Cytoskeletal re-arrangements leading to correct neuron polarisation during vertebrate CNS development

Neuronal differentiation during embryonic development is a fundamentally important process that ultimately results in the formation of functional neural circuitry. Differentiating neurons in the developing vertebrate spinal cord undergo the regulated process of apical abscission, resulting in acute loss of cell polarity, dismantling of the primary cilium and retention of the centrosome as they delaminate from the neuroepithelium (Das and Storey, Science, 2014). This loss of polarity results in a potentially hazardous cell state, requiring tight control, as the nascent neuron must now rapidly re-establish its polarity. We have recently discovered that delaminating neurons rapidly reassemble a new primary cilium, which facilitates a switch in the mode of Shh signalling that now drives polarisation and axon extension (Toro-Tapia and Das, Science Advances, 2020). As newborn neurons polarise in response to the extracellular Shh gradient, they must ultimately undergo cytoskeletal remodelling to effect the morphological changes characteristic of axon extension. However, the mechanisms through which primary cilium reassembly facilitates this cytoskeletal remodelling remains unknown.

This project will utilise cutting-edge, long-term high-resolution live-tissue imaging and advanced super-resolution STED fixed-tissue imaging to determine the normal dynamics of actin cytoskeletal remodelling during neuron polarisation and axon extension. The function of the primary cilium will then be disrupted by using targeted optogenetic methods and the effect on actin remodelling observed using live imaging. As the centrosome has previously been implicated as an actin organising centre in vitro, we will investigate if this function is recapitulated in vivo using live imaging and super resolution STED microscopy. Finally, the molecular mechanisms through which the primary cilium mediates normal actin remodelling during neuron repolarization will be examined.

Fit with BBSRC remit:
This project aims to investigate the fundamental mechanisms of neuron polarisation in the context of the developing spinal cord, and therefore fits the BBSRC remit of funding work investigating the normal biological mechanisms operating in animals. Furthermore, this work utilises novel and innovative live imaging techniques which are unique to the lab, which facilitate documentation and targeted disruption of cell behaviour in the context of a developing tissue. This work is therefore at the forefront of modern biological research and aligns well with the BBSRC DTP theme of world class underpinning biosciences, and facilitates new ways of understanding fundamental cell behaviour driving neuronal differentiation.