Limits...
Slug is a direct Notch target required for initiation of cardiac cushion cellularization.

Niessen K, Fu Y, Chang L, Hoodless PA, McFadden D, Karsan A - J. Cell Biol. (2008)

Bottom Line: Slug deficiency results in impaired cellularization of the cardiac cushion at embryonic day (E)-9.5 but is compensated by increased Snail expression at E10.5, which restores cardiac cushion EMT.We further demonstrate that Slug, but not Snail, is directly up-regulated by Notch in endothelial cells and that Slug expression is required for Notch-mediated repression of the vascular endothelial cadherin promoter and for promoting migration of transformed endothelial cells.Collectively, our data suggest that combined expression of Slug and Snail is required for EMT in cardiac cushion morphogenesis.

View Article: PubMed Central - PubMed

Affiliation: Department of Medical Biophysics, British Columbia Cancer Agency, Vancouver V5Z 1L3, Canada.

ABSTRACT
Snail family proteins are key regulators of epithelial-mesenchymal transition, but their role in endothelial-to-mesenchymal transition (EMT) is less well studied. We show that Slug, a Snail family member, is expressed by a subset of endothelial cells as well as mesenchymal cells of the atrioventricular canal and outflow tract during cardiac cushion morphogenesis. Slug deficiency results in impaired cellularization of the cardiac cushion at embryonic day (E)-9.5 but is compensated by increased Snail expression at E10.5, which restores cardiac cushion EMT. We further demonstrate that Slug, but not Snail, is directly up-regulated by Notch in endothelial cells and that Slug expression is required for Notch-mediated repression of the vascular endothelial cadherin promoter and for promoting migration of transformed endothelial cells. In contrast, transforming growth factor beta (TGF-beta) induces Snail but not Slug. Interestingly, activation of Notch in the context of TGF-beta stimulation results in synergistic up-regulation of Snail in endothelial cells. Collectively, our data suggest that combined expression of Slug and Snail is required for EMT in cardiac cushion morphogenesis.

Show MeSH

Related in: MedlinePlus

Slug expression during cardiac cushion development. (A) β-galactosidase staining (representing Slug expression) of whole-mounted hearts from Slug-lacZ+/− embryos from E9.5 to 11.5. Arrows point to the AV canal or OFT at E11.5. Bars, 250 μm. (B) Sections through the AV canal and OFT of β-galactosidase–stained Slug-lacZ+/− hearts from E9.5 to 11.5. Bars, 100 μm. (C) Immunofluorescence staining for Slug (red) and CD31 (green) in E11.5 embryonic mouse hearts. Arrowheads point to cells coexpressing Slug and CD31. Bars, 25 μm. (D) In situ hybridization for Snail and Slug in a 65-d human embryonic heart. Arrows point to the mitral and tricuspid valves, arrowheads indicate the interatrial septum, and the asterisk marks the AV septum. A higher magnification image of the heart valve is shown in the right panel of each analysis. Bars, 1 mm.
© Copyright Policy
Related In: Results  -  Collection

License 1 - License 2
getmorefigures.php?uid=PMC2483533&req=5

fig2: Slug expression during cardiac cushion development. (A) β-galactosidase staining (representing Slug expression) of whole-mounted hearts from Slug-lacZ+/− embryos from E9.5 to 11.5. Arrows point to the AV canal or OFT at E11.5. Bars, 250 μm. (B) Sections through the AV canal and OFT of β-galactosidase–stained Slug-lacZ+/− hearts from E9.5 to 11.5. Bars, 100 μm. (C) Immunofluorescence staining for Slug (red) and CD31 (green) in E11.5 embryonic mouse hearts. Arrowheads point to cells coexpressing Slug and CD31. Bars, 25 μm. (D) In situ hybridization for Snail and Slug in a 65-d human embryonic heart. Arrows point to the mitral and tricuspid valves, arrowheads indicate the interatrial septum, and the asterisk marks the AV septum. A higher magnification image of the heart valve is shown in the right panel of each analysis. Bars, 1 mm.

Mentions: It has been reported that Slug mRNA is not expressed at E9.5 in the cardiac cushion (Timmerman et al., 2004), which suggests that it is dispensable for cardiac EMT. Because Notch activation has been shown to be critical for EMT during cardiac cushion development (Noseda et al., 2004; Timmerman et al., 2004), we were interested in defining the expression of Slug during the period of cellularization of the mammalian cardiac cushions. We thus examined Slug-lacZ mice, which have the lacZ gene inserted into the Slug locus with concomitant deletion of the zinc finger DNA binding motifs. Expression of lacZ in this model has been shown to faithfully match expression of Slug mRNA in all tissues analyzed, as determined by in situ hybridization (Jiang et al., 1998). β-galactosidase staining of E9.5–11.5 embryos showed that Slug-expressing cells were abundant in the heart (Fig. 2 A) with expression within a subset of endothelial cells as well as the mesenchymal cells of the AV canal and OFT at E9.5, with increasing expression over E10.5 and 11.5 (Fig. 2 B). Detailed analysis of Slug expression around E9.5 revealed that Slug expression is induced in the AV canal at the 25–28-somite stage at the initiation of EMT (Fig. S2, available at http://www.jcb.org/cgi/content/full/jcb.200710067/DC1). Immunofluorescent staining for Slug protein at E11.5 confirmed expression in the cardiac cushion mesenchyme and a subset of endothelial cells that costained for CD31 (Fig. 2 C, arrowheads).


