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Regulation of interkinetic nuclear migration by cell cycle-coupled active and passive mechanisms in the developing brain.

Kosodo Y, Suetsugu T, Suda M, Mimori-Kiyosue Y, Toida K, Baba SA, Kimura A, Matsuzaki F - EMBO J. (2011)

Bottom Line: Here, we show that INM proceeds through the cell cycle-dependent linkage of cell-autonomous and non-autonomous mechanisms.In contrast, in vivo observations of implanted microbeads, acute S-phase arrest of surrounding cells and computational modelling suggest that the basal migration of G1-phase nuclei depends on a displacement effect by G2-phase nuclei migrating apically.Our model for INM explains how the dynamics of neural progenitors harmonize their extensive proliferation with the epithelial architecture in the developing brain.

View Article: PubMed Central - PubMed

Affiliation: Laboratory for Cell Asymmetry, RIKEN Center for Developmental Biology, Kobe, Japan. kosodo@med.kawasaki-m.ac.jp

ABSTRACT
A hallmark of neurogenesis in the vertebrate brain is the apical-basal nuclear oscillation in polarized neural progenitor cells. Known as interkinetic nuclear migration (INM), these movements are synchronized with the cell cycle such that nuclei move basally during G1-phase and apically during G2-phase. However, it is unknown how the direction of movement and the cell cycle are tightly coupled. Here, we show that INM proceeds through the cell cycle-dependent linkage of cell-autonomous and non-autonomous mechanisms. During S to G2 progression, the microtubule-associated protein Tpx2 redistributes from the nucleus to the apical process, and promotes nuclear migration during G2-phase by altering microtubule organization. Thus, Tpx2 links cell-cycle progression and autonomous apical nuclear migration. In contrast, in vivo observations of implanted microbeads, acute S-phase arrest of surrounding cells and computational modelling suggest that the basal migration of G1-phase nuclei depends on a displacement effect by G2-phase nuclei migrating apically. Our model for INM explains how the dynamics of neural progenitors harmonize their extensive proliferation with the epithelial architecture in the developing brain.

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Related in: MedlinePlus

Arresting the cell cycle in G1-phase by introduction of p18Ink4c induces accumulation of nuclei in the basal region of the VZ. (A) Immunostaining of E14.5 mouse brain treated with p18Ink4c to arrest the cell cycle in G1-phase with a Ki67 antibody (proliferative marker, red). (a) Contralateral side. (b, b′) E13.5 mouse lateral cortex in which p18Ink4c (b) and NLS-GFP (b′; green) were co-electroporated and incubated for 24 h. Bar=50 μm. (B) Expression of GFP (green) with electroporation of either control vector (a) or p18Ink4c (b, b′) into E10.5 mouse telencephalon followed by 18 h of whole-embryo culture. Immunostaining using a p18Ink4c antibody was performed (b′; red). Bar=10 μm. (C) Positions of NLS-GFP-expressing nuclei with control vector (a) or p18Ink4c (b) introduced by electroporation into E13.5 mouse cortex followed by a 24-h incubation. Co-staining using a Tuj1 antibody (neuronal marker, red) and DAPI (DNA, blue) was performed. Bar=10 μm. (D) Positions of NLS-GFP-expressing nuclei relative to the apical surface with control vector (a) or p18Ink4c (b). Sums of numbers (N, x-coordinate) counted in 15 electroporated brain sections (nuclei from each section are indicated by white or dotted boxes) are shown according to their distance from the apical surface (y-coordinate). For (A–C), apical surface is down.
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f2: Arresting the cell cycle in G1-phase by introduction of p18Ink4c induces accumulation of nuclei in the basal region of the VZ. (A) Immunostaining of E14.5 mouse brain treated with p18Ink4c to arrest the cell cycle in G1-phase with a Ki67 antibody (proliferative marker, red). (a) Contralateral side. (b, b′) E13.5 mouse lateral cortex in which p18Ink4c (b) and NLS-GFP (b′; green) were co-electroporated and incubated for 24 h. Bar=50 μm. (B) Expression of GFP (green) with electroporation of either control vector (a) or p18Ink4c (b, b′) into E10.5 mouse telencephalon followed by 18 h of whole-embryo culture. Immunostaining using a p18Ink4c antibody was performed (b′; red). Bar=10 μm. (C) Positions of NLS-GFP-expressing nuclei with control vector (a) or p18Ink4c (b) introduced by electroporation into E13.5 mouse cortex followed by a 24-h incubation. Co-staining using a Tuj1 antibody (neuronal marker, red) and DAPI (DNA, blue) was performed. Bar=10 μm. (D) Positions of NLS-GFP-expressing nuclei relative to the apical surface with control vector (a) or p18Ink4c (b). Sums of numbers (N, x-coordinate) counted in 15 electroporated brain sections (nuclei from each section are indicated by white or dotted boxes) are shown according to their distance from the apical surface (y-coordinate). For (A–C), apical surface is down.

