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Attenuation of Notch and Hedgehog signaling is required for fate specification in the spinal cord.

Huang P, Xiong F, Megason SG, Schier AF - PLoS Genet. (2012)

Bottom Line: Hh signaling is required in LFP progenitors for KA″ fate specification, but prolonged Hh signaling interferes with KA″ differentiation.Notch signaling acts permissively to maintain LFP progenitor cells: activation of Notch signaling prevents differentiation, whereas inhibition of Notch signaling results in differentiation of ectopic KA″ cells.These results indicate that neural progenitors depend on Notch signaling to maintain Hh responsiveness and rely on Hh signaling to induce fate identity, whereas proper differentiation depends on the attenuation of both Notch and Hh signaling.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Cellular Biology, Center for Brain Science, Harvard Stem Cell Institute, Broad Institute, Center for Systems Biology, Harvard University, Cambridge, Massachusetts, USA. huang@mcb.harvard.edu

ABSTRACT
During the development of the spinal cord, proliferative neural progenitors differentiate into postmitotic neurons with distinct fates. How cells switch from progenitor states to differentiated fates is poorly understood. To address this question, we studied the differentiation of progenitors in the zebrafish spinal cord, focusing on the differentiation of Kolmer-Agduhr″ (KA″) interneurons from lateral floor plate (LFP) progenitors. In vivo cell tracking demonstrates that KA″ cells are generated from LFP progenitors by both symmetric and asymmetric cell divisions. A photoconvertible reporter of signaling history (PHRESH) reveals distinct temporal profiles of Hh response: LFP progenitors continuously respond to Hh, while KA″ cells lose Hh response upon differentiation. Hh signaling is required in LFP progenitors for KA″ fate specification, but prolonged Hh signaling interferes with KA″ differentiation. Notch signaling acts permissively to maintain LFP progenitor cells: activation of Notch signaling prevents differentiation, whereas inhibition of Notch signaling results in differentiation of ectopic KA″ cells. These results indicate that neural progenitors depend on Notch signaling to maintain Hh responsiveness and rely on Hh signaling to induce fate identity, whereas proper differentiation depends on the attenuation of both Notch and Hh signaling.

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Notch signaling in KA″ specification.(A) Wild type embryos at 18 hpf were double labeled with tal2 (red) and hes5, her12, her2, or her4 (green). tal2-expressing KA″ cells (arrows) lack expression of hes5, her12, her2, or her4. Note that hes5, her12, her2, and her4 are expressed in only a subset of LFP cells, which might reflect the oscillating nature of these genes in neural progenitor cells as previously reported [49]. Dorsal views are shown. (B) hsp-Gal4; UAS-NICD embryos and non-transgenic control embryos were heat-shocked at 10 hpf, and stained at 25 hpf for the expression of tal2 and nkx2.9 (top panel). Embryos were treated with DMSO or compound E at 13 hpf, and stained at 25 hpf for the expression of tal2 and nkx2.9 (bottom panel). tal2 and nkx2.9 staining is shown in lateral and dorsal views, respectively. Brackets indicate the extent of the LFP domain in lateral views, and arrows denote the two rows of LFP domains in dorsal views. (C) Quantification of KA″ specification under different conditions. KA″ cells were scored by the expression of tal2 at 25 hpf. Note that the fewer data points in compound E treatment at 13 hpf is due to the fact that most compound E-treated embryos had many tal2-positive cells in close clusters which prevents reliable scoring. Data shown therefore underestimate the total number of KA″ cells under this condition. (D) Ptc1-Kaede embryos were treated with DMSO or compound E at 14 hpf, photoconverted at 25 hpf, and stained with the GABA antibody (blue) at 37 hpf. Compound E treatment induced ectopic GABA-positive cells in the LFP domain (arrows), and abolished the expression of Ptc1-Kaedegreen (middle panel). Scale bars: 20 µm in A, D and 50 µm in B.
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pgen-1002762-g006: Notch signaling in KA″ specification.(A) Wild type embryos at 18 hpf were double labeled with tal2 (red) and hes5, her12, her2, or her4 (green). tal2-expressing KA″ cells (arrows) lack expression of hes5, her12, her2, or her4. Note that hes5, her12, her2, and her4 are expressed in only a subset of LFP cells, which might reflect the oscillating nature of these genes in neural progenitor cells as previously reported [49]. Dorsal views are shown. (B) hsp-Gal4; UAS-NICD embryos and non-transgenic control embryos were heat-shocked at 10 hpf, and stained at 25 hpf for the expression of tal2 and nkx2.9 (top panel). Embryos were treated with DMSO or compound E at 13 hpf, and stained at 25 hpf for the expression of tal2 and nkx2.9 (bottom panel). tal2 and nkx2.9 staining is shown in lateral and dorsal views, respectively. Brackets indicate the extent of the LFP domain in lateral views, and arrows denote the two rows of LFP domains in dorsal views. (C) Quantification of KA″ specification under different conditions. KA″ cells were scored by the expression of tal2 at 25 hpf. Note that the fewer data points in compound E treatment at 13 hpf is due to the fact that most compound E-treated embryos had many tal2-positive cells in close clusters which prevents reliable scoring. Data shown therefore underestimate the total number of KA″ cells under this condition. (D) Ptc1-Kaede embryos were treated with DMSO or compound E at 14 hpf, photoconverted at 25 hpf, and stained with the GABA antibody (blue) at 37 hpf. Compound E treatment induced ectopic GABA-positive cells in the LFP domain (arrows), and abolished the expression of Ptc1-Kaedegreen (middle panel). Scale bars: 20 µm in A, D and 50 µm in B.

