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Two Notch ligands, Dll1 and Jag1, are differently restricted in their range of action to control neurogenesis in the mammalian spinal cord.

Ramos C, Rocha S, Gaspar C, Henrique D - PLoS ONE (2010)

Bottom Line: In the presence of a functional Dll1 allele, V1 neurogenesis is restored to the levels detected in single Jag1 mutants, while dI6 neurogenesis returns to normal, thereby confirming that Dll1-mediated signalling compensates for Jag1 deletion in V1 and dI6 domains.Our results reveal that Dll1 and Jag1 are functionally equivalent in controlling the rate of neurogenesis within their expression domains.However, Jag1 can only activate Notch signalling within the V1 and dI6 domains, whereas Dll1 can signal to neural progenitors both inside and outside its domains of expression.

View Article: PubMed Central - PubMed

Affiliation: Faculdade de Medicina de Lisboa, Instituto de Medicina Molecular, Lisboa, Portugal.

ABSTRACT

Background: Notch signalling regulates neuronal differentiation in the vertebrate nervous system. In addition to a widespread function in maintaining neural progenitors, Notch signalling has also been involved in specific neuronal fate decisions. These functions are likely mediated by distinct Notch ligands, which show restricted expression patterns in the developing nervous system. Two ligands, in particular, are expressed in non-overlapping complementary domains of the embryonic spinal cord, with Jag1 being restricted to the V1 and dI6 progenitor domains, while Dll1 is expressed in the remaining domains. However, the specific contribution of different ligands to regulate neurogenesis in vertebrate embryos is still poorly understood.

Methodology/principal findings: In this work, we investigated the role of Jag1 and Dll1 during spinal cord neurogenesis, using conditional knockout mice where the two genes are deleted in the neuroepithelium, singly or in combination. Our analysis showed that Jag1 deletion leads to a modest increase in V1 interneurons, while dI6 neurogenesis was unaltered. This mild Jag1 phenotype contrasts with the strong neurogenic phenotype detected in Dll1 mutants and led us to hypothesize that neighbouring Dll1-expressing cells signal to V1 and dI6 progenitors and restore neurogenesis in the absence of Jag1. Analysis of double Dll1;Jag1 mutant embryos revealed a stronger increase in V1-derived interneurons and overproduction of dI6 interneurons. In the presence of a functional Dll1 allele, V1 neurogenesis is restored to the levels detected in single Jag1 mutants, while dI6 neurogenesis returns to normal, thereby confirming that Dll1-mediated signalling compensates for Jag1 deletion in V1 and dI6 domains.

Conclusions/significance: Our results reveal that Dll1 and Jag1 are functionally equivalent in controlling the rate of neurogenesis within their expression domains. However, Jag1 can only activate Notch signalling within the V1 and dI6 domains, whereas Dll1 can signal to neural progenitors both inside and outside its domains of expression.

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Nestin-Cre driver efficiently inactivates Jag1 in V1 and dI6 domains.(A, B) Immunofluorescence in Jag1f/f;Rosa26YFP/+;NesCre E11.5 embryos shows widespread YFP expression, indicating that most of the cells had undergone Cre-mediated recombination. Since YFP signal fades away during longer fixations, IF with an anti-GFP antibody was used to detect YFP expression. (A′, B′) The two characteristic stripes of Jag1-protein expression are absent in Jag1f/f;Rosa26YFP/+;NesCre embryos. (A″, B″) Merge of YFP and Jag1 expression. Scale bar 50 µm. Strong red signal in B′, B″ is due to erythrocytes.
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pone-0015515-g004: Nestin-Cre driver efficiently inactivates Jag1 in V1 and dI6 domains.(A, B) Immunofluorescence in Jag1f/f;Rosa26YFP/+;NesCre E11.5 embryos shows widespread YFP expression, indicating that most of the cells had undergone Cre-mediated recombination. Since YFP signal fades away during longer fixations, IF with an anti-GFP antibody was used to detect YFP expression. (A′, B′) The two characteristic stripes of Jag1-protein expression are absent in Jag1f/f;Rosa26YFP/+;NesCre embryos. (A″, B″) Merge of YFP and Jag1 expression. Scale bar 50 µm. Strong red signal in B′, B″ is due to erythrocytes.

Mentions: To exclude that the mild neurogenic phenotype found in Jag1f/f;NesCre embryos was due to poor Cre recombinase activity driven by the Nestin-Cre driver, we evaluated the extent of Nestin Cre-mediated recombination in the embryonic spinal cord of Jag1 mutants. To assess this, a Rosa26-derived reporter line that conditionally expresses the YFP gene (Rosa26-YFP) was bred into the Jag1f/f;NesCre line, allowing us to identify cells where Cre-mediated recombination is active [19]. E11.5 Jag1f/f;R26-YFP/+;NesCre embryos were collected and exhibited an intense YFP immunofluorescence along the whole DV axis of the developing spinal cord, indicating widespread Cre-mediated recombination in the neuroepithelium (Fig. 4 A,B and A″,B″). In addition, we have used immunofluorescence to detect the presence of the Jag1 protein in control and Jag1 mutant embryos. Our results show that Jag1 is completely absent from the dI6 and V1 domains of Jag1 mutants, demonstrating that the Nestin-Cre driver effectively deletes Jag1 in the embryonic spinal cord (Fig. 4 A′,B′ and A″,B″).


