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Dev Biol
2009 Jan 01;3251:249-62. doi: 10.1016/j.ydbio.2008.10.031.
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Maternal Interferon Regulatory Factor 6 is required for the differentiation of primary superficial epithelia in Danio and Xenopus embryos.
Sabel JL, d'Alençon C, O'Brien EK, Van Otterloo E, Lutz K, Cuykendall TN, Schutte BC, Houston DW, Cornell RA.
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Early in the development of animal embryos, superficial cells of the blastula form a distinct lineage and adopt an epithelial morphology. In different animals, the fate of these primary superficial epithelial (PSE) cells varies, and it is unclear whether pathways governing segregation of blastomeres into the PSE lineage are conserved. Mutations in the gene encoding Interferon Regulatory Factor 6 (IRF6) are associated with syndromic and non-syndromic forms of cleft lip and palate, consistent with a role for Irf6 in development of oral epithelia, and mouse Irf6 targeted null mutant embryos display abnormal differentiation of oral epithelia and skin. In Danio rerio (zebrafish) and Xenopus laevis (African clawed frog) embryos, zygotic irf6 transcripts are present in many epithelial tissues including the presumptive PSE cells and maternal irf6 transcripts are present throughout all cells at the blastula stage. Injection of antisense oligonucleotides with ability to disrupt translation of irf6 transcripts caused little or no effect on development. By contrast, injection of RNA encoding a putative dominant negative Irf6 caused epiboly arrest, loss of gene expression characteristic of the EVL, and rupture of the embryo at late gastrula stage. The dominant negative Irf6 disrupted EVL gene expression in a cell autonomous fashion. These results suggest that Irf6 translated in the oocyte or unfertilized egg suffices for early development. Supporting the importance of maternal Irf6, we show that depletion of maternal irf6 transcripts in X. laevis embryos leads to gastrulation defects and rupture of the superficialepithelium. These experiments reveal a conserved role for maternally-encoded Irf6 in differentiation of a simple epithelium in X. laevis and D. rerio. This epithelium constitutes a novel model tissue in which to explore the Irf6 regulatory pathway.
Fig. 5. Expression of irf6 in Xenopus laevis. (A) RT-PCR analysis of irf6 expression at different stages of development. Developmental stages are indicated at the top (Nieuwkoop and Faber, 1956). − RT, negative control reaction without reverse transcriptase. (B–G) Whole mount in situ hybridization for irf6. (B) Stage 10.5, left is vegetal view, arrow indicates expression in the dorsal marginal zone. Right is animal pole view. (C) Stage 11, expression is throughout marginal zone, but beginning to be lost from dorsal midline. (D) Stage 12.5, expression around blastopore, absent from prospective notochord. (E) Stage 18, expression flanking prospective tailbud region, arrow. (F) Stage 28 and (G) stage 30, notable expression in tailbud, arrowheads. (H) Section of animal cap hybridized with an irrelevant control probe (xnr3). (I) Section of animal cap hybridized with an antisense irf6 probe. Diffuse expression is found in both superficial and deep layers. (J) Quantitative RT-PCR of irf6 (green) and esr6e (blue) expression in ectodermal explants; st. 10 (stage 10 whole embryo), DEL (deep layer explant), SE (superficial layer explant).
itln2 (intelectin 2) gene expression in Xenopus laevis embryo, assayed via in situ hybridization, NF stage 14, dorsal view, anteriorleft.
irf6 (interferon regulatory factor 6) gene expression in Xenopus laevis embryos, assayed via in situ hybridization, NF stage 10.5, vegetal/blastoporal view, dorsal up (left) and animal pole view ( right).
irf6 (interferon regulatory factor 6) gene expression in Xenopus laevis embryos, assayed via in situ hybridization, NF stage 11, vegetal/blastoporal view.
irf6 (interferon regulatory factor 6) gene expression in Xenopus laevis embryos, assayed via in situ hybridization, NF stage 12.5, dorso-posterior view, anterior up.
irf6 (interferon regulatory factor 6) gene expression in Xenopus laevis embryos, assayed via in situ hybridization, NF stage 18, posterior view, dorsal up.
irf6 (interferon regulatory factor 6) gene expression in bisected Xenopus laevis embryo, mid-sagittal section, assayed via in situ hybridization, NF stage 10.5, animal hemisphere up.
irf6 (interferon regulatory factor 6) gene expression in Xenopus laevis embryo, assayed via in situ hybridization, NF stage 27/28, lateral view, anteriorleft, dorsal up.
