Camilla Bosone

The Zfp423/ZNF423 gene encodes a 30-zinc-finger transcription factor involved in key developmental pathways. Although null Zfp423 mutants develop cerebellar malformations, the underlying mechanism remains unknown. ZNF423 mutations are associated with Joubert Syndrome, a ciliopathy causing cerebellar vermis hypoplasia and ataxia. ZNF423 participates in the DNA-damage response (DDR), raising questions regarding its role as a regulator of neural progenitor cell cycle progression in cerebellar development. To characterize in vivo the function of ZFP423 in neurogenesis, we analyzed allelic murine mutants in which distinct functional domains are deleted. One deletion impairs mitotic spindle orientation, leading to premature cell cycle exit and Purkinje cell (PC) progenitor pool deletion. The other deletion impairs PC differentiation. In both mutants, cell cycle progression is remarkably delayed and DDR markers are upregulated in cerebellar ventricular zone progenitors. Our in vivo evidence sheds light on the domain-specific roles played by ZFP423 in different aspects of PC progenitor development, and at the same time strengthens the emerging notion that an impaired DDR may be a key factor in the pathogenesis of JS and other ciliopathies.

Organoids generating major cortical cell types in distinct compartments are used to study cortical development, evolution and disorders. However, the lack of morphogen gradients imparting cortical positional information and topography in current systems hinders the investigation of complex phenotypes. Here, we engineer human cortical assembloids by fusing an organizer-like structure expressing fibroblast growth factor 8 (FGF8) with an elongated organoid to enable the controlled modulation of FGF8 signaling along the longitudinal organoid axis. These polarized cortical assembloids mount a position-dependent transcriptional program that in part matches the in vivo rostrocaudal gene expression patterns and that is lost upon mutation in the FGFR3 gene associated with temporal lobe malformations and intellectual disability. By producing spatially oriented cell populations with signatures related to frontal and temporal area identity within individual assembloids, this model recapitulates in part the early transcriptional divergence embedded in the protomap and enables the study of cortical area-relevant alterations underlying human disorders.

Gap junctions (GJs) are integral membrane proteins that enable the direct cytoplasmic exchange of ions and low molecular weight metabolites between adjacent cells. They are formed by the apposition of two connexons belonging to adjacent cells. Each connexon is formed by six proteins, named connexins (Cxs). Current evidence suggests that gap junctions play an important part in ensuring normal embryo development. Mutations in connexin genes have been linked to a variety of human diseases, although the precise role and the cell biological mechanisms of their action remain almost unknown. Among the big family of Cxs, several are expressed in nervous tissue but just a few are expressed in the anterior neural tube of vertebrates. Many efforts have been made to elucidate the molecular bases of Cxs cell biology and how they influence the morphogenetic signal activity produced by brain signaling centers. These centers, orchestrated by transcription factors and morphogenes determine the axial patterning of the mammalian brain during its specification and regionalization. The present review revisits the findings of GJ composed by Cx43 and Cx36 in neural tube patterning and discuss Cx43 putative enrollment in the control of Fgf8 signal activity coming from the well known secondary organizer, the isthmic organizer.

Cortical organoids generating all major cortical cell types in distinct compartments are widely used to mechanistically study cortical development, evolution and related disorders. However, the lack of morphogen gradients imparting cortical positional information and topography in current systems hinders the solid investigation of complex cortical phenotypes. Here, we describe an assembloid culture protocol that combines an organizer-like structure expressing the morphogen FGF8 with an elongated organoid, thus introducing a polarized source of FGF8 morphogen within individual organoids. This system, referred to as polarized cortical assembloid (polCA), allows for the establishment of FGF8-dependent positional identity along the longitudinal axis of individual organoids and the generation of spatially oriented populations with distinct transcriptional signatures relevant for the cortical protomap. This protocol can be adapted to include organizer-like structures expressing various morphogens, allowing for a reliable analysis of their effect on cortical positional identity.