Deficient adaptation to centrosome duplication defects in neural progenitors causes microcephaly and subcortical heterotopias
José González-Martínez, … , Sagrario Ortega, Marcos Malumbres
José González-Martínez, … , Sagrario Ortega, Marcos Malumbres
Published July 8, 2021
Citation Information: JCI Insight. 2021;6(16):e146364. https://doi.org/10.1172/jci.insight.146364.
View: Text | PDF
Research Article Cell biology Development

Deficient adaptation to centrosome duplication defects in neural progenitors causes microcephaly and subcortical heterotopias

Abstract

Congenital microcephaly (MCPH) is a neurodevelopmental disease associated with mutations in genes encoding proteins involved in centrosomal and chromosomal dynamics during mitosis. Detailed MCPH pathogenesis at the cellular level is still elusive, given the diversity of MCPH genes and lack of comparative in vivo studies. By generating a series of CRISPR/Cas9-mediated genetic KOs, we report here that — whereas defects in spindle pole proteins (ASPM, MCPH5) result in mild MCPH during development — lack of centrosome (CDK5RAP2, MCPH3) or centriole (CEP135, MCPH8) regulators induces delayed chromosome segregation and chromosomal instability in neural progenitors (NPs). Our mouse model of MCPH8 suggests that loss of CEP135 results in centriole duplication defects, TP53 activation, and cell death of NPs. Trp53 ablation in a Cep135-deficient background prevents cell death but not MCPH, and it leads to subcortical heterotopias, a malformation seen in MCPH8 patients. These results suggest that MCPH in some MCPH patients can arise from the lack of adaptation to centriole defects in NPs and may lead to architectural defects if chromosomally unstable cells are not eliminated during brain development.

Authors

José González-Martínez, Andrzej W. Cwetsch, Diego Martínez-Alonso, Luis R. López-Sainz, Jorge Almagro, Anna Melati, Jesús Gómez, Manuel Pérez-Martínez, Diego Megías, Jasminka Boskovic, Javier Gilabert-Juan, Osvaldo Graña-Castro, Alessandra Pierani, Axel Behrens, Sagrario Ortega, Marcos Malumbres

×

Figure 6

Genetic ablation of Trp53 in Cep135-KO embryos.

Options: View larger image (or click on image) Download as PowerPoint
Genetic ablation of Trp53 in Cep135-KO embryos. (A) IHC staining for act...
(A) IHC staining for active caspase 3 (AC3; brown) in the indicated models. Scale bars: 1 mm (whole embryos) and 100 μm (insets). In the quantification, data are mean SEM (3 embryos). **P < 0.01; ***P < 0.001 by Student’s t test. (B) H&E staining of the medial aspect of the neocortex of E14.5 embryos with the indicated genotypes. Scale bars: 20 μm (upper left), 10 μm (lower), and 100 μm (right). Arrows indicate aberrant mitotic figures of apical progenitors. Scale: 10 μm. (C) Histological sections of E14.5 neocortices of the indicated genotypes showing cortical heterotopias and malformations (arrowheads). Scale bars: 100 μm (upper) and 50 μm (insets). (D) Confocal imaging of E14.5 neocortices stained with the indicated antibodies (upper), and higher magnification confocal images showing the neocortical area delimited by slashed boxes (middle). Yellow arrowhead indicates a subcortical heterotopia. Higher-magnification inset in the middle panel shows CTIP2+ heterotopias. Bottom panels show APs with aberrant mitotic figures (arrowheads). Scale bars: 100 μm (upper), 50 μm (middle), and 10 μm (lower). (E) Confocal images of APs in the neocortices shown in D stained with the indicated antibodies. Scale bars: 10 μm. (F) Confocal micrographs of E14.5 neocortices stained with the indicated antibodies. The middle panels show higher-magnification images of the innermost APs with representative mitoses (insets) and monopolar spindles (yellow arrowheads), and acentromal cells (white arrowheads). Scale bars: 50 μm (left), 10 μm (middle), 10 μm (right). Asterisks indicate comparison with Cep135(+/+); Trp53(+/+) controls. Data are mean SEM from 3 embryos (1 for Cep135; Trp53 homozygous mutants). *P < 0.01; ***P < 0.001; ****P < 0.0001; 1-way ANOVA with Tukey’s multiple-comparison test.