Mentions: Notch signaling plays an important role in maintaining neural progenitor cells. We therefore hypothesized that active Notch signaling might not only maintain LFP progenitor state but also maintain Hh responsiveness. To test this model, we first analyzed endogenous Notch signaling activity by examining the expression pattern of known Notch target genes in the LFP domain, including hes5, her12, her2, and her4[32]. Strikingly, transcripts of all four genes were largely absent in tal2-expressing cells at 18 hpf (Figure 6A). This result suggests that Notch signaling activity is attenuated in KA″ cells.


Attenuation of Notch and Hedgehog signaling is required for fate specification in the spinal cord.

Huang P, Xiong F, Megason SG, Schier AF - PLoS Genet. (2012)

Notch signaling in KA″ specification.(A) Wild type embryos at 18 hpf were double labeled with tal2 (red) and hes5, her12, her2, or her4 (green). tal2-expressing KA″ cells (arrows) lack expression of hes5, her12, her2, or her4. Note that hes5, her12, her2, and her4 are expressed in only a subset of LFP cells, which might reflect the oscillating nature of these genes in neural progenitor cells as previously reported [49]. Dorsal views are shown. (B) hsp-Gal4; UAS-NICD embryos and non-transgenic control embryos were heat-shocked at 10 hpf, and stained at 25 hpf for the expression of tal2 and nkx2.9 (top panel). Embryos were treated with DMSO or compound E at 13 hpf, and stained at 25 hpf for the expression of tal2 and nkx2.9 (bottom panel). tal2 and nkx2.9 staining is shown in lateral and dorsal views, respectively. Brackets indicate the extent of the LFP domain in lateral views, and arrows denote the two rows of LFP domains in dorsal views. (C) Quantification of KA″ specification under different conditions. KA″ cells were scored by the expression of tal2 at 25 hpf. Note that the fewer data points in compound E treatment at 13 hpf is due to the fact that most compound E-treated embryos had many tal2-positive cells in close clusters which prevents reliable scoring. Data shown therefore underestimate the total number of KA″ cells under this condition. (D) Ptc1-Kaede embryos were treated with DMSO or compound E at 14 hpf, photoconverted at 25 hpf, and stained with the GABA antibody (blue) at 37 hpf. Compound E treatment induced ectopic GABA-positive cells in the LFP domain (arrows), and abolished the expression of Ptc1-Kaedegreen (middle panel). Scale bars: 20 µm in A, D and 50 µm in B.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC3369957&req=5

