Specific polar subpopulations of astral microtubules control spindle orientation and symmetric neural stem cell division
- Max Planck Institute of Molecular Cell Biology and Genetics, Germany
- RIKEN Center for Developmental Biology, Japan
Figures
Figure 1
Dynamic spindle orientation variability increases when progenitors become neurogenic.
Live tissue imaging of spindle orientation as reported by chromosome plate orientation in organotypic slice culture of coronal sections from E14.5 Tis21::GFP mouse dorsolateral telencephalon. …
Figure 2
Fewer astral microtubules when progenitors become neurogenic.
(A) Cartoons of the mitotic spindle in APs as appearing upon ‘Front’ and ‘Side’ viewing of the same chromosome plate, rotating around its apico-basal axis. Apical- and basal-reaching astrals …
Figure 3
Fewer apical/basal astral microtubules in progenitors as neurogenesis progresses.
A, B) α-Tubulin immunofluorescence (maximum intensity projections of two 0.75 μm optical sections) of E11.5 and E16.5 mouse dorsolateral telencephalon showing mitotic microtubules in APs. Dashed …
Figure 4
Fewer astral microtubules in LGN KO progenitors and upon dominant-negative LGN-C overexpression.
(A–D) α-Tubulin immunofluorescence (maximum intensity projections of two 0.75 μm optical sections) of E14.5 heterozygous (LGN +/−) vs homozygous (LGN −/−) E14.5 LGN KO mouse dorsolateral …
Figure 5
Less cell cortical LGN in neurogenic progenitors.
(A–C) Double immunofluorescence for LGN and GFP, with DNA staining (DAPI), of E14.5 Tis21::GFP mouse dorsolateral telencephalon showing representative examples of a proliferating AP (Tis21::GFP–) vs …
Figure 6 with 1 supplement
Changing the number of apical/basal astral microtubules in APs.
A–E) Controlled decrease in the apical/basal astrals of proliferating APs under minimal nocodazole. E14.5 forebrains from Tis21::GFP mice were incubated for 2.5 hr in BRO culture, either with …
Figure 6—figure supplement 1
No increase in apical/basal astral microtubules of BPs upon wt LGN overexpression combined with minimal taxol incubation.
E13.5 wt forebrains were subjected to in utero electroporation of 6Myc-LGN-wt and 18 hr later (E14.25) incubated for 2.5 hr in BRO culture with either solvent (DMSO) only (Control) or with 25 pM …
Figure 7 with 1 supplement
Key spindle and cell features in APs are not altered by 30 pM nocodazole.
All measurements were performed on the same cell images from which the astral microtubule data were obtained (see Figures 2 & 4). (A) Cartoon and example image (α-tubulin immunofluorescence) of the …
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Figure 7—source data 1
Table with the values for the graphs in Figure 7.
- https://doi.org/10.7554/eLife.02875.014
Figure 7—figure supplement 1
Key subcellular, cellular, and tissue features in APs are not significantly altered by 30 pM nocodazole.
E14.5 forebrains from wt mice were incubated for 2.5 hr in BRO culture either with solvent (DMSO) only (control) or with 30 pM nocodazole, and immunofluorescences of dorsolateral telencephalon …
Figure 8 with 1 supplement
Dynamic spindle orientation variability decreases with minimal taxol incubation combined with LGN-wt overexpression, but not with taxol alone.
A–D) E14.5 forebrains from Tis21::GFP mice were incubated on the microscope stage with either solvent (DMSO) only (Control) or with 25 pM taxol. Live tissue imaging of spindle orientation, as …
Figure 8—figure supplement 1
No increase in dynamic spindle orientation variability of BPs upon wt LGN overexpression combined with minimal taxol incubation.
E13.5 wt forebrains were subjected to in utero electroporation of 6Myc-LGN-wt and 18 hr later (E14.25) incubated for 2.5 hr in BRO culture with either solvent (DMSO) only (Control) or with 25 pM …
Figure 9
Dynamic spindle orientation variability and asymmetric AP divisions increase with minimal nocodazole.
E14.5 forebrains from Tis21::GFP mice were incubated on the microscope stage with either solvent (DMSO) only (Control) or with 30 pM nocodazole (Noc). (A–D) Live tissue imaging of spindle …
Figure 10 with 1 supplement
Increase in basal progenitors, basal mitoses, and neurogenesis with minimal nocodazole.
E13.5 forebrains from wt mice were incubated for 24 hr in BRO culture, either with solvent (DMSO) only (Control) or with 30 pM nocodazole, followed by immunofluorescence of coronal sections of the …
Figure 10—figure supplement 1
BRO culture with 30 pM nocodazole increases Tis21-positive VZ cells and basal mitoses, but not apoptosis.
