The scaffold protein Nde1 safeguards the brain genome during S phase of early neural progenitor differentiation
- Northwestern University Feinberg School of Medicine, United States
Figures
Figure 1 with 3 supplements
The correlation of DNA damage and apoptosis with neural progenitor early fate restriction in Nde1 mutant brains.
(A) Immunohistological analysis of γH2AX (red) and cleaved caspase 3 (CC3, green) reveals the co-existence of DNA damage and apoptosis in the neocortex of Nde1 mutants at E12.5. Higher-magnification …
Figure 1—figure supplement 1
The correlation of DNA damage and apoptosis with neural progenitor early fate restriction in Nde1 mutants.
Immunohistological analysis with antibodies to γH2AX (red) and cleaved caspase 3 (CC3, green) shows the spatiotemporal concurrence of DNA damage and apoptosis with early stages of neuronal …
Figure 1—figure supplement 2
DNA damage and apoptosis along the transverse neurogenetic gradient (TNG) in Nde1−/− brains.
Immunohistological analysis of serial coronal sections of Nde1−/− brains at early and late E12, respectively, to demonstrate the spatial correlation of cleaved caspase 3 (CC3, green) with TNG …
Figure 1—figure supplement 3
The correlation of DNA damage and apoptosis with neuronal differentiation in the Nde1 mutant spinal cord from E9.5 to E13.5.
Immunohistological analysis of γH2AX (red) and cleaved caspase 3 (CC3, green) of the developing spinal cord of Nde1−/−Lis1+/− (M) and control (C, Nde1+/− or wild type) littermates from E9.5 to …
Figure 2
Co-activation of γH2AX with p53 in Nde1 mutant neocortices.
(A) Results from comet assay with cortical cells isolated from Nde1−/− or Nde1−/−Lis1+/− mutants and their Nde1+/− littermates at E12.5. The distribution, mean, and median values of comet tail …
Figure 3 with 1 supplement
Restoration of the size and structure of the Nde1−/− brain by abrogating p53.
(A) Immunohistological analyses of cleaved caspase 3 (CC3, green) in wild-type, Nde1−/−, Nde1+/−Trp53−/−, and Nde1−/−Trp53−/− neocortices at E12.5. (B) Brain weight of Nde1−/−, Nde1−/−Trp53−/− …
Figure 3—figure supplement 1
Restoration of brain size and structure of Nde1−/− mutants by abrogating p53.
(A) Representative brain images of wild-type, Nde1−/−, and Nde1−/−Trp53−/− mice at post-natal day 23. (B) Immunostaining with parvalbumin antibody (PV) showing the normal number and distribution of …
Figure 4 with 2 supplements
DNA damage caused by Nde1 mutation occurs concurrently with DNA replication.
(A) Co-immunostaining of γH2AX (red) and multipotent/primary progenitor marker Pax6 (green) on cortical sections at E12.5. (B) Co-immunostaining of γH2AX (red) and BrdU (green); cortical sections …
Figure 4—figure supplement 1
The cell type and cell cycle specificity of DNA damage in Nde1 mutant brains.
(A) Double immunohistological staining E12.5 cortical sections with γH2AX (green) and the new neuron marker DCX (red). (B) Double immunohistological staining E13.5 cortical sections with γH2AX (red) …
Figure 4—figure supplement 2
Stalled or delayed DNA replication in Nde1 mutant neural progenitors.
(A) Triple immunofluorescence histological staining of neocortical sections with B44 (green), BU1/75 (red), and PH3 (blue), showing the lack of PH3 immunoreactivity in the arrow-indicated B44+BU1− …
Figure 5
Stalled DNA replication during mid-late S phase at heterochromatic domains in Nde1−/− mutant neural progenitors.
(A) Quantitative analysis of IdU+CldU− (red), IdU+CldU+ (yellow), and IdU−CldU+ (green) cell fractions (%) by IdU (2 hr) and CldU (30 min) sequential labeling. Data are presented as mean ± SD. n.s.: …
Figure 6 with 1 supplement
Persistent cell cycle stress and genotoxicity in Nde1 mutants after p53 abrogation.
(A) Representative images of B44 (green, recognizes both IdU and BrdU) and BU1/75 (red, recognizes only BrdU) double immunostained neocortical sections from IdU (2 hr), BrdU (30 min) sequential …
Figure 6—figure supplement 1
Lymphomagenesis and remarkably increased p53 in Nde1−/−Trp53+/− thymic lymphoma.
(A) Representative T cell thymic lymphoma of Nde1−/−Trp53+/− mice at 3 months. (B) Representative image of H&E stained Nde1−/−Trp53+/− lymphoma. Apoptotic cells with fragmented nuclei are indicated …
Figure 7 with 2 supplements
Identification of a nuclear pool of Nde1 that interacts with the cohesin complex.
(A) Double immunohistological staining with antibodies to Nde1 (red) and Pax6 (green) reveals the presence of Nde1 in the nucleus of cells in the neocortical VZ. Notice the detection of Ndel1 in the …
Figure 7—figure supplement 1
Blocking Nde1–cohesin interaction results in apoptosis.
Immunofluorescence analysis of 293T cells transfected by SMC3 Flag-NBD (red) showing increased apoptosis indicated by cleaved caspase 3 immunoreactivities (CC3, green) and micronuclei indicated by …
Figure 7—figure supplement 2
Increased genomic instability of Nde1 mutant MEFs.
(A) Karyotyping analysis shows the rapid development of aneuploidy in Nde1−/− and Nde1−/−Trp53−/− MEFs. Chromosome number distributions (% of total diploid cells) of wild-type and Nde1−/− MEF lines …
Figure 8
Nde1's role in maintaining genome integrity during early neural differentiation.
During early neural differentiation, Nde1 mutant progenitors experience catastrophic DNA damage concurrent with mid-late S phase, when heterochromatic replication occurs. This evokes a DNA damage …
