Cdh-2 modulates tissue stiffness and cortical F-actin distribution in neuroepithelial cells to drive forebrain roof plate invagination

The invagination of the embryonic forebrain roof plate is critical for cerebral hemisphere formation. Disruption of this process leads to holoprosencephaly (HPE), a severe congenital brain malformation. Although mutations in several genes have been linked to HPE and studies using the chick embryo have implicated certain signaling pathways, yet the mechanisms by which mechanical signals are translated into tissue morphological changes remain poorly understood. To address this lacuna, we employed atomic force microscopy to map spatiotemporal stiffness gradients across the dorsal forebrain in the chick embryo during roof plate invagination. Our analysis revealed that the initial uniformly elevated levels of stiffness across the roof plate are dramatically altered during morphogenesis. A pronounced stiffness gradient emerges, whereby the roof plate midline becomes markedly more compliant than dorsolateral regions. Through expression screening, we identified Cdh-2 as a candidate mechanical regulator whose spatiotemporal expression pattern precisely mirrors the observed stiffness gradient. Both loss- and gain-of-function perturbations of Cdh-2 disrupted the normal stiffness gradient and caused significant architectural alterations, such as evagination or a V-shaped invagination of the roof plate. Mechanistically, Cdh-2 modulates tissue stiffness by regulating adherens junction stability and cortical F-actin distribution along the apicobasal axis of neuroepithelial cells. These findings highlight the importance of Cdh-2 and provide critical insights into the mechanical forces and molecular interactions governing forebrain roof plate morphogenesis while shedding light on the of pathogenesis of HPE.