockdown of which also inhibited NKX2-5 in PEER cells. Analysis of nuclear matrix attachment (NMA) in PEER cells showed enhanced attachment near to the most effectual enhancer cluster which was alleviated by TSA-treatment. Interestingly, the juxtapositional genomic regions of “active” ins(14;5) rearrangements driving NKX2-5 expression exhibited tight NMA, forming structures reminiscent of “active chromatin hubs”. These findings lead us to propose that HMGA1 and PU.1 co-regulate ectopic homeobox gene expression in t(5;14) T-ALL by interactions mediated at the nuclear matrix, possibly mediated by SATB1 binding. Our data document homeobox gene dysregulation by a novel regulatory region at 3′-BCL11B responsive to HDAC-inhibition and highlight a novel class of potential therapeutic target amid “junk” DNA.

Download Full-text