Chromatin Dynamics in Developing Somites – Identification and Validation of Novel Regulatory Elements

Smith, Emily (2024) Chromatin Dynamics in Developing Somites – Identification and Validation of Novel Regulatory Elements. Doctoral thesis, University of East Anglia.

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Abstract

Paraxial mesoderm in developing vertebrae embryos gives rise to paired somites, forming on either side of the neural tube, which generate important cell lineages of the musculoskeletal system. Components of developmental signalling pathways regulating somite patterning and differentiation have been well characterised, however the genome-wide regulatory landscape is less well understood. I aimed to identify and characterise cis-regulatory elements (CREs) and associated transcription factors that form gene regulatory networks (GRN) important for the formation of musculoskeletal cell lineages.

Previously, dynamic changes in the chromatin landscape of paraxial mesoderm along the A-P axis of chicken embryos have been captured by combining ATAC-sequencing and RNA-sequencing datasets. Utilising these datasets, I identified differentially accessible regions of chromatin focussing on candidate CREs proximal to genes involved in paraxial mesoderm development and somite patterning. I validated novel CREs associated with the Tbx6 and Msgn1 genes in vivo, by electroporation of citrine-reporter plasmids into gastrula stage chick embryos. Further characterisation of the Tbx6 enhancer region by mutation of transcription factor motifs identified Cdx, Hox and Retinoic Acid Receptors as important for Tbx6 enhancer activation. These Citrine labelled CREs were then used to identify Tbx6, Msgn1 and Meox1 positive cells. I used these CREs to isolate citrine positive cells by FACS from chick embryos for mini-bulk ATAC-sequencing. This dataset further refined the chromatin accessibility signatures of different paraxial mesoderm cell types and was used to identify more CREs active in paraxial mesoderm cell types.

Item Type:	Thesis (Doctoral)
Faculty \ School:	Faculty of Science > School of Biological Sciences
Depositing User:	Kitty Laine
Date Deposited:	07 Apr 2025 15:01
Last Modified:	07 Apr 2025 15:01
URI:	https://ueaeprints.uea.ac.uk/id/eprint/98981
DOI:

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