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Role of the gut endoderm in relaying left-right patterning in mice.

Viotti M, Niu L, Shi SH, Hadjantonakis AK - PLoS Biol. (2012)

Bottom Line: Iontophoretic dye injection experiments revealed planar gap junction coupling across the gut endoderm in wild-type but not Sox17 mutant embryos.They also revealed uncoupling of left and right sides of the gut endoderm in an isolated domain of gap junction intercellular communication at the midline, which in principle could function as a barrier to communication between the left and right sides of the embryo.The role for gap junction communication in LR patterning was confirmed by pharmacological inhibition, which molecularly recapitulated the mutant phenotype.

View Article: PubMed Central - PubMed

Affiliation: Developmental Biology Program, Sloan-Kettering Institute, New York, New York, United States of America.

ABSTRACT
Establishment of left-right (LR) asymmetry occurs after gastrulation commences and utilizes a conserved cascade of events. In the mouse, LR symmetry is broken at a midline structure, the node, and involves signal relay to the lateral plate, where it results in asymmetric organ morphogenesis. How information transmits from the node to the distantly situated lateral plate remains unclear. Noting that embryos lacking Sox17 exhibit defects in both gut endoderm formation and LR patterning, we investigated a potential connection between these two processes. We observed an endoderm-specific absence of the critical gap junction component, Connexin43 (Cx43), in Sox17 mutants. Iontophoretic dye injection experiments revealed planar gap junction coupling across the gut endoderm in wild-type but not Sox17 mutant embryos. They also revealed uncoupling of left and right sides of the gut endoderm in an isolated domain of gap junction intercellular communication at the midline, which in principle could function as a barrier to communication between the left and right sides of the embryo. The role for gap junction communication in LR patterning was confirmed by pharmacological inhibition, which molecularly recapitulated the mutant phenotype. Collectively, our data demonstrate that Cx43-mediated communication across gap junctions within the gut endoderm serves as a mechanism for information relay between node and lateral plate in a process that is critical for the establishment of LR asymmetry in mice.

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Sox17 mutants fail to establish LR asymmetry.Panels A-B″ and E-H′ depict wholemount mRNA in situ hybridizations, and panels C-D′ depict wholemount beta-galactosidase staining. (A and B) Nodal in ∼4 somite stage wild-type embryo (N = 14), localized around node (black asterisk) and along left LPM (white asterisks). (A′–B″) Nodal in ∼4 somite stage Sox17 mutants, around node, and either absent (A′ and B′, N = 3/8) or posteriorly restricted in left LPM (A″ and B″, N = 5/8), and present in ectopic patches on either side of midline (black arrowheads). (C and D) Nodal-LacZ in left LPM and around node of wild-type embryos (N = 6/6). (C′ and D′) Nodal-LacZ around node, absent from left LPM, and in ectopic patches on either side of midline in mutants (N = 4/4). (E and F) Lefty1/2 localized along left LPM and midline in wild-type embryo (N = 14/15). (E′–F″) Sox17 mutants exhibited absence (E′ and F′, N = 4/6) or reduction (E″ and F″, N = 5/9) of Lefty1/2 in left LPM. (G and H) Pitx2 in left LPM of wild-type embryo (N = 7/7). (G′ and H′) In the mutant, Pitx2 was absent from left LPM and punctate bilaterally (N = 5/5). (I) Diagram summarizing domains of Nodal, Lefty1/2, and Pitx2 expression in wild-type and Sox17 mutants. A, anterior; L, left; P, posterior; R, right.
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pbio-1001276-g001: Sox17 mutants fail to establish LR asymmetry.Panels A-B″ and E-H′ depict wholemount mRNA in situ hybridizations, and panels C-D′ depict wholemount beta-galactosidase staining. (A and B) Nodal in ∼4 somite stage wild-type embryo (N = 14), localized around node (black asterisk) and along left LPM (white asterisks). (A′–B″) Nodal in ∼4 somite stage Sox17 mutants, around node, and either absent (A′ and B′, N = 3/8) or posteriorly restricted in left LPM (A″ and B″, N = 5/8), and present in ectopic patches on either side of midline (black arrowheads). (C and D) Nodal-LacZ in left LPM and around node of wild-type embryos (N = 6/6). (C′ and D′) Nodal-LacZ around node, absent from left LPM, and in ectopic patches on either side of midline in mutants (N = 4/4). (E and F) Lefty1/2 localized along left LPM and midline in wild-type embryo (N = 14/15). (E′–F″) Sox17 mutants exhibited absence (E′ and F′, N = 4/6) or reduction (E″ and F″, N = 5/9) of Lefty1/2 in left LPM. (G and H) Pitx2 in left LPM of wild-type embryo (N = 7/7). (G′ and H′) In the mutant, Pitx2 was absent from left LPM and punctate bilaterally (N = 5/5). (I) Diagram summarizing domains of Nodal, Lefty1/2, and Pitx2 expression in wild-type and Sox17 mutants. A, anterior; L, left; P, posterior; R, right.