Two Notch ligands, Dll1 and Jag1, are differently restricted in their range of action to control neurogenesis in the mammalian spinal cord.

Ramos C, Rocha S, Gaspar C, Henrique D - PLoS ONE (2010)

Nestin-Cre driver efficiently inactivates Jag1 in V1 and dI6 domains.(A, B) Immunofluorescence in Jag1f/f;Rosa26YFP/+;NesCre E11.5 embryos shows widespread YFP expression, indicating that most of the cells had undergone Cre-mediated recombination. Since YFP signal fades away during longer fixations, IF with an anti-GFP antibody was used to detect YFP expression. (A′, B′) The two characteristic stripes of Jag1-protein expression are absent in Jag1f/f;Rosa26YFP/+;NesCre embryos. (A″, B″) Merge of YFP and Jag1 expression. Scale bar 50 µm. Strong red signal in B′, B″ is due to erythrocytes.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2991363&req=5

pone-0015515-g004: Nestin-Cre driver efficiently inactivates Jag1 in V1 and dI6 domains.(A, B) Immunofluorescence in Jag1f/f;Rosa26YFP/+;NesCre E11.5 embryos shows widespread YFP expression, indicating that most of the cells had undergone Cre-mediated recombination. Since YFP signal fades away during longer fixations, IF with an anti-GFP antibody was used to detect YFP expression. (A′, B′) The two characteristic stripes of Jag1-protein expression are absent in Jag1f/f;Rosa26YFP/+;NesCre embryos. (A″, B″) Merge of YFP and Jag1 expression. Scale bar 50 µm. Strong red signal in B′, B″ is due to erythrocytes.
Mentions: To exclude that the mild neurogenic phenotype found in Jag1f/f;NesCre embryos was due to poor Cre recombinase activity driven by the Nestin-Cre driver, we evaluated the extent of Nestin Cre-mediated recombination in the embryonic spinal cord of Jag1 mutants. To assess this, a Rosa26-derived reporter line that conditionally expresses the YFP gene (Rosa26-YFP) was bred into the Jag1f/f;NesCre line, allowing us to identify cells where Cre-mediated recombination is active [19]. E11.5 Jag1f/f;R26-YFP/+;NesCre embryos were collected and exhibited an intense YFP immunofluorescence along the whole DV axis of the developing spinal cord, indicating widespread Cre-mediated recombination in the neuroepithelium (Fig. 4 A,B and A″,B″). In addition, we have used immunofluorescence to detect the presence of the Jag1 protein in control and Jag1 mutant embryos. Our results show that Jag1 is completely absent from the dI6 and V1 domains of Jag1 mutants, demonstrating that the Nestin-Cre driver effectively deletes Jag1 in the embryonic spinal cord (Fig. 4 A′,B′ and A″,B″).

Bottom Line: In the presence of a functional Dll1 allele, V1 neurogenesis is restored to the levels detected in single Jag1 mutants, while dI6 neurogenesis returns to normal, thereby confirming that Dll1-mediated signalling compensates for Jag1 deletion in V1 and dI6 domains.Our results reveal that Dll1 and Jag1 are functionally equivalent in controlling the rate of neurogenesis within their expression domains.However, Jag1 can only activate Notch signalling within the V1 and dI6 domains, whereas Dll1 can signal to neural progenitors both inside and outside its domains of expression.

View Article: PubMed Central - PubMed

Affiliation: Faculdade de Medicina de Lisboa, Instituto de Medicina Molecular, Lisboa, Portugal.

ABSTRACT

Background: Notch signalling regulates neuronal differentiation in the vertebrate nervous system. In addition to a widespread function in maintaining neural progenitors, Notch signalling has also been involved in specific neuronal fate decisions. These functions are likely mediated by distinct Notch ligands, which show restricted expression patterns in the developing nervous system. Two ligands, in particular, are expressed in non-overlapping complementary domains of the embryonic spinal cord, with Jag1 being restricted to the V1 and dI6 progenitor domains, while Dll1 is expressed in the remaining domains. However, the specific contribution of different ligands to regulate neurogenesis in vertebrate embryos is still poorly understood.

Methodology/principal findings: In this work, we investigated the role of Jag1 and Dll1 during spinal cord neurogenesis, using conditional knockout mice where the two genes are deleted in the neuroepithelium, singly or in combination. Our analysis showed that Jag1 deletion leads to a modest increase in V1 interneurons, while dI6 neurogenesis was unaltered. This mild Jag1 phenotype contrasts with the strong neurogenic phenotype detected in Dll1 mutants and led us to hypothesize that neighbouring Dll1-expressing cells signal to V1 and dI6 progenitors and restore neurogenesis in the absence of Jag1. Analysis of double Dll1;Jag1 mutant embryos revealed a stronger increase in V1-derived interneurons and overproduction of dI6 interneurons. In the presence of a functional Dll1 allele, V1 neurogenesis is restored to the levels detected in single Jag1 mutants, while dI6 neurogenesis returns to normal, thereby confirming that Dll1-mediated signalling compensates for Jag1 deletion in V1 and dI6 domains.

Conclusions/significance: Our results reveal that Dll1 and Jag1 are functionally equivalent in controlling the rate of neurogenesis within their expression domains. However, Jag1 can only activate Notch signalling within the V1 and dI6 domains, whereas Dll1 can signal to neural progenitors both inside and outside its domains of expression.

Show MeSH
Related in: MedlinePlus