Mentions: Individual phases of INM are tightly correlated with phases of the cell cycle, but it has not been determined how migration depends on cell-cycle progression. To address this question, we first examined whether INM depends on G1- to S-phase progression. The cell cycle of neural progenitors was arrested at G1-phase by overexpression of p18Ink4c, a cyclin-dependent kinase inhibitor (Guan et al, 1994; Sherr and Roberts, 1999). Introduction of p18Ink4c by in utero electroporation resulted in a decrease in the number of cells expressing Ki67, a marker for the proliferative state (Figure 2A). The electroporated cells were neither labelled by BrdU, which is incorporated into DNA during S-phase (Supplementary Figure S1A), nor observed histologically to be in M-phase. These cells, therefore, had passed through M-phase and were arrested in G1-phase by the time of analysis (18 h after electroporation). Interestingly, at E10.5, when proliferative cells are dominant, the cell bodies of the p18Ink4c-electroporated cells accumulated in the basal region of the VZ, with their long apical processes extended toward the apical surface (Figure 2B). This phenomenon is not specific to this developmental stage, as statistical measurements showed basal accumulation of G1-arrested nuclei in the VZ at E14.5 as well (Figure 2C and D). The basal nuclear localization of p18Ink4c-expressing cells may be due to differentiation of G1-arrested progenitor cells into neurons that do not migrate to the apical surface. However, we confirmed that the progenitor state is not affected in p18Ink4c-expressing cells based on expression of Sox2 (Supplementary Figure S1B) and Pax6 (Supplementary Figure S1C), markers for apical neural progenitor cells (Götz et al, 1998; Graham et al, 2003). Furthermore, we did not observe any significant changes in the pattern of Tuj1 staining, a marker for neurons, nor any increase in expression of Tbr2, a marker for differentiating intermediate progenitor cells (Kowalczyk et al, 2009), 24 h after electroporation (Supplementary Figure S1D and E). These results indicate that the nuclei of neural progenitor cells do not migrate in the apical direction when they are arrested in G1-phase and suggest that entry into S-phase is a prerequisite for basal-to-apical nuclear migration.


Regulation of interkinetic nuclear migration by cell cycle-coupled active and passive mechanisms in the developing brain.

Kosodo Y, Suetsugu T, Suda M, Mimori-Kiyosue Y, Toida K, Baba SA, Kimura A, Matsuzaki F - EMBO J. (2011)

