Genome editing tool for studying Ciona robusta nervous system differentiation

Olivo, Paola (2023). Genome editing tool for studying Ciona robusta nervous system differentiation. PhD thesis The Open University.



The development of the central nervous system (CNS) depends on complex gene regulatory networks (GRN) that orchestrate the specification, patterning and differentiation of neural cell types. Taking advantage of the unique characteristics of a simple chordate, the ascidian Ciona robusta, I identified a group of genes, expressed in the nervous system of Ciona, to investigate the network that controls specification inside the central and peripheral nervous system. The approach used to study functionally these genes has been the gene editing, performed by CRISPR/cas9 technique, indeed in this work I had the opportunity to evaluate the whole process of this emerging technique, preparing and testing several sgRNAs on the selected genes. Moreover, I focused my attention on a type of sensory neuron, belonging to the peripheral nervous system, Bipolar tail neurons (BTNs), through the investigation of two poorly studied genes, Rimbp and LZTS, both expressed in the BTNs neurons. This thesis amplified the knowledge on their involvement in the gene regulatory network of BTNs during Ciona nervous system development. Here I showed that CRISPR/Cas9-mediated knockout of LZTS in the epidermis results in extra BTNs, suggesting LZTS functions as a repressor during differentiation and specification of BTNs. All these data provide new insight into the development of the Ciona nervous system, encouraging further studies to clarify and confirm LZTS role in the Ciona nervous system development.

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