Genetic Deletion of Afadin Causes Hydrocephalus by Destruction of Adherens Junctions in Radial Glial and Ependymal Cells in the Midbrain

Abstract

Adherens junctions (AJs) play a role in mechanically connecting adjacent cells to maintain tissue structure, particularly in epithelial cells. The major cell–cell adhesion molecules at AJs are cadherins and nectins. Afadin binds to both nectins and α-catenin and recruits the cadherin-β-catenin complex to the nectin-based cell–cell adhesion site to form AJs. To explore the role of afadin in radial glial and ependymal cells in the brain, we generated mice carrying a nestin-Cre-mediated conditional knockout (cKO) of the afadin gene. Newborn afadin-cKO mice developed hydrocephalus and died neonatally. The afadin-cKO brain displayed enlarged lateral ventricles and cerebral aqueduct, resulting from stenosis of the caudal end of the cerebral aqueduct and obliteration of the ventral part of the third ventricle. Afadin deficiency further caused the loss of ependymal cells from the ventricular and aqueductal surfaces. During development, radial glial cells, which terminally differentiate into ependymal cells, scattered from the ventricular zone and were replaced by neurons that eventually covered the ventricular and aqueductal surfaces of the afadin-cKO midbrain. Moreover, the denuded ependymal cells were only occasionally observed in the third ventricle and the cerebral aqueduct of the afadin-cKO midbrain. Afadin was co-localized with nectin-1 and N-cadherin at AJs of radial glial and ependymal cells in the control midbrain, but these proteins were not concentrated at AJs in the afadin-cKO midbrain. Thus, the defects in the afadin-cKO midbrain most likely resulted from the destruction of AJs, because AJs in the midbrain were already established before afadin was genetically deleted. These results indicate that afadin is essential for the maintenance of AJs in radial glial and ependymal cells in the midbrain and is required for normal morphogenesis of the cerebral aqueduct and ventral third ventricle in the midbrain.

Impairment of radial glial scaffold-dependent neuronal migration and formation of double cortex by genetic ablation of afadin

“…Several lines of evidence shown here indicate that double cortex was formed by an abnormal radial glial scaffold likely resulting from the disruption of AJs. We previously showed that the Nestin-Cre-afadin-cKO mice develop severe hydrocephalus due to the defects in the midbrain (Yamamoto et al, 2013), making it hard to analyze the architecture of the afadin-ablated cerebral cortex. In contrast to nestin, which is expressed in many regions including the telencephalon and the midbrain (Wiese et al, 2004), Emx1 is exclusively expressed in the dorsal telencephalon, but not in the midbrain (Simeone et al, 1992a(Simeone et al, , 1992b.…”

“…In vivo, afadin systemic KO mice show developmental defects at stages during and after gastrulation and that these mice die at early embryonic stages (Ikeda et al, 1999;Zhadanov et al, 1999). Furthermore, Nestin-Cre-mediated central nervous system-specific afadin cKO mice die perinatally, because they develop severe hydrocephalus caused by the disruption of AJs of radial glial and ependymal cells in the midbrain (Yamamoto et al, 2013). Therefore, it was difficult to analyze the roles of afadin for the formation of the cerebral cortex using these afadin mutant mice.…”

Genetics and mechanisms leading to human cortical malformations

“…Related genes, such as α-E-catenin, Ccdc85C, Rapgef2/Rapgef6, Afadin and RhoA induce similar, severe phenotypes (e.g. SBH or PVHlike phenotypes) in the mouse when they are down-regulated or suppressed during cortical development (see Table 6) [195,[272][273][274][275][276]. However, no mutations in these latter genes have been reported in human, and in some cases constitutive phenotypes are lethal in the mouse (See Table 6).…”

Cooperative Roles of Nectins with Cadherins in Physiological and Pathological Processes

“…The nectin-afadin system maintains the formation of AJs between RGCs in the midbrain and the dorsal telencephalon and between ependymal cells in the midbrain in concert with N-cadherin for the formation of the layer structure in the developing cerebral cortex (Gil-Sanz et al 2014;Yamamoto et al 2013Yamamoto et al , 2015. In the midbrain, loss of afadin causes hydrocephalus by mislocalization of RGCs in the ventricular and intermediate zones, mislocalization of neurons at the surface of the cerebral aqueduct, and loss of ependymal cells from the ventricular and aqueductal surfaces, accompanied by the reduced localization of nectin-1 and N-cadherin and the disruption of AJs.…”