Biology subjects

Constable, S.

Publications and source records attributed to Constable, S..

3 recordsLinked to original sources

bioRxiv2023

Programmed withdrawal of cilia maintenance followed by centriole capping leads to permanent cilia loss during cerebellar granule cell neurogenesis

Primary cilia in brain neurons provide a privileged compartment for binding and responding to extracellular ligands such as sonic hedgehog. Paradoxically, cilia in differentiating cerebellar granule cells are deconstructed during neurogenesis. To identify mechanisms underlying this newly defined cilia deconstruction pathway, we used single cell transcriptomic and immunocytological analyses to compare the transcript and protein signatures of differentiating and progenitor granule cells. We found that differentiating granule cells lacked transcripts for key regulators of pre-mitotic cilia resorption, suggesting cilia disassembly in differentiating cells was distinct from pre-mitotic cilia resorption. Further analysis revealed that during differentiation, transcription of genes required for cilia maintenance decreased. Specifically, protein components of intraflagellar transport complexes, pericentrosomal material and centriolar satellites all decreased as granule cells matured. The changes in transcription and translation correlated with the downregulation of sonic hedgehog signaling at the onset of differentiation. We also found binding of centriolar cap proteins to the mother centrioles as granule cell neurons matured. These data indicate that global, developmentally programmed, diminution of cilium maintenance caused cilia deconstruction in differentiating granule cells. Furthermore, the capping of docked mother centrioles prevents cilia regrowth likely blocking dysregulated sonic hedgehog signaling and tumorigenesis.

cell biology↗

bioRxiv2023

Permanent deconstruction of intracellular primary cilia in differentiating granule cell neurons

Primary cilia on granule cell neuron progenitors in the developing cerebellum detect sonic hedgehog to facilitate proliferation. Following differentiation, cerebellar granule cells become the most abundant neuronal cell type in the brain. While essential during early developmental stages, the fate of granule cell cilia is unknown. Here, we provide nanoscopic resolution of ciliary dynamics in situ by studying developmental changes in granule cell cilia using large-scale electron microscopy volumes and immunostaining of mouse cerebella. We found that many granule cell primary cilia were intracellular and concealed from the external environment. Cilia were disassembed in differentiating granule cell neurons in a process we call cilia deconstruction that was distinct from pre-mitotic cilia resorption in proliferating progenitors. In differentiating granule cells, ciliary loss involved unique disassembly intermediates, and, as maturation progressed, mother centriolar docking at the plasma membrane. Cilia did not reform from the docked centrioles, rather, in adult mice granule cell neurons remained unciliated. Many neurons in other brain regions require cilia to regulate function and connectivity. In contrast, our results show that granule cell progenitors had concealed cilia that underwent deconstruction potentially to prevent mitogenic hedgehog responsiveness. The ciliary deconstruction mechanism we describe could be paradigmatic of cilia removal during differentiation in other tissues.

cell biology↗

bioRxiv2019

Ciliary phosphatidylinositol phosphatase Inpp5e plays positive and negative regulatory roles in Shh signaling

Sonic hedgehog (Shh) signal transduction specifies ventral cell fates in the neural tube and is mediated by the Gli transcription factors that play both activator (GliA) and repressor (GliR) roles. Cilia are essential for Shh signal transduction and the ciliary phosphatidylinositol phosphatase, Inpp5e, is linked to Shh regulation. In the course of a forward genetic screen for recessive mouse mutants, we identified a functional null allele of Inpp5e, ridge top (rdg), with expanded ventral neural cell fates at E10.5. By E12.5, Inpp5erdg/rdg embryos displayed normal neural patterning and this correction over time required Gli3, the predominant repressor in neural patterning. Inpp5erdg function largely depended on the presence of cilia and on Smoothened, the obligate transducer of Shh signaling, indicating Inpp5e functions within the cilium to regulate the pathway. These data indicate that Inpp5e plays a more complicated role in Shh signaling than previously appreciated. We propose that Inpp5e attenuates Shh signaling in the neural tube through regulation of the relative timing of GliA and GliR production, which is important in understanding how duration of Shh signaling regulates neural tube patterning.\n\nSummary statementInpp5e attenuates Sonic hedgehog signal transduction through a combination of positive and negative regulatory roles that likely control the relative timing of Gli processing.

developmental biology↗