Mentions: Sox17 mutant embryos exhibit a dysmorphic heart, have an open ventral body wall, and fail to turn (Figure S1A–S1D′) [17]–[19]. Since these features are characteristic of mutants with defects in LR patterning [8], they prompted us to determine whether LR asymmetry is established in Sox17 mutants. To do so, we analyzed the expression of components of the core circuitry controlling the establishment of LR asymmetry in mice: genes encoding the TGFβ family proteins Nodal and Lefty2, and the homeodomain protein Pitx2. In E8.5 wild-type embryos, Nodal was expressed around the node and along the left LPM (Figure 1A and 1B). In stage-matched Sox17 mutants, Nodal was present around the node, but absent (three out of eight embryos analyzed) or reduced and restricted posteriorly (five out of eight embryos analyzed) in the left LPM (Figure 1A′–1B″). Sox17 mutants also exhibited ectopic patchy domains of Nodal expression on both the left and right sides. Notably, these ectopic patches of gene expression were not in cells of the mesoderm layer; instead, they were located superficially and were confined to the endoderm layer of the embryo, as analyzed below.


Role of the gut endoderm in relaying left-right patterning in mice.

Viotti M, Niu L, Shi SH, Hadjantonakis AK - PLoS Biol. (2012)

Sox17 mutants fail to establish LR asymmetry.Panels A-B″ and E-H′ depict wholemount mRNA in situ hybridizations, and panels C-D′ depict wholemount beta-galactosidase staining. (A and B) Nodal in ∼4 somite stage wild-type embryo (N = 14), localized around node (black asterisk) and along left LPM (white asterisks). (A′–B″) Nodal in ∼4 somite stage Sox17 mutants, around node, and either absent (A′ and B′, N = 3/8) or posteriorly restricted in left LPM (A″ and B″, N = 5/8), and present in ectopic patches on either side of midline (black arrowheads). (C and D) Nodal-LacZ in left LPM and around node of wild-type embryos (N = 6/6). (C′ and D′) Nodal-LacZ around node, absent from left LPM, and in ectopic patches on either side of midline in mutants (N = 4/4). (E and F) Lefty1/2 localized along left LPM and midline in wild-type embryo (N = 14/15). (E′–F″) Sox17 mutants exhibited absence (E′ and F′, N = 4/6) or reduction (E″ and F″, N = 5/9) of Lefty1/2 in left LPM. (G and H) Pitx2 in left LPM of wild-type embryo (N = 7/7). (G′ and H′) In the mutant, Pitx2 was absent from left LPM and punctate bilaterally (N = 5/5). (I) Diagram summarizing domains of Nodal, Lefty1/2, and Pitx2 expression in wild-type and Sox17 mutants. A, anterior; L, left; P, posterior; R, right.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3295824&req=5

