Developmental roles of p73 in Cajal-Retzius cells and cortical patterning - PubMed (original) (raw)
Developmental roles of p73 in Cajal-Retzius cells and cortical patterning
Gundela Meyer et al. J Neurosci. 2004.
Abstract
To examine the role of the p53 homolog p73 in brain development, we studied p73-/-, p73+/-, E2F1-/-, and reeler mutant mice. p73 in developing brain is expressed in Cajal-Retzius (CR) cells, the cortical hem, and the choroid plexus. p73-expressing CR cells are lost in p73-/- embryos, although Reelin is faintly expressed in the marginal zone. Ectopic neurons in the p73-/- preplate and cortical hem at embryonic day 12 implicate p73 in the early developmental program of the cortex; however, preplate partition and early cortical plate formation are not disturbed. Postnatal p73-/- mice show a mild hypoplasia of the rostral cortex and a severely disrupted architecture of the posterior telencephalon. In the developing p73-/- hippocampus, the most striking abnormality is the absence of the hippocampal fissure, suggesting a role of p73 in cortical folding. p73+/- mice have a less severe cortical phenotype; they display a dorsal shift of the entorhinal cortex and a reduced size of occipital and posterior temporal areas, which acquire entorhinal-like features such as Reelin-positive cells in layer II. CR cells appear unaffected by heterozygosity. We relate the malformations of the posterior pole in p73 mutant mice to alterations of p73 expression in the cortical hem and suggest that p73 forms part of an early signaling network that controls neocortical and archicortical regionalization. In mice deficient for the transcription factor E2F1, a main activator of the TAp73 (transactivating p73) isoform, we find a defect of the caudal cortical architecture resembling the p73+/- phenotype along with reduced TAp73 protein levels and propose that an E2F1-TAp73 dependent pathway is involved in cortical patterning.
Figures
Figure 1.
The p73+/- and _p73_-/- brain phenotypes. A, P3 WT mice. B, P3 +/- mice. C, P3 -/- mice. D, P12 -/- mice, Nissl stain. B, The p73+/- brain displays abnormal shape and cytoarchitecture of caudal cortical areas and posterior cortical nucleus of the amygdala (CNA). A, B, Arrows point to the periamigdaloid fissure, which is shifted dorsally; the arrowhead in A points to the rhinal fissure, which is unrecognizable in B. C, D, The variability in the malformations of hippocampal architecture in _p73_-/- mice. All of the sections are at comparable levels. DG, Dentate gyrus; H, hilus. Scale bars, 200 μm.
Figure 2.
The posterior pole of the telencephalon. A, P7 WT mice. B, P7 -/- mice, parasagittal sections immunostained with GFAP. Cortical areas at the posterior pole are reduced in size in the _p73_-/- mouse. The white matter (WM) area is also smaller, and the hippocampal fissure (HF) is missing. F, Fimbria; RS, retrosplenial cortex; T, thalamus. C-F, The posterior pole of p73 heterozygous (D, F) and WT (C, E) mice at E18 in coronal Nissl (C, D) and Reelin (E, F) stained sections. Sections are at the same level to show the size reduction of the caudal telencephalon and the dorsal shift of entorhinal cortex. The proliferative subventricular zone (SVZ) is still visible in C (WT) but not in D (p73+/-). E, F, The number and distribution of Reelin-IR CR cells in the hippocampal fissure and in the cortical marginal zone are similar in WT and p73+/- mice. Scale bars, 200 μm.
Figure 3.
The cortical phenotype of _p73_-/- mice. A, B, P12 mice, Nissl stain. The frontal (motor) cortex of p73+/- mice (A) displays normal cytoarchitecture. In the _p73_-/- motor cortex (B), basic lamination and cytoarchitecture are preserved, but the cortex is smaller and white matter (wm) is reduced. _C, D, p73_-/- (same case as in B), stained with Nissl (C) and Reelin (D). In occipital cortex, lamination is lost and cell density is reduced. Irregular clusters of Reelin-IR neurons are present in the profoundly altered cortex. Scale bars: A, B, 200 μm; C, D, 100 μm.
Figure 4.
The preplate stage. A, B, D, E12 mice, ISH using specific probes for exon 3 (TAp73) in WT (A), exon 3 in _p73_-/- (B), and exon 3bis (DNp73) in WT (D) mice. TAp73 transcripts are expressed in the cortical hem of WT and _p73_-/- mice, whereas DNp73 transcripts are present in the WT cortical hem and in CR cells of the cortical preplate. C, E, Nissl-stained sections of a control brain at corresponding levels; C for A and B and E for D. The cortical hem is indicated by arrowheads. F, G, E12 mice, calretinin IHC, horizontal sections. F, WT mice. _G, p73_-/- embryo. The mutant preplate presents increased numbers of calretinin-IR pioneer neurons compared with the WT preplate. In addition, there are ectopic calretinin-IR neurons in the cortical hem (arrowheads) that are absent in the WT embryo. Arrows indicate the orientation; R, rostral; L, lateral; A, amygdala; E, eye; GE, ganglionic eminence; T, thalamus. Scale bars: A-E, 200 μm; F, G, 100 μm.
