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Neural stem and progenitor cells shorten S-phase on commitment to neuron production.

Arai Y, Pulvers JN, Haffner C, Schilling B, Nüsslein I, Calegari F, Huttner WB - Nat Commun (2011)

Bottom Line: We found that G1 lengthening was associated with the transition from stem cell-like apical progenitors to fate-restricted basal (intermediate) progenitors.Comparative genome-wide gene expression analysis of expanding versus committed progenitor cells revealed changes in key factors of cell-cycle regulation, DNA replication and repair and chromatin remodelling.Our findings suggest that expanding neural stem and progenitor cells invest more time during S-phase into quality control of replicated DNA than those committed to neuron production.

View Article: PubMed Central - PubMed

Affiliation: Max Planck Institute of Molecular Cell Biology and Genetics, Pfotenhauerstrasse 108, 01307 Dresden, Germany.

ABSTRACT
During mammalian cerebral cortex development, the G1-phase of the cell cycle is known to lengthen, but it has been unclear which neural stem and progenitor cells are affected. In this paper, we develop a novel approach to determine cell-cycle parameters in specific classes of neural stem and progenitor cells, identified by molecular markers rather than location. We found that G1 lengthening was associated with the transition from stem cell-like apical progenitors to fate-restricted basal (intermediate) progenitors. Unexpectedly, expanding apical and basal progenitors exhibit a substantially longer S-phase than apical and basal progenitors committed to neuron production. Comparative genome-wide gene expression analysis of expanding versus committed progenitor cells revealed changes in key factors of cell-cycle regulation, DNA replication and repair and chromatin remodelling. Our findings suggest that expanding neural stem and progenitor cells invest more time during S-phase into quality control of replicated DNA than those committed to neuron production.

No MeSH data available.


Related in: MedlinePlus

Classification of proliferative and neurogenic APs and BPs on the basis of differential marker expression.(a) Pax6 (red), Tbr2 (blue), Tis21-GFP (green) and DAPI (white) staining. Red arrowheads, Pax6+/Tbr2−/Tis21-GFP− nuclei (APs); yellow arrowheads, Pax6+/Tbr2+/Tis21-GFP+ nuclei (BPs); blue arrowheads, Pax6−/Tbr2+/Tis21-GFP+ nucleus (BP). Scale bar, 50 μm. (b) Quantification of Tbr2+/Pax6+ (blue–red) and Tbr2−/Pax6+ (red) nuclei in VZ, SVZ and VZ+SVZ, each expressed as a percentage of total Pax6+ nuclei. (c) Quantification of the percentage of Tis21-GFP+ (green) and Tis21-GFP− (white) Tbr2−/Pax6+ nuclei in VZ+SVZ. (d, e) Pax6 (red), Tbr1 (d) or Tuj1 (e) (green) and DAPI (white) staining. Scale bar, 50 μm. (f) Tbr2 (red), Tbr1 (blue), Tis21-GFP (green) and DAPI (white) staining. Red arrowheads, Tbr2+/Tbr1−/Tis21-GFP+ nucleus (BP); yellow arrowheads, Tbr2+/Tbr1+/Tis21-GFP+ nucleus (neuron); blue arrowheads, Tbr2−/Tbr1+/Tis21-GFP+ nucleus (neuron). Scale bar, 50 μm. (g) Quantification of Tbr1+/Tbr2+ (blue–red) and Tbr1−/Tbr2+ (red) nuclei in VZ, SVZ and VZ+SVZ, each expressed as a percentage of total Tbr2+ nuclei. (h) Quantification of the percentage of Tis21-GFP+ (green) and Tis21-GFP− (white) Tbr1−/Tbr2+ nuclei in VZ+SVZ. (a, d, e, f) White lines at margins, VZ and SVZ boundaries. (b, c, g, h) Data are the mean of three 225-μm-wide fields, each from a different brain and litter; error bars indicate s.e.m. Images in (a, d, f) and data in (b, c, g, h) are from embryos subjected to cumulative EdU labelling for 5 h (a, d), 1 h (f) and 5, 9 and 12 h (b, c, g, h; compare Supplementary Fig. S4).
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f1: Classification of proliferative and neurogenic APs and BPs on the basis of differential marker expression.(a) Pax6 (red), Tbr2 (blue), Tis21-GFP (green) and DAPI (white) staining. Red arrowheads, Pax6+/Tbr2−/Tis21-GFP− nuclei (APs); yellow arrowheads, Pax6+/Tbr2+/Tis21-GFP+ nuclei (BPs); blue arrowheads, Pax6−/Tbr2+/Tis21-GFP+ nucleus (BP). Scale bar, 50 μm. (b) Quantification of Tbr2+/Pax6+ (blue–red) and Tbr2−/Pax6+ (red) nuclei in VZ, SVZ and VZ+SVZ, each expressed as a percentage of total Pax6+ nuclei. (c) Quantification of the percentage of Tis21-GFP+ (green) and Tis21-GFP− (white) Tbr2−/Pax6+ nuclei in VZ+SVZ. (d, e) Pax6 (red), Tbr1 (d) or Tuj1 (e) (green) and DAPI (white) staining. Scale bar, 50 μm. (f) Tbr2 (red), Tbr1 (blue), Tis21-GFP (green) and DAPI (white) staining. Red arrowheads, Tbr2+/Tbr1−/Tis21-GFP+ nucleus (BP); yellow arrowheads, Tbr2+/Tbr1+/Tis21-GFP+ nucleus (neuron); blue arrowheads, Tbr2−/Tbr1+/Tis21-GFP+ nucleus (neuron). Scale bar, 50 μm. (g) Quantification of Tbr1+/Tbr2+ (blue–red) and Tbr1−/Tbr2+ (red) nuclei in VZ, SVZ and VZ+SVZ, each expressed as a percentage of total Tbr2+ nuclei. (h) Quantification of the percentage of Tis21-GFP+ (green) and Tis21-GFP− (white) Tbr1−/Tbr2+ nuclei in VZ+SVZ. (a, d, e, f) White lines at margins, VZ and SVZ boundaries. (b, c, g, h) Data are the mean of three 225-μm-wide fields, each from a different brain and litter; error bars indicate s.e.m. Images in (a, d, f) and data in (b, c, g, h) are from embryos subjected to cumulative EdU labelling for 5 h (a, d), 1 h (f) and 5, 9 and 12 h (b, c, g, h; compare Supplementary Fig. S4).