Arresting the cell cycle in G1-phase by introduction of p18Ink4c induces accumulation of nuclei in the basal region of the VZ. (A) Immunostaining of E14.5 mouse brain treated with p18Ink4c to arrest the cell cycle in G1-phase with a Ki67 antibody (proliferative marker, red). (a) Contralateral side. (b, b′) E13.5 mouse lateral cortex in which p18Ink4c (b) and NLS-GFP (b′; green) were co-electroporated and incubated for 24 h. Bar=50 μm. (B) Expression of GFP (green) with electroporation of either control vector (a) or p18Ink4c (b, b′) into E10.5 mouse telencephalon followed by 18 h of whole-embryo culture. Immunostaining using a p18Ink4c antibody was performed (b′; red). Bar=10 μm. (C) Positions of NLS-GFP-expressing nuclei with control vector (a) or p18Ink4c (b) introduced by electroporation into E13.5 mouse cortex followed by a 24-h incubation. Co-staining using a Tuj1 antibody (neuronal marker, red) and DAPI (DNA, blue) was performed. Bar=10 μm. (D) Positions of NLS-GFP-expressing nuclei relative to the apical surface with control vector (a) or p18Ink4c (b). Sums of numbers (N, x-coordinate) counted in 15 electroporated brain sections (nuclei from each section are indicated by white or dotted boxes) are shown according to their distance from the apical surface (y-coordinate). For (A–C), apical surface is down.
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f2: Arresting the cell cycle in G1-phase by introduction of p18Ink4c induces accumulation of nuclei in the basal region of the VZ. (A) Immunostaining of E14.5 mouse brain treated with p18Ink4c to arrest the cell cycle in G1-phase with a Ki67 antibody (proliferative marker, red). (a) Contralateral side. (b, b′) E13.5 mouse lateral cortex in which p18Ink4c (b) and NLS-GFP (b′; green) were co-electroporated and incubated for 24 h. Bar=50 μm. (B) Expression of GFP (green) with electroporation of either control vector (a) or p18Ink4c (b, b′) into E10.5 mouse telencephalon followed by 18 h of whole-embryo culture. Immunostaining using a p18Ink4c antibody was performed (b′; red). Bar=10 μm. (C) Positions of NLS-GFP-expressing nuclei with control vector (a) or p18Ink4c (b) introduced by electroporation into E13.5 mouse cortex followed by a 24-h incubation. Co-staining using a Tuj1 antibody (neuronal marker, red) and DAPI (DNA, blue) was performed. Bar=10 μm. (D) Positions of NLS-GFP-expressing nuclei relative to the apical surface with control vector (a) or p18Ink4c (b). Sums of numbers (N, x-coordinate) counted in 15 electroporated brain sections (nuclei from each section are indicated by white or dotted boxes) are shown according to their distance from the apical surface (y-coordinate). For (A–C), apical surface is down.
Mentions: Individual phases of INM are tightly correlated with phases of the cell cycle, but it has not been determined how migration depends on cell-cycle progression. To address this question, we first examined whether INM depends on G1- to S-phase progression. The cell cycle of neural progenitors was arrested at G1-phase by overexpression of p18Ink4c, a cyclin-dependent kinase inhibitor (Guan et al, 1994; Sherr and Roberts, 1999). Introduction of p18Ink4c by in utero electroporation resulted in a decrease in the number of cells expressing Ki67, a marker for the proliferative state (Figure 2A). The electroporated cells were neither labelled by BrdU, which is incorporated into DNA during S-phase (Supplementary Figure S1A), nor observed histologically to be in M-phase. These cells, therefore, had passed through M-phase and were arrested in G1-phase by the time of analysis (18 h after electroporation). Interestingly, at E10.5, when proliferative cells are dominant, the cell bodies of the p18Ink4c-electroporated cells accumulated in the basal region of the VZ, with their long apical processes extended toward the apical surface (Figure 2B). This phenomenon is not specific to this developmental stage, as statistical measurements showed basal accumulation of G1-arrested nuclei in the VZ at E14.5 as well (Figure 2C and D). The basal nuclear localization of p18Ink4c-expressing cells may be due to differentiation of G1-arrested progenitor cells into neurons that do not migrate to the apical surface. However, we confirmed that the progenitor state is not affected in p18Ink4c-expressing cells based on expression of Sox2 (Supplementary Figure S1B) and Pax6 (Supplementary Figure S1C), markers for apical neural progenitor cells (Götz et al, 1998; Graham et al, 2003). Furthermore, we did not observe any significant changes in the pattern of Tuj1 staining, a marker for neurons, nor any increase in expression of Tbr2, a marker for differentiating intermediate progenitor cells (Kowalczyk et al, 2009), 24 h after electroporation (Supplementary Figure S1D and E). These results indicate that the nuclei of neural progenitor cells do not migrate in the apical direction when they are arrested in G1-phase and suggest that entry into S-phase is a prerequisite for basal-to-apical nuclear migration.

Bottom Line: Here, we show that INM proceeds through the cell cycle-dependent linkage of cell-autonomous and non-autonomous mechanisms.In contrast, in vivo observations of implanted microbeads, acute S-phase arrest of surrounding cells and computational modelling suggest that the basal migration of G1-phase nuclei depends on a displacement effect by G2-phase nuclei migrating apically.Our model for INM explains how the dynamics of neural progenitors harmonize their extensive proliferation with the epithelial architecture in the developing brain.

View Article: PubMed Central - PubMed

Affiliation: Laboratory for Cell Asymmetry, RIKEN Center for Developmental Biology, Kobe, Japan. kosodo@med.kawasaki-m.ac.jp

ABSTRACT
A hallmark of neurogenesis in the vertebrate brain is the apical-basal nuclear oscillation in polarized neural progenitor cells. Known as interkinetic nuclear migration (INM), these movements are synchronized with the cell cycle such that nuclei move basally during G1-phase and apically during G2-phase. However, it is unknown how the direction of movement and the cell cycle are tightly coupled. Here, we show that INM proceeds through the cell cycle-dependent linkage of cell-autonomous and non-autonomous mechanisms. During S to G2 progression, the microtubule-associated protein Tpx2 redistributes from the nucleus to the apical process, and promotes nuclear migration during G2-phase by altering microtubule organization. Thus, Tpx2 links cell-cycle progression and autonomous apical nuclear migration. In contrast, in vivo observations of implanted microbeads, acute S-phase arrest of surrounding cells and computational modelling suggest that the basal migration of G1-phase nuclei depends on a displacement effect by G2-phase nuclei migrating apically. Our model for INM explains how the dynamics of neural progenitors harmonize their extensive proliferation with the epithelial architecture in the developing brain.

Show MeSH
Related in: MedlinePlus