Figure 5.
Early development of the cortical plate. A, E15_p73_+/- mice. B, E15_p73_-/- mice, Reelin. A, CR cells form a dense cell layer below the pial surface. B, Only few faintly Reelin-positive cells are in the marginal zone (MZ). C, E15 p73+/- mice, high expression of p73 in CR cells. E, E15_p73_+/- mice. _F,p73_-/- mice, calretinin. E, CR cells at this stage are calretinin positive and are absent in the p73 knock-out brain. Asterisks mark calretinin-IR pioneer cells above and below the cortical plate (CP), and arrows point to thalamocortical fibers terminating in the subplate (SP); both elements are unchanged in the _p73_-/- brain. D, G, E16 Relnrl-Orl mice. D, Reelin and p73 are coexpressed in CR cells. Inset, p73 in the nucleus and Reelin in the cytoplasm of the CR cells. G, Calretinin stains superplate elements and aberrant thalamocortical fibers (arrows) traversing the abnormal CP. Compare with F, in which CP and thalamocortical fibers have normal position. Scale bars, 25 μm.
Figure 6.
Early development of the hippocampal fissure. A-C, E15 WT mice. D, E, E15 _p73_-/- mice. F, E16 Relnrl-Orl mice. CR cells express calretinin (A), Reelin (B), and p73 (C) and occupy the fissure (arrows), which begins to fold. In E15 _p73_-/- embryos (D, calretinin; E, Reelin), CR cells are absent and the fissure does not fold. F, Reelin. In E16 Relnrl-Orl mice, the hippocampal fissure develops normally. Scale bars, 100 μm.
Figure 7.
The postnatal hippocampal fissure (HF). A, B, Reelin ISH in WT (A) and _p73_-/- (B) mice at P2. B, CR cells are absent, and the molecular layers of dentate gyrus (DG) and CA1/CA2 are fused. C, D, Calretinin ISH in WT (C) and _p73_-/- (D) mice at P2. In the mutant, calretinin-positive hilus (H) cells have abnormal distribution. E, F, GFAP IHC in WT (E) and _p73_-/- (F) mice at P2. GFAP stains subpial astrocytes in the HF and the incipient radial glia scaffold in the dentate gyrus. In the mutant, astrocytes are found only along the medial brain surface. G, H, GFAP in WT (G) and _p73_-/- (H) mice at P12. The radial glia scaffold is fully established in the WT dentate; in the mutant, astrocytes are sparse and lack radial orientation. Arrowheads mark the entrance of the fissure, which fails to fold in the mutant. S, Subiculum. Scale bars: A, B, 200 μm; C-H, 100 μm.
Figure 8.
Architecture of the p73+/- telencephalon. Reelin in P3 p73+/- (A), P12 p73+/- (B), and P12 WT (C) brains. Reelin-IR cells in layer II mark the entorhinal cortex in the WT brain. In p73+/- brains, they are dorsally displaced and expanded, whereas the neocortex is reduced in size. The posterior cortical nucleus of the amygdala (CNA) is hyperplastic. The rhinal fissure (RF) is not visible in the p73+/- brains. D, E, Higher magnification of the boxed areas in A and D, respectively, to show ectopic Reelin-IR cells in layer II of the abnormal cortex and CR cells in layer I. F, At P3, CR cells in the abnormal p73+/- cortex are intensely p73 IR. Scale bars: A-C, 200 μm; D, 100 μm; E, F, 50 μm.
Figure 9.
The _E2F1_-/- cortex. Calretinin immunostaining in WT (A) and _E2F1_-/- (B) cortex at P1 shows that the caudal neocortex is transformed and acquires entorhinal-like features. Calretinin-IR interneurons are distributed differently in WT and _E2F1_-/- cortex, and the posterior cortical nucleus of the amygdala (CNA) is slightly larger in the mutant. C, In WT, Reelin-IR cells in layer II stop at the rhinal fissure (arrowheads in A and C). D, in the mutant, they extend dorsally into the altered neocortex. Scale bars: A, B, 200 μm; C, D, 100 μm.
Figure 10.
Expression of p73 in E2F1 knock-out mice. A, C, Immunoblot analysis of p73 in cortical tissue extracts probed with N-terminal (A) and C-terminal (C) antibodies. Note the important reduction of TAp73 levels at 82 kDa (TAp73α) and at 72 kDa (probably representing TAp73β) in mutant mice compared with the controls. After incubation with a C-terminal antibody, a DNp73 isoform at 66 kDa shows only a slight reduction in the E2F1 mutant (C). B, D, α-Tubulin controls to test the amount of protein analyzed.
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