Mentions: To determine the extent of intermingling of AP and BP interphase nuclei in the VZ and SVZ, and the proportion of APs committed to the neurogenic lineage, we performed triple immunofluorescence for Pax6, Tbr2 and Tis21-GFP, which is specifically expressed in the sub-population of APs that generate neurons or neurogenic BPs69 (Fig. 1a). With regard to APs, virtually all interphase nuclei in the VZ were Pax6+, as reported previously273031. Counter immunofluorescence for Tbr2 revealed that ≈30% of Pax6+ interphase nuclei in the VZ were Tbr2+ (Fig. 1b), indicating that APs contributed ≈70% and newborn BPs ≈30% to VZ interphase nuclei (Supplementary Fig. S2).


Neural stem and progenitor cells shorten S-phase on commitment to neuron production.

Arai Y, Pulvers JN, Haffner C, Schilling B, Nüsslein I, Calegari F, Huttner WB - Nat Commun (2011)

Classification of proliferative and neurogenic APs and BPs on the basis of differential marker expression.(a) Pax6 (red), Tbr2 (blue), Tis21-GFP (green) and DAPI (white) staining. Red arrowheads, Pax6+/Tbr2−/Tis21-GFP− nuclei (APs); yellow arrowheads, Pax6+/Tbr2+/Tis21-GFP+ nuclei (BPs); blue arrowheads, Pax6−/Tbr2+/Tis21-GFP+ nucleus (BP). Scale bar, 50 μm. (b) Quantification of Tbr2+/Pax6+ (blue–red) and Tbr2−/Pax6+ (red) nuclei in VZ, SVZ and VZ+SVZ, each expressed as a percentage of total Pax6+ nuclei. (c) Quantification of the percentage of Tis21-GFP+ (green) and Tis21-GFP− (white) Tbr2−/Pax6+ nuclei in VZ+SVZ. (d, e) Pax6 (red), Tbr1 (d) or Tuj1 (e) (green) and DAPI (white) staining. Scale bar, 50 μm. (f) Tbr2 (red), Tbr1 (blue), Tis21-GFP (green) and DAPI (white) staining. Red arrowheads, Tbr2+/Tbr1−/Tis21-GFP+ nucleus (BP); yellow arrowheads, Tbr2+/Tbr1+/Tis21-GFP+ nucleus (neuron); blue arrowheads, Tbr2−/Tbr1+/Tis21-GFP+ nucleus (neuron). Scale bar, 50 μm. (g) Quantification of Tbr1+/Tbr2+ (blue–red) and Tbr1−/Tbr2+ (red) nuclei in VZ, SVZ and VZ+SVZ, each expressed as a percentage of total Tbr2+ nuclei. (h) Quantification of the percentage of Tis21-GFP+ (green) and Tis21-GFP− (white) Tbr1−/Tbr2+ nuclei in VZ+SVZ. (a, d, e, f) White lines at margins, VZ and SVZ boundaries. (b, c, g, h) Data are the mean of three 225-μm-wide fields, each from a different brain and litter; error bars indicate s.e.m. Images in (a, d, f) and data in (b, c, g, h) are from embryos subjected to cumulative EdU labelling for 5 h (a, d), 1 h (f) and 5, 9 and 12 h (b, c, g, h; compare Supplementary Fig. S4).
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f1: Classification of proliferative and neurogenic APs and BPs on the basis of differential marker expression.(a) Pax6 (red), Tbr2 (blue), Tis21-GFP (green) and DAPI (white) staining. Red arrowheads, Pax6+/Tbr2−/Tis21-GFP− nuclei (APs); yellow arrowheads, Pax6+/Tbr2+/Tis21-GFP+ nuclei (BPs); blue arrowheads, Pax6−/Tbr2+/Tis21-GFP+ nucleus (BP). Scale bar, 50 μm. (b) Quantification of Tbr2+/Pax6+ (blue–red) and Tbr2−/Pax6+ (red) nuclei in VZ, SVZ and VZ+SVZ, each expressed as a percentage of total Pax6+ nuclei. (c) Quantification of the percentage of Tis21-GFP+ (green) and Tis21-GFP− (white) Tbr2−/Pax6+ nuclei in VZ+SVZ. (d, e) Pax6 (red), Tbr1 (d) or Tuj1 (e) (green) and DAPI (white) staining. Scale bar, 50 μm. (f) Tbr2 (red), Tbr1 (blue), Tis21-GFP (green) and DAPI (white) staining. Red