Fig. 6. Defects in development of superficialepithelium in irf6DBD-injected X. laevis embryos. (A) Animal and, (C) vegetal views of stage 11 uninjected controls (Un). (B) Animal and (D) vegetal views of stage 11 embryos injected with 2Â ng D. rerio irf6DBD mRNA. (E) Uninjected (Un) and (F) irf6DBD-injected stage 14 embryos. Arrow in (F) indicates lesion in the ectoderm. (G) RT-PCR analysis of SE markers, esr6e and grhl3 in control (Un) and irf6DBD-injected embryos and explants. St. 11, whole embryo stage 11; cap, stage 11 animal cap; DEL, deep layer explants; SE, superficial layer explants.
Fig. 7. Maternal Irf6 is required for SE specification in X. laevis. (A) Control (left side) and Irf6-depleted (right side) embryos at stage 12. Upper row is a vegetal view; lower row is an animal view. (B) Control (left embryo) and Irf6-depleted (right four embryos) embryos at stage 24. Arrow indicates epidermal lesion in an Irf6-depleted embryo. (C) Control uninjected embryos. (D) embryos injected with 24 ng irf6 MO at the 1–2 cell stage. (E) Quantitative RT-PCR of esr6e, grhl3, sox11, irf6, sox2 and xbra expression in uninjected embryos (Un) and Irf6-depleted embryos (irf6 MO) at the indicated stages. Graphs represent data from a single PCR run.
Fig. 8. Maternal Irf6 is required for SE differentiation in X. laevis. (A, B) Whole mount in situ hybridization of stage 14, control (A) uninjected (Un) and (B) Irf6-depleted (irf6 MO) embryos for intelectin 2 (intel2). Lateral view is shown, anterior is to the left. Inset shows close up views of the epidermis in a bisected embryo. (C, D) Sections of (C) uninjected and (D) Irf6-depleted stage 13 embryos stained with mAb LP3K. (E, F) Sections of, (E) control and, (F) Irf6-depleted stage 22 embryos stained with Azan. Dotted lines indicate the thickness of the ectoderm; arrows indicate the location of embryonic pigment.
Fig. 9. Specificity of Irf6 depletion in Xenopus. (A) Quantitative RT-PCR of esr6e and grhl3 expression in controls (Un), Irf6-depleted embryos (irf6 MO) and depleted embryos injected with 50 pg irf6.2 mRNA (MO + RNA), or (B) with 250 pg irf6.2 mRNA (MO + RNA). (C– E) Whole mount in situ hybridization for intelectin 2 at stage 14, (C) control uninjected (Un), (D) Irf6-depleted (irf6 MO) and (E) rescued embryos (irf6 MO + irf6 RNA).
itln2 (intelectin 2) gene expression in Xenopus laevis embryo, assayed via in situ hybridization, NF stage 14, lateral view, anteriorleft, and in section (inset)
Bailey,
Interferon regulatory factor 6 promotes cell cycle arrest and is regulated by the proteasome in a cell cycle-dependent manner.
2008, Pubmed
Bailey,
Interferon regulatory factor 6 promotes cell cycle arrest and is regulated by the proteasome in a cell cycle-dependent manner.
2008,
Pubmed Bailey,
Mammary serine protease inhibitor (Maspin) binds directly to interferon regulatory factor 6: identification of a novel serpin partnership.
2005,
Pubmed Ben,
Genomic, cDNA and embryonic expression analysis of zebrafish IRF6, the gene mutated in the human oral clefting disorders Van der Woude and popliteal pterygium syndromes.
2005,
Pubmed Betchaku,
Contact relations, surface activity, and cortical microfilaments of marginal cells of the enveloping layer and of the yolk syncytial and yolk cytoplasmic layers of fundulus before and during epiboly.
1978,
Pubmed Bouvet,
Enveloping layer and periderm of the trout embryo (Salmo trutta fario L.)20U.
1976,
Pubmed Braissant,
Differential expression of peroxisome proliferator-activated receptor-alpha, -beta, and -gamma during rat embryonic development.
1998,
Pubmed Bruce,
The maternally expressed zebrafish T-box gene eomesodermin regulates organizer formation.
2003,
Pubmed
,
Xenbase Butler,
Nonradioactive in situ hybridization to xenopus tissue sections.
2001,
Pubmed
,
Xenbase Chalmers,
Grainyhead-like 3, a transcription factor identified in a microarray screen, promotes the specification of the superficial layer of the embryonic epidermis.
2006,
Pubmed
,
Xenbase Chalmers,
Oriented cell divisions asymmetrically segregate aPKC and generate cell fate diversity in the early Xenopus embryo.
2003,
Pubmed
,
Xenbase Chalmers,
Intrinsic differences between the superficial and deep layers of the Xenopus ectoderm control primary neuronal differentiation.
2002,
Pubmed
,
Xenbase Chalmers,
aPKC, Crumbs3 and Lgl2 control apicobasal polarity in early vertebrate development.
2005,
Pubmed
,
Xenbase Choi,
Xenopus Cdc42 regulates convergent extension movements during gastrulation through Wnt/Ca2+ signaling pathway.
