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The integrator complex subunit 6 (Ints6) confines the dorsal organizer in vertebrate embryogenesis.

Kapp LD, Abrams EW, Marlow FL, Mullins MC - PLoS Genet. (2013)

Bottom Line: We have isolated a recessive dorsalizing maternal-effect mutation disrupting the gene encoding Integrator Complex Subunit 6 (Ints6).Limiting Nodal signaling or restoring BMP signaling restores wild-type patterning to affected embryos.Our results are consistent with a novel role for Ints6 in restricting the vertebrate organizer to a dorsal domain in embryonic patterning.

View Article: PubMed Central - PubMed

Affiliation: Perelman School of Medicine at the University of Pennsylvania, Department of Cell and Developmental Biology, Philadelphia, Pennsylvania, United States of America.

ABSTRACT
Dorsoventral patterning of the embryonic axis relies upon the mutual antagonism of competing signaling pathways to establish a balance between ventralizing BMP signaling and dorsal cell fate specification mediated by the organizer. In zebrafish, the initial embryo-wide domain of BMP signaling is refined into a morphogenetic gradient following activation dorsally of a maternal Wnt pathway. The accumulation of β-catenin in nuclei on the dorsal side of the embryo then leads to repression of BMP signaling dorsally and the induction of dorsal cell fates mediated by Nodal and FGF signaling. A separate Wnt pathway operates zygotically via Wnt8a to limit dorsal cell fate specification and maintain the expression of ventralizing genes in ventrolateral domains. We have isolated a recessive dorsalizing maternal-effect mutation disrupting the gene encoding Integrator Complex Subunit 6 (Ints6). Due to widespread de-repression of dorsal organizer genes, embryos from mutant mothers fail to maintain expression of BMP ligands, fail to fully express vox and ved, two mediators of Wnt8a, display delayed cell movements during gastrulation, and severe dorsalization. Consistent with radial dorsalization, affected embryos display multiple independent axial domains along with ectopic dorsal forerunner cells. Limiting Nodal signaling or restoring BMP signaling restores wild-type patterning to affected embryos. Our results are consistent with a novel role for Ints6 in restricting the vertebrate organizer to a dorsal domain in embryonic patterning.

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Related in: MedlinePlus

Excessive axial mesoderm at the expense of ventrolateral mesoderm in p18ahub embryos.WT (A, n = 19) and stage-matched p18ahub (B, n = 20) embryos at a mid blastula stage, exhibited similar nodal related 1 (ndr1) expression. WT (C, n = 14) and age-matched p18ahub (D, n = 13) embryos at 6 hpf after synchronized matings displayed similar lefty1 (lft1) expression, although blastoderm involution was not evident in the mutants. (A–D) are animal pole views, dorsal to right. (E–L) are mid gastrula stage, except (G and H) are 3–5 somite stage or equivalent. Dorsal views, animal to top, except (H–H″) are animal pole views. WT embryos (E, n = 6) displayed marginal and axial ntl expression. In contrast, some p18ahub embryos displayed only axial ntl expression distributed circumferentially (F, n = 16), while others displayed broadened axial and reduced ventrolateral ntl expression (F inset, n = 8). In WT embryos (G, n = 22) ntl was expressed in the notochord and tail bud mesenchyme. In equivalent stage p18ahub embryos, (H–H″, n = 21), ntl expression was observed in multiple notochords terminating in smaller tail bud-like domains. Consistent with ectopic axial ntl expression, the expression of floating head (flh), a marker of notochord precursors, confined dorsally in WT embryos (I, n = 20), was expressed around the entire embryonic margin in p18ahub embryos (J, n = 16). In WT (K, n = 18) sox17 is expressed in endodermal precursor cells and in a single cluster of dorsal forerunner cells (arrow). In p18ahub embryos (L, n = 16) sox17 expression indicates the presence of ectopic clusters of dorsal forerunner cells (arrows).
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pgen-1003822-g005: Excessive axial mesoderm at the expense of ventrolateral mesoderm in p18ahub embryos.WT (A, n = 19) and stage-matched p18ahub (B, n = 20) embryos at a mid blastula stage, exhibited similar nodal related 1 (ndr1) expression. WT (C, n = 14) and age-matched p18ahub (D, n = 13) embryos at 6 hpf after synchronized matings displayed similar lefty1 (lft1) expression, although blastoderm involution was not evident in the mutants. (A–D) are animal pole views, dorsal to right. (E–L) are mid gastrula stage, except (G and H) are 3–5 somite stage or equivalent. Dorsal views, animal to top, except (H–H″) are animal pole views. WT embryos (E, n = 6) displayed marginal and axial ntl expression. In contrast, some p18ahub embryos displayed only axial ntl expression distributed circumferentially (F, n = 16), while others displayed broadened axial and reduced ventrolateral ntl expression (F inset, n = 8). In WT embryos (G, n = 22) ntl was expressed in the notochord and tail bud mesenchyme. In equivalent stage p18ahub embryos, (H–H″, n = 21), ntl expression was observed in multiple notochords terminating in smaller tail bud-like domains. Consistent with ectopic axial ntl expression, the expression of floating head (flh), a marker of notochord precursors, confined dorsally in WT embryos (I, n = 20), was expressed around the entire embryonic margin in p18ahub embryos (J, n = 16). In WT (K, n = 18) sox17 is expressed in endodermal precursor cells and in a single cluster of dorsal forerunner cells (arrow). In p18ahub embryos (L, n = 16) sox17 expression indicates the presence of ectopic clusters of dorsal forerunner cells (arrows).