Slug is a direct Notch target required for initiation of cardiac cushion cellularization.

Niessen K, Fu Y, Chang L, Hoodless PA, McFadden D, Karsan A - J. Cell Biol. (2008)

Slug expression during cardiac cushion development. (A) β-galactosidase staining (representing Slug expression) of whole-mounted hearts from Slug-lacZ+/− embryos from E9.5 to 11.5. Arrows point to the AV canal or OFT at E11.5. Bars, 250 μm. (B) Sections through the AV canal and OFT of β-galactosidase–stained Slug-lacZ+/− hearts from E9.5 to 11.5. Bars, 100 μm. (C) Immunofluorescence staining for Slug (red) and CD31 (green) in E11.5 embryonic mouse hearts. Arrowheads point to cells coexpressing Slug and CD31. Bars, 25 μm. (D) In situ hybridization for Snail and Slug in a 65-d human embryonic heart. Arrows point to the mitral and tricuspid valves, arrowheads indicate the interatrial septum, and the asterisk marks the AV septum. A higher magnification image of the heart valve is shown in the right panel of each analysis. Bars, 1 mm.
© Copyright Policy
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC2483533&req=5

fig2: Slug expression during cardiac cushion development. (A) β-galactosidase staining (representing Slug expression) of whole-mounted hearts from Slug-lacZ+/− embryos from E9.5 to 11.5. Arrows point to the AV canal or OFT at E11.5. Bars, 250 μm. (B) Sections through the AV canal and OFT of β-galactosidase–stained Slug-lacZ+/− hearts from E9.5 to 11.5. Bars, 100 μm. (C) Immunofluorescence staining for Slug (red) and CD31 (green) in E11.5 embryonic mouse hearts. Arrowheads point to cells coexpressing Slug and CD31. Bars, 25 μm. (D) In situ hybridization for Snail and Slug in a 65-d human embryonic heart. Arrows point to the mitral and tricuspid valves, arrowheads indicate the interatrial septum, and the asterisk marks the AV septum. A higher magnification image of the heart valve is shown in the right panel of each analysis. Bars, 1 mm.
Mentions: It has been reported that Slug mRNA is not expressed at E9.5 in the cardiac cushion (Timmerman et al., 2004), which suggests that it is dispensable for cardiac EMT. Because Notch activation has been shown to be critical for EMT during cardiac cushion development (Noseda et al., 2004; Timmerman et al., 2004), we were interested in defining the expression of Slug during the period of cellularization of the mammalian cardiac cushions. We thus examined Slug-lacZ mice, which have the lacZ gene inserted into the Slug locus with concomitant deletion of the zinc finger DNA binding motifs. Expression of lacZ in this model has been shown to faithfully match expression of Slug mRNA in all tissues analyzed, as determined by in situ hybridization (Jiang et al., 1998). β-galactosidase staining of E9.5–11.5 embryos showed that Slug-expressing cells were abundant in the heart (Fig. 2 A) with expression within a subset of endothelial cells as well as the mesenchymal cells of the AV canal and OFT at E9.5, with increasing expression over E10.5 and 11.5 (Fig. 2 B). Detailed analysis of Slug expression around E9.5 revealed that Slug expression is induced in the AV canal at the 25–28-somite stage at the initiation of EMT (Fig. S2, available at http://www.jcb.org/cgi/content/full/jcb.200710067/DC1). Immunofluorescent staining for Slug protein at E11.5 confirmed expression in the cardiac cushion mesenchyme and a subset of endothelial cells that costained for CD31 (Fig. 2 C, arrowheads).

Bottom Line: Slug deficiency results in impaired cellularization of the cardiac cushion at embryonic day (E)-9.5 but is compensated by increased Snail expression at E10.5, which restores cardiac cushion EMT.We further demonstrate that Slug, but not Snail, is directly up-regulated by Notch in endothelial cells and that Slug expression is required for Notch-mediated repression of the vascular endothelial cadherin promoter and for promoting migration of transformed endothelial cells.Collectively, our data suggest that combined expression of Slug and Snail is required for EMT in cardiac cushion morphogenesis.

View Article: PubMed Central - PubMed

Affiliation: Department of Medical Biophysics, British Columbia Cancer Agency, Vancouver V5Z 1L3, Canada.

ABSTRACT
Snail family proteins are key regulators of epithelial-mesenchymal transition, but their role in endothelial-to-mesenchymal transition (EMT) is less well studied. We show that Slug, a Snail family member, is expressed by a subset of endothelial cells as well as mesenchymal cells of the atrioventricular canal and outflow tract during cardiac cushion morphogenesis. Slug deficiency results in impaired cellularization of the cardiac cushion at embryonic day (E)-9.5 but is compensated by increased Snail expression at E10.5, which restores cardiac cushion EMT. We further demonstrate that Slug, but not Snail, is directly up-regulated by Notch in endothelial cells and that Slug expression is required for Notch-mediated repression of the vascular endothelial cadherin promoter and for promoting migration of transformed endothelial cells. In contrast, transforming growth factor beta (TGF-beta) induces Snail but not Slug. Interestingly, activation of Notch in the context of TGF-beta stimulation results in synergistic up-regulation of Snail in endothelial cells. Collectively, our data suggest that combined expression of Slug and Snail is required for EMT in cardiac cushion morphogenesis.

Show MeSH
Related in: MedlinePlus