pbio-1001276-g001: Sox17 mutants fail to establish LR asymmetry.Panels A-B″ and E-H′ depict wholemount mRNA in situ hybridizations, and panels C-D′ depict wholemount beta-galactosidase staining. (A and B) Nodal in ∼4 somite stage wild-type embryo (N = 14), localized around node (black asterisk) and along left LPM (white asterisks). (A′–B″) Nodal in ∼4 somite stage Sox17 mutants, around node, and either absent (A′ and B′, N = 3/8) or posteriorly restricted in left LPM (A″ and B″, N = 5/8), and present in ectopic patches on either side of midline (black arrowheads). (C and D) Nodal-LacZ in left LPM and around node of wild-type embryos (N = 6/6). (C′ and D′) Nodal-LacZ around node, absent from left LPM, and in ectopic patches on either side of midline in mutants (N = 4/4). (E and F) Lefty1/2 localized along left LPM and midline in wild-type embryo (N = 14/15). (E′–F″) Sox17 mutants exhibited absence (E′ and F′, N = 4/6) or reduction (E″ and F″, N = 5/9) of Lefty1/2 in left LPM. (G and H) Pitx2 in left LPM of wild-type embryo (N = 7/7). (G′ and H′) In the mutant, Pitx2 was absent from left LPM and punctate bilaterally (N = 5/5). (I) Diagram summarizing domains of Nodal, Lefty1/2, and Pitx2 expression in wild-type and Sox17 mutants. A, anterior; L, left; P, posterior; R, right.
Mentions: Sox17 mutant embryos exhibit a dysmorphic heart, have an open ventral body wall, and fail to turn (Figure S1A–S1D′) [17]–[19]. Since these features are characteristic of mutants with defects in LR patterning [8], they prompted us to determine whether LR asymmetry is established in Sox17 mutants. To do so, we analyzed the expression of components of the core circuitry controlling the establishment of LR asymmetry in mice: genes encoding the TGFβ family proteins Nodal and Lefty2, and the homeodomain protein Pitx2. In E8.5 wild-type embryos, Nodal was expressed around the node and along the left LPM (Figure 1A and 1B). In stage-matched Sox17 mutants, Nodal was present around the node, but absent (three out of eight embryos analyzed) or reduced and restricted posteriorly (five out of eight embryos analyzed) in the left LPM (Figure 1A′–1B″). Sox17 mutants also exhibited ectopic patchy domains of Nodal expression on both the left and right sides. Notably, these ectopic patches of gene expression were not in cells of the mesoderm layer; instead, they were located superficially and were confined to the endoderm layer of the embryo, as analyzed below.

Bottom Line: Iontophoretic dye injection experiments revealed planar gap junction coupling across the gut endoderm in wild-type but not Sox17 mutant embryos.They also revealed uncoupling of left and right sides of the gut endoderm in an isolated domain of gap junction intercellular communication at the midline, which in principle could function as a barrier to communication between the left and right sides of the embryo.The role for gap junction communication in LR patterning was confirmed by pharmacological inhibition, which molecularly recapitulated the mutant phenotype.

View Article: PubMed Central - PubMed

Affiliation: Developmental Biology Program, Sloan-Kettering Institute, New York, New York, United States of America.

ABSTRACT
Establishment of left-right (LR) asymmetry occurs after gastrulation commences and utilizes a conserved cascade of events. In the mouse, LR symmetry is broken at a midline structure, the node, and involves signal relay to the lateral plate, where it results in asymmetric organ morphogenesis. How information transmits from the node to the distantly situated lateral plate remains unclear. Noting that embryos lacking Sox17 exhibit defects in both gut endoderm formation and LR patterning, we investigated a potential connection between these two processes. We observed an endoderm-specific absence of the critical gap junction component, Connexin43 (Cx43), in Sox17 mutants. Iontophoretic dye injection experiments revealed planar gap junction coupling across the gut endoderm in wild-type but not Sox17 mutant embryos. They also revealed uncoupling of left and right sides of the gut endoderm in an isolated domain of gap junction intercellular communication at the midline, which in principle could function as a barrier to communication between the left and right sides of the embryo. The role for gap junction communication in LR patterning was confirmed by pharmacological inhibition, which molecularly recapitulated the mutant phenotype. Collectively, our data demonstrate that Cx43-mediated communication across gap junctions within the gut endoderm serves as a mechanism for information relay between node and lateral plate in a process that is critical for the establishment of LR asymmetry in mice.

Show MeSH
Related in: MedlinePlus