Molecular mechanisms of NMC plasticity and fate commitment

Their combined loss phenocopies the Wnt3a null phenotype, implicating them in the Wnt3a-Tbxt autoregulatory loop and, by extension, the maintenance of NMC self-renewal and mesoderm formation. Yet, details regarding the mechanisms by which Sp proteins interface with the wider NMC-GRN remains unclear.<br>In this thesis, we illustrate that Sp proteins are critical regulators of NMC homeostasis throughout axis elongation. We find that Sp5 and Sp8, together with other NMC-GRN regulators, control the network through reciprocal interactions and converge on a cis-regulatory element (cRE) downstream of Wnt3a.

In gastruloid models, we show this element is essential for Wnt3a expression, and its deletion collapses the Wnt3a-Tbxt autoregulatory loop, driving premature depletion of axial progenitors. Using single-cell multi-omics, we show that Sp8 is required to sustain NMC homeostasis during later stages of axis elongation. Loss of Sp8 was shown to remodel the epigenetic landscape and induce a transcriptional response that destabilizes the NMC-GRN, biasing NMCs toward neural lineages at the expense of mesoderm and self-renewal.

Importantly, we link this bias and the resulting exhaustion of the NMC pool to the absence of tail vertebrae and an expanded posterior neural plate, a feature we propose underlies the spina bifida–like phenotype in Sp8 mutants. Together, our findings establish Sp proteins as central components of the NMC-GRN and provide insight linking the fundamental molecular regulation of axial progenitors to the aetiology of clinically relevant phenotypes.<br><br><i>Thesis is embargoed until 31 July 2031.</i><br>