E13.5 forebrains from wt mice were incubated for 24 hr in BRO culture, either with solvent (DMSO) only (control) or with 30 pM nocodazole, followed by immunofluorescence of coronal sections of the …
Author response image 1
No LGN immunoreactivity in LGN –/– mice.
E13.5 LGN+/– (top) and LGN–/– (bottom) mice (as in Konno et al, 2008). 1-μm confocal sections of immunofluorescences for endogenous LGN (red), together with DNA (DAPI, cyan) staining, of coronal …
Videos
Video 1
Dynamic spindle orientation variability in a representative proliferating AP. Related to Figure 1A.
Live tissue imaging of spindle orientation, as reported by the chromosome plate (DNA) orientation, in organotypic slice culture of dorsolateral telencephalon coronal sections, from an E14.5 …
Video 2
Dynamic spindle orientation variability in a representative neurogenic AP. Related to Figure 1B.
Live tissue imaging of spindle orientation, as reported by the chromosome plate (DNA) orientation, in organotypic slice culture of dorsolateral telencephalon coronal sections, from an E14.5 …
Video 3
Dynamic spindle orientation variability in a representative neurogenic BP. Related to Figure 1C.
Live tissue imaging of spindle orientation, as reported by the chromosome plate (DNA) orientation, in organotypic slice culture of dorsolateral telencephalon coronal sections, from an E14.5 …
Video 4
Dynamic spindle orientation variability in a representative proliferating AP incubated with DMSO only at 1.5 min time resolution. Related to Figure 8A.
Live tissue imaging of spindle orientation, as reported by the chromosome plate (DNA) orientation, in organotypic slice culture of dorsolateral telencephalon coronal sections, from an E14.5 …
Video 5
Dynamic spindle orientation variability in a representative proliferating AP incubated with 25 pM taxol at 1.5 min time resolution. Related to Figure 8B.
Live tissue imaging of spindle orientation, as reported by the chromosome plate (DNA) orientation, in organotypic slice culture of dorsolateral telencephalon coronal sections, from an E14.5 …
Video 6
Dynamic spindle orientation variability in a representative neurogenic AP incubated with DMSO only at 1.5 min time resolution.
Related to Figure 8C. Live tissue imaging of spindle orientation, as reported by the chromosome plate (DNA) orientation, in organotypic slice culture of dorsolateral telencephalon coronal sections, …
Video 7
Dynamic spindle orientation variability in a representative neurogenic AP incubated with 25 pM taxol at 1.5 min time resolution.
Related to Figure 8D. Live tissue imaging of spindle orientation, as reported by the chromosome plate (DNA) orientation, in organotypic slice culture of dorsolateral telencephalon coronal sections, …
Video 8
Dynamic spindle orientation variability in a representative AP incubated with DMSO only and co-electroporated with GAP43-EGFP and 6Myc-LGN-wt.
Related to Figure 8E. Live tissue imaging of spindle orientation, as reported by the chromosome plate (DNA) orientation, in organotypic slice culture of dorsolateral telencephalon coronal sections, …
Video 9
Dynamic spindle orientation variability in a representative AP incubated with 25 pM taxol and co-electroporated with GAP43-EGFP and 6Myc-LGN-wt.
Related to Figure 8F. Live tissue imaging of spindle orientation, as reported by the chromosome plate (DNA) orientation, in organotypic slice culture of dorsolateral telencephalon coronal sections, …
Video 10
Dynamic spindle orientation variability in a representative proliferating AP incubated with DMSO only.
Related to Figure 9A. Live tissue imaging of spindle orientation, as reported by the chromosome plate (DNA) orientation, in organotypic slice culture of dorsolateral telencephalon coronal sections, …
Video 11
Dynamic spindle orientation variability in a representative proliferating AP incubated with DMSO only.
Related to Figure 9B. Live tissue imaging of spindle orientation, as reported by the chromosome plate (DNA) orientation, in organotypic slice culture of dorsolateral telencephalon coronal sections, …
Video 12
Dynamic spindle orientation variability in a representative neurogenic AP incubated with 30 pM nocodazole.
Related to Figure 9C. Live tissue imaging of spindle orientation, as reported by the chromosome plate (DNA) orientation, in organotypic slice culture of dorsolateral telencephalon coronal sections, …
Video 13
Dynamic spindle orientation variability in a representative neurogenic AP incubated with 30 pM nocodazole.
Related to Figure 9D. Live tissue imaging of spindle orientation, as reported by the chromosome plate (DNA) orientation, in organotypic slice culture of dorsolateral telencephalon coronal sections, …