pgen-1002762-g006: Notch signaling in KA″ specification.(A) Wild type embryos at 18 hpf were double labeled with tal2 (red) and hes5, her12, her2, or her4 (green). tal2-expressing KA″ cells (arrows) lack expression of hes5, her12, her2, or her4. Note that hes5, her12, her2, and her4 are expressed in only a subset of LFP cells, which might reflect the oscillating nature of these genes in neural progenitor cells as previously reported [49]. Dorsal views are shown. (B) hsp-Gal4; UAS-NICD embryos and non-transgenic control embryos were heat-shocked at 10 hpf, and stained at 25 hpf for the expression of tal2 and nkx2.9 (top panel). Embryos were treated with DMSO or compound E at 13 hpf, and stained at 25 hpf for the expression of tal2 and nkx2.9 (bottom panel). tal2 and nkx2.9 staining is shown in lateral and dorsal views, respectively. Brackets indicate the extent of the LFP domain in lateral views, and arrows denote the two rows of LFP domains in dorsal views. (C) Quantification of KA″ specification under different conditions. KA″ cells were scored by the expression of tal2 at 25 hpf. Note that the fewer data points in compound E treatment at 13 hpf is due to the fact that most compound E-treated embryos had many tal2-positive cells in close clusters which prevents reliable scoring. Data shown therefore underestimate the total number of KA″ cells under this condition. (D) Ptc1-Kaede embryos were treated with DMSO or compound E at 14 hpf, photoconverted at 25 hpf, and stained with the GABA antibody (blue) at 37 hpf. Compound E treatment induced ectopic GABA-positive cells in the LFP domain (arrows), and abolished the expression of Ptc1-Kaedegreen (middle panel). Scale bars: 20 µm in A, D and 50 µm in B.
Mentions: Notch signaling plays an important role in maintaining neural progenitor cells. We therefore hypothesized that active Notch signaling might not only maintain LFP progenitor state but also maintain Hh responsiveness. To test this model, we first analyzed endogenous Notch signaling activity by examining the expression pattern of known Notch target genes in the LFP domain, including hes5, her12, her2, and her4[32]. Strikingly, transcripts of all four genes were largely absent in tal2-expressing cells at 18 hpf (Figure 6A). This result suggests that Notch signaling activity is attenuated in KA″ cells.

Bottom Line: Hh signaling is required in LFP progenitors for KA″ fate specification, but prolonged Hh signaling interferes with KA″ differentiation.Notch signaling acts permissively to maintain LFP progenitor cells: activation of Notch signaling prevents differentiation, whereas inhibition of Notch signaling results in differentiation of ectopic KA″ cells.These results indicate that neural progenitors depend on Notch signaling to maintain Hh responsiveness and rely on Hh signaling to induce fate identity, whereas proper differentiation depends on the attenuation of both Notch and Hh signaling.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Cellular Biology, Center for Brain Science, Harvard Stem Cell Institute, Broad Institute, Center for Systems Biology, Harvard University, Cambridge, Massachusetts, USA. huang@mcb.harvard.edu

ABSTRACT
During the development of the spinal cord, proliferative neural progenitors differentiate into postmitotic neurons with distinct fates. How cells switch from progenitor states to differentiated fates is poorly understood. To address this question, we studied the differentiation of progenitors in the zebrafish spinal cord, focusing on the differentiation of Kolmer-Agduhr″ (KA″) interneurons from lateral floor plate (LFP) progenitors. In vivo cell tracking demonstrates that KA″ cells are generated from LFP progenitors by both symmetric and asymmetric cell divisions. A photoconvertible reporter of signaling history (PHRESH) reveals distinct temporal profiles of Hh response: LFP progenitors continuously respond to Hh, while KA″ cells lose Hh response upon differentiation. Hh signaling is required in LFP progenitors for KA″ fate specification, but prolonged Hh signaling interferes with KA″ differentiation. Notch signaling acts permissively to maintain LFP progenitor cells: activation of Notch signaling prevents differentiation, whereas inhibition of Notch signaling results in differentiation of ectopic KA″ cells. These results indicate that neural progenitors depend on Notch signaling to maintain Hh responsiveness and rely on Hh signaling to induce fate identity, whereas proper differentiation depends on the attenuation of both Notch and Hh signaling.

Show MeSH
Related in: MedlinePlus