Mentions: Along with the maternal Wnt pathway, the dorsal organizer also depends on Nodal signaling for its induction (reviewed in [61]). Thus, we investigated the status of the Nodal pathway in p18ahub embryos. We examined expression of the Nodal ligand nodal-related 1 (ndr1, squint) in mutant and WT embryos [62]. ndr1 induction is initiated on the dorsal side of the embryo (Figure 5A) and requires Wnt signaling similarly to boz[2]. At a mid blastula stage, we observed no significant differences in the expression of ndr1 between WT and p18ahub embryos (Figure 5B) and ndr1 expression was never observed more animally outside of its normal marginal domain through an early gastrula stage equivalent (not shown).


The integrator complex subunit 6 (Ints6) confines the dorsal organizer in vertebrate embryogenesis.

Kapp LD, Abrams EW, Marlow FL, Mullins MC - PLoS Genet. (2013)

Excessive axial mesoderm at the expense of ventrolateral mesoderm in p18ahub embryos.WT (A, n = 19) and stage-matched p18ahub (B, n = 20) embryos at a mid blastula stage, exhibited similar nodal related 1 (ndr1) expression. WT (C, n = 14) and age-matched p18ahub (D, n = 13) embryos at 6 hpf after synchronized matings displayed similar lefty1 (lft1) expression, although blastoderm involution was not evident in the mutants. (A–D) are animal pole views, dorsal to right. (E–L) are mid gastrula stage, except (G and H) are 3–5 somite stage or equivalent. Dorsal views, animal to top, except (H–H″) are animal pole views. WT embryos (E, n = 6) displayed marginal and axial ntl expression. In contrast, some p18ahub embryos displayed only axial ntl expression distributed circumferentially (F, n = 16), while others displayed broadened axial and reduced ventrolateral ntl expression (F inset, n = 8). In WT embryos (G, n = 22) ntl was expressed in the notochord and tail bud mesenchyme. In equivalent stage p18ahub embryos, (H–H″, n = 21), ntl expression was observed in multiple notochords terminating in smaller tail bud-like domains. Consistent with ectopic axial ntl expression, the expression of floating head (flh), a marker of notochord precursors, confined dorsally in WT embryos (I, n = 20), was expressed around the entire embryonic margin in p18ahub embryos (J, n = 16). In WT (K, n = 18) sox17 is expressed in endodermal precursor cells and in a single cluster of dorsal forerunner cells (arrow). In p18ahub embryos (L, n = 16) sox17 expression indicates the presence of ectopic clusters of dorsal forerunner cells (arrows).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3814294&req=5