2002,
Pubmed
,
Xenbase Ciruna,
Production of maternal-zygotic mutant zebrafish by germ-line replacement.
2002,
Pubmed Cooper,
A cluster of noninvoluting endocytic cells at the margin of the zebrafish blastoderm marks the site of embryonic shield formation.
1996,
Pubmed Cornell,
Delta signaling mediates segregation of neural crest and spinal sensory neurons from zebrafish lateral neural plate.
2000,
Pubmed D'Amico,
Morphogenetic domains in the yolk syncytial layer of axiating zebrafish embryos.
2001,
Pubmed
,
Xenbase Deblandre,
A two-step mechanism generates the spacing pattern of the ciliated cells in the skin of Xenopus embryos.
1999,
Pubmed
,
Xenbase Detrich,
Intraembryonic hematopoietic cell migration during vertebrate development.
1995,
Pubmed Dollar,
Regulation of Lethal giant larvae by Dishevelled.
2005,
Pubmed
,
Xenbase Doyon,
Heritable targeted gene disruption in zebrafish using designed zinc-finger nucleases.
2008,
Pubmed Faure,
Xenopus p21-activated kinase 5 regulates blastomeres' adhesive properties during convergent extension movements.
2005,
Pubmed
,
Xenbase Godsave,
Intermediate filaments in the Xenopus oocyte: the appearance and distribution of cytokeratin-containing filaments.
1984,
Pubmed
,
Xenbase Guttormsen,
Disruption of epidermal specific gene expression and delayed skin development in AP-2 gamma mutant mice.
2008,
Pubmed Hartenstein,
Early neurogenesis in Xenopus: the spatio-temporal pattern of proliferation and cell lineages in the embryonic spinal cord.
1989,
Pubmed
,
Xenbase Hatada,
An interferon regulatory factor-related gene (xIRF-6) is expressed in the posterior mesoderm during the early development of Xenopus laevis.
1997,
Pubmed
,
Xenbase Hayes,
Identification of novel ciliogenesis factors using a new in vivo model for mucociliary epithelial development.
2007,
Pubmed
,
Xenbase Heasman,
Fertilization of cultured Xenopus oocytes and use in studies of maternally inherited molecules.
1991,
Pubmed
,
Xenbase Helfrich,
Role of aPKC isoforms and their binding partners Par3 and Par6 in epidermal barrier formation.
2007,
Pubmed Hens,
Regulation of Xenopus embryonic cell adhesion by the small GTPase, rac.
2002,
Pubmed
,
Xenbase Hensey,
A developmental timer that regulates apoptosis at the onset of gastrulation.
1997,
Pubmed
,
Xenbase Ho,
Cell movements and cell fate during zebrafish gastrulation.
1992,
Pubmed Houston,
Maternal Xenopus Zic2 negatively regulates Nodal-related gene expression during anteroposterior patterning.
2005,
Pubmed
,
Xenbase Houston,
A critical role for Xdazl, a germ plasm-localized RNA, in the differentiation of primordial germ cells in Xenopus.
2000,
Pubmed
,
Xenbase Hu,
Abnormal morphogenesis but intact IKK activation in mice lacking the IKKalpha subunit of IkappaB kinase.
1999,
Pubmed Ikegami,
Developmental activation of the capability to undergo checkpoint-induced apoptosis in the early zebrafish embryo.
1999,
Pubmed
,
Xenbase Imboden,
Cytokeratin 8 is a suitable epidermal marker during zebrafish development.
1997,
Pubmed Ingraham,
Abnormal skin, limb and craniofacial morphogenesis in mice deficient for interferon regulatory factor 6 (Irf6).
2006,
Pubmed Johnson,
Lineage allocation and cell polarity during mouse embryogenesis.
2004,
Pubmed Jones,
Development of the ectoderm in Xenopus: tissue specification and the role of cell association and division.
1986,
Pubmed
,
Xenbase Kaji,
dlx3b and dlx4b function in the development of Rohon-Beard sensory neurons and trigeminal placode in the zebrafish neurula.
2004,
Pubmed
,
Xenbase Kane,
Mutations in half baked/E-cadherin block cell behaviors that are necessary for teleost epiboly.
2005,
Pubmed Kane,
The zebrafish epiboly mutants.
1996,
Pubmed Keller,
Rearrangement of enveloping layer cells without disruption of the epithelial permeability barrier as a factor in Fundulus epiboly.
1987,
Pubmed Keller,
The cellular basis of epiboly: an SEM study of deep-cell rearrangement during gastrulation in Xenopus laevis.
1980,
Pubmed
,
Xenbase Kimmel,
The shaping of pharyngeal cartilages during early development of the zebrafish.
1998,
Pubmed Kimmel,
Origin and organization of the zebrafish fate map.
