Sammut, Michele; (2022) Key cell cycle regulators are implicated in both proliferative and direct glia-to-neuron cell fate switches in C. elegans. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

In the vertebrate nervous system, stably differentiated quiescent radial glial cells act as neural progenitors, for the first time we describe the production of neurons from glia outside of vertebrate models. Here we show that two sets of differentiated glial cells are capable of sex-specifically producing neurons in the male C. elegans. The two glia-to-neuron switches in cell fate, occur by distinct developmental mechanisms; the amphid socket (AMso) glial cells, undergo asymmetric division to produce the interneurons, the mystery cells of the male (MCMs), while the phasmid socket 1 (PHso1) glial cells directly transdifferentiate into the sensory phasmid neurons D (PHDs). In order to identify genetic factors that regulate the production of the MCMs, we have isolated nine no mystery cell (nom) mutants from a GFP-based forward genetic screen, in which the MCMs fail to be specified. Using a novel mapping-by-sequencing method, designed for male specific phenotypes, we find that two mutant alleles nom-5(drp5) and nom-8(drp8) which block AMso division, map to the cdk-4 locus on the X chromosome. Genetic analysis by rescue and complementation confirms they are alleles of cdk-4. CDK-4 is a key cell cycle regulator and intriguingly, although cdk-4 is required for all postembryonic cell divisions cdk-4(drp5) and cdk-4(drp8) present no obvious pleiotropies, hinting that novel cell cycle pathways may be involved in this specific division. The development of the MCMs and PHD share molecular features in common with vertebrate neural development, such as the expression of hlh-14, the homologue of the proneural factor ASCL1. Also, analysis of cki-1, a commonly used marker of cell cycle quiescence and G1/S regulator, shows that it is strongly expressed in both the AMso and PHso1 throughout their life prior to the production of neurons. A further surprising finding is that S-phase genes are expressed during the direct PHso1-to-PHD transdifferentiation. The role of S-phase genes in a direct transdifferentiation, that may not require DNA synthesis, is unknown. An understanding of fate plasticity is an often overlooked but key component to the processes of neurogenesis from glia, this model promises to deepen our understanding. How the progenitor cells of neurons control their ability to either re-enter or withdraw from the cell cycle and how this decision is integrated with the terminal differentiation process is an outstanding question. To this end we begin to establish the C. elegans male as a model in which to study the molecular mechanisms of late-in-development cell fate plasticity at a single cell level.

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