title: Single-cell dissection of regulatory landscapes during embryogenesis
creator: Secchia, Stefano
description: Gene regulatory networks control the precise spatial and temporal execution of  developmental gene expression programs during embryogenesis. Enhancers are core  components of these regulatory networks and they serve as integration platforms for  the developmental signals delivered by transcriptions factors (TFs). Dissecting the  function of gene regulatory networks and their underlying components is a major goal  of modern biology. In this thesis, I leverage the recent advances in single-cell genomics  methods to assess the impact of perturbations on developmental regulatory networks,  using Drosophila melanogaster embryogenesis as a well-studied model system.    I started by optimizing a protocol to profile single-cell chromatin accessibility by sci-  ATAC-seq in Drosophila embryos and applied it to profile a dense time-course of over  20,000 cells during mesoderm development. The time-course comprised eight  overlapping time-points spanning half of embryogenesis and captured a continuum of  regulatory transitions as cells move from multipotency to different developmental  lineages. I used this dataset to reconstruct developmental trajectories of all major celltypes,  and uncover both the TFs and enhancers involved.    I then present two approaches that integrate the single-cell resolution of sci-ATAC-seq  with perturbations of TFs and enhancers, as a means to dissect regulatory networks.  First, I perturbed regulatory networks in trans, by mutating four key developmental TFs  that drive Drosophila mesoderm development and used sci-ATAC-seq to jointly assess  the regulatory outcome at both the cellular and molecular level. I demonstrate that this  approach not only recovers previously described high-level phenotypes, but also more  subtle alterations in cell fate, while simultaneously providing information on the TF’s  input at enhancers. Second, I perturbed regulatory networks in cis by exploiting natural  sequence variation as a means for large-scale enhancer disruption. By profiling  chromatin accessibility in hybrid embryos obtained from mating genetically diverse  parent lines, I show how sci-ATAC-seq can be used to discover the cellular context  affected by genetic variants and I discuss upcoming in vivo experiments to assess their  impact on enhancer activity.
date: 2022
type: Dissertation
type: info:eu-repo/semantics/doctoralThesis
type: NonPeerReviewed
format: application/pdf
identifier: https://archiv.ub.uni-heidelberg.de/volltextserverhttps://archiv.ub.uni-heidelberg.