1990,
Pubmed Kimmel,
Stages of embryonic development of the zebrafish.
1995,
Pubmed
,
Xenbase Klein,
Genetic and genomic tools for Xenopus research: The NIH Xenopus initiative.
2002,
Pubmed
,
Xenbase Knight,
Developmental expression analysis of the mouse and chick orthologues of IRF6: the gene mutated in Van der Woude syndrome.
2006,
Pubmed Kondo,
Mutations in IRF6 cause Van der Woude and popliteal pterygium syndromes.
2002,
Pubmed Köppen,
Coordinated cell-shape changes control epithelial movement in zebrafish and Drosophila.
2006,
Pubmed Lane,
Keratin antigens in differentiating skin.
1985,
Pubmed Li,
Redundant activities of Tfap2a and Tfap2c are required for neural crest induction and development of other non-neural ectoderm derivatives in zebrafish embryos.
2007,
Pubmed Li,
IKK1-deficient mice exhibit abnormal development of skin and skeleton.
1999,
Pubmed Li,
Identification of 14-3-3sigma mutation causing cutaneous abnormality in repeated-epilation mutant mouse.
2005,
Pubmed Lyons,
Molecular cloning, genetic mapping, and expression analysis of four zebrafish c/ebp genes.
2001,
Pubmed Mathavan,
Transcriptome analysis of zebrafish embryogenesis using microarrays.
2005,
Pubmed McFarland,
Genetic locus half baked is necessary for morphogenesis of the ectoderm.
2005,
Pubmed Meng,
Targeted gene inactivation in zebrafish using engineered zinc-finger nucleases.
2008,
Pubmed Nagafuchi,
Molecular architecture of adherens junctions.
2001,
Pubmed Nakaya,
Meiotic maturation induces animal-vegetal asymmetric distribution of aPKC and ASIP/PAR-3 in Xenopus oocytes.
2000,
Pubmed
,
Xenbase Nasevicius,
Effective targeted gene 'knockdown' in zebrafish.
2000,
Pubmed O'Brien,
Transcription factor Ap-2alpha is necessary for development of embryonic melanophores, autonomic neurons and pharyngeal skeleton in zebrafish.
2004,
Pubmed Ossipova,
PAR1 specifies ciliated cells in vertebrate ectoderm downstream of aPKC.
2007,
Pubmed
,
Xenbase Oteíza,
Origin and shaping of the laterality organ in zebrafish.
2008,
Pubmed Paun,
The IRF family, revisited.
2007,
Pubmed Pei,
An early requirement for maternal FoxH1 during zebrafish gastrulation.
2007,
Pubmed Postlethwait,
Vertebrate genome evolution and the zebrafish gene map.
1998,
Pubmed Richardson,
Irf6 is a key determinant of the keratinocyte proliferation-differentiation switch.
2006,
Pubmed Sagerström,
Specification of the enveloping layer and lack of autoneuralization in zebrafish embryonic explants.
2005,
Pubmed
,
Xenbase Schulte-Merker,
no tail (ntl) is the zebrafish homologue of the mouse T (Brachyury) gene.
1994,
Pubmed Solnica-Krezel,
Microtubule arrays of the zebrafish yolk cell: organization and function during epiboly.
1994,
Pubmed Solnica-Krezel,
Gastrulation in zebrafish -- all just about adhesion?
2006,
Pubmed Strähle,
Ultraviolet irradiation impairs epiboly in zebrafish embryos: evidence for a microtubule-dependent mechanism of epiboly.
1993,
Pubmed Suzuki,
The PAR-aPKC system: lessons in polarity.
2006,
Pubmed Taniguchi,
IRF family of transcription factors as regulators of host defense.
2001,
Pubmed Torpey,
Function of maternal cytokeratin in Xenopus development.
1992,
Pubmed
,
Xenbase Truong,
Control of keratinocyte proliferation and differentiation by p63.
2007,
Pubmed Uribe,
A review of actin binding proteins: new perspectives.
2009,
Pubmed Wagner,
Maternal control of development at the midblastula transition and beyond: mutants from the zebrafish II.
2004,
Pubmed Wang,
AP-2alpha: a regulator of EGF receptor signaling and proliferation in skin epidermis.
2006,
Pubmed Warga,
Cell movements during epiboly and gastrulation in zebrafish.
1990,
Pubmed Wünnenberg-Stapleton,
Involvement of the small GTPases XRhoA and XRnd1 in cell adhesion and head formation in early Xenopus development.
1999,
Pubmed
,
Xenbase Zalik,
Cell adhesion and the actin cytoskeleton of the enveloping layer in the zebrafish embryo during epiboly.
1999,
Pubmed Zucchero,
Interferon regulatory factor 6 (IRF6) gene variants and the risk of isolated cleft lip or palate.
2004,
Pubmed
