A differential requirement for ciliary transition zone proteins in human and mouse neural progenitor fate specification

Abstract

AbstractStudying developmental processes in the context of the human central nervous system is essential to understand neurodevelopmental diseases. In this paper we perform a comparative functional study of the ciliopathy geneRPGRIP1Lin human and mouse spinal development usingin vitro3D differentiation of pluripotent stem cells.RPGRIP1L, a causal gene of severe neurodevelopmental ciliopathies such as Joubert and Meckel syndromes, encodes a scaffolding protein of the ciliary transition zone involved in ciliary gating. Previous work has identified a major role forRpgrip1lin mouse brain and spinal cord development, via controlling the Sonic Hedgehog (SHH)/GLI pathway. We show that spinal organoids derived fromRpgrip1lmutant mouse embryonic stem cells faithfully recapitulate the loss of motoneurons and the strong reduction of SHH signaling observed in the mutant mice. In contrast, human induced pluripotent stem cells mutant forRPGRIP1Lproduce motoneurons and activate the SHH pathway at levels similar to wild types, a property shared by human iPSCs mutant for another ciliopathy geneTMEM67. Moreover, we show that, in humanRPGRIP1Lmutant organoids, motoneurons acquire a more anterior identity, expressingHOXgenes and proteins normally present in the hindbrain while motoneurons from wild type organoids strictly display spinal identity. By performing a temporal transcriptome analysis throughout the differentiation process, we find that the anteroposterior specification defect arises in early axial progenitors and correlates with the loss of cilia in these cells. Thus, this study uncovers distinct functions in humans and mice for ciliopathy proteins and a novel role for RPGRIP1L in human spinal anteroposterior patterning. These findings have important implications for understanding the role of cilia in human spinal cord development and the pathogenic mechanisms of neurodevelopmental ciliopathies.