pgen-1003822-g005: Excessive axial mesoderm at the expense of ventrolateral mesoderm in p18ahub embryos.WT (A, n = 19) and stage-matched p18ahub (B, n = 20) embryos at a mid blastula stage, exhibited similar nodal related 1 (ndr1) expression. WT (C, n = 14) and age-matched p18ahub (D, n = 13) embryos at 6 hpf after synchronized matings displayed similar lefty1 (lft1) expression, although blastoderm involution was not evident in the mutants. (A–D) are animal pole views, dorsal to right. (E–L) are mid gastrula stage, except (G and H) are 3–5 somite stage or equivalent. Dorsal views, animal to top, except (H–H″) are animal pole views. WT embryos (E, n = 6) displayed marginal and axial ntl expression. In contrast, some p18ahub embryos displayed only axial ntl expression distributed circumferentially (F, n = 16), while others displayed broadened axial and reduced ventrolateral ntl expression (F inset, n = 8). In WT embryos (G, n = 22) ntl was expressed in the notochord and tail bud mesenchyme. In equivalent stage p18ahub embryos, (H–H″, n = 21), ntl expression was observed in multiple notochords terminating in smaller tail bud-like domains. Consistent with ectopic axial ntl expression, the expression of floating head (flh), a marker of notochord precursors, confined dorsally in WT embryos (I, n = 20), was expressed around the entire embryonic margin in p18ahub embryos (J, n = 16). In WT (K, n = 18) sox17 is expressed in endodermal precursor cells and in a single cluster of dorsal forerunner cells (arrow). In p18ahub embryos (L, n = 16) sox17 expression indicates the presence of ectopic clusters of dorsal forerunner cells (arrows).
Mentions: Along with the maternal Wnt pathway, the dorsal organizer also depends on Nodal signaling for its induction (reviewed in [61]). Thus, we investigated the status of the Nodal pathway in p18ahub embryos. We examined expression of the Nodal ligand nodal-related 1 (ndr1, squint) in mutant and WT embryos [62]. ndr1 induction is initiated on the dorsal side of the embryo (Figure 5A) and requires Wnt signaling similarly to boz[2]. At a mid blastula stage, we observed no significant differences in the expression of ndr1 between WT and p18ahub embryos (Figure 5B) and ndr1 expression was never observed more animally outside of its normal marginal domain through an early gastrula stage equivalent (not shown).

Bottom Line: We have isolated a recessive dorsalizing maternal-effect mutation disrupting the gene encoding Integrator Complex Subunit 6 (Ints6).Limiting Nodal signaling or restoring BMP signaling restores wild-type patterning to affected embryos.Our results are consistent with a novel role for Ints6 in restricting the vertebrate organizer to a dorsal domain in embryonic patterning.

View Article: PubMed Central - PubMed

Affiliation: Perelman School of Medicine at the University of Pennsylvania, Department of Cell and Developmental Biology, Philadelphia, Pennsylvania, United States of America.

ABSTRACT
Dorsoventral patterning of the embryonic axis relies upon the mutual antagonism of competing signaling pathways to establish a balance between ventralizing BMP signaling and dorsal cell fate specification mediated by the organizer. In zebrafish, the initial embryo-wide domain of BMP signaling is refined into a morphogenetic gradient following activation dorsally of a maternal Wnt pathway. The accumulation of β-catenin in nuclei on the dorsal side of the embryo then leads to repression of BMP signaling dorsally and the induction of dorsal cell fates mediated by Nodal and FGF signaling. A separate Wnt pathway operates zygotically via Wnt8a to limit dorsal cell fate specification and maintain the expression of ventralizing genes in ventrolateral domains. We have isolated a recessive dorsalizing maternal-effect mutation disrupting the gene encoding Integrator Complex Subunit 6 (Ints6). Due to widespread de-repression of dorsal organizer genes, embryos from mutant mothers fail to maintain expression of BMP ligands, fail to fully express vox and ved, two mediators of Wnt8a, display delayed cell movements during gastrulation, and severe dorsalization. Consistent with radial dorsalization, affected embryos display multiple independent axial domains along with ectopic dorsal forerunner cells. Limiting Nodal signaling or restoring BMP signaling restores wild-type patterning to affected embryos. Our results are consistent with a novel role for Ints6 in restricting the vertebrate organizer to a dorsal domain in embryonic patterning.

Show MeSH
Related in: MedlinePlus