arrowheads, Tbr2+/Tbr1−/Tis21-GFP+ nucleus (BP); yellow arrowheads, Tbr2+/Tbr1+/Tis21-GFP+ nucleus (neuron); blue arrowheads, Tbr2−/Tbr1+/Tis21-GFP+ nucleus (neuron). Scale bar, 50 μm. (g) Quantification of Tbr1+/Tbr2+ (blue–red) and Tbr1−/Tbr2+ (red) nuclei in VZ, SVZ and VZ+SVZ, each expressed as a percentage of total Tbr2+ nuclei. (h) Quantification of the percentage of Tis21-GFP+ (green) and Tis21-GFP− (white) Tbr1−/Tbr2+ nuclei in VZ+SVZ. (a, d, e, f) White lines at margins, VZ and SVZ boundaries. (b, c, g, h) Data are the mean of three 225-μm-wide fields, each from a different brain and litter; error bars indicate s.e.m. Images in (a, d, f) and data in (b, c, g, h) are from embryos subjected to cumulative EdU labelling for 5 h (a, d), 1 h (f) and 5, 9 and 12 h (b, c, g, h; compare Supplementary Fig. S4).
Mentions: To determine the extent of intermingling of AP and BP interphase nuclei in the VZ and SVZ, and the proportion of APs committed to the neurogenic lineage, we performed triple immunofluorescence for Pax6, Tbr2 and Tis21-GFP, which is specifically expressed in the sub-population of APs that generate neurons or neurogenic BPs69 (Fig. 1a). With regard to APs, virtually all interphase nuclei in the VZ were Pax6+, as reported previously273031. Counter immunofluorescence for Tbr2 revealed that ≈30% of Pax6+ interphase nuclei in the VZ were Tbr2+ (Fig. 1b), indicating that APs contributed ≈70% and newborn BPs ≈30% to VZ interphase nuclei (Supplementary Fig. S2).

Bottom Line: We found that G1 lengthening was associated with the transition from stem cell-like apical progenitors to fate-restricted basal (intermediate) progenitors.Comparative genome-wide gene expression analysis of expanding versus committed progenitor cells revealed changes in key factors of cell-cycle regulation, DNA replication and repair and chromatin remodelling.Our findings suggest that expanding neural stem and progenitor cells invest more time during S-phase into quality control of replicated DNA than those committed to neuron production.

View Article: PubMed Central - PubMed

Affiliation: Max Planck Institute of Molecular Cell Biology and Genetics, Pfotenhauerstrasse 108, 01307 Dresden, Germany.

ABSTRACT
During mammalian cerebral cortex development, the G1-phase of the cell cycle is known to lengthen, but it has been unclear which neural stem and progenitor cells are affected. In this paper, we develop a novel approach to determine cell-cycle parameters in specific classes of neural stem and progenitor cells, identified by molecular markers rather than location. We found that G1 lengthening was associated with the transition from stem cell-like apical progenitors to fate-restricted basal (intermediate) progenitors. Unexpectedly, expanding apical and basal progenitors exhibit a substantially longer S-phase than apical and basal progenitors committed to neuron production. Comparative genome-wide gene expression analysis of expanding versus committed progenitor cells revealed changes in key factors of cell-cycle regulation, DNA replication and repair and chromatin remodelling. Our findings suggest that expanding neural stem and progenitor cells invest more time during S-phase into quality control of replicated DNA than those committed to neuron production.

No MeSH data available.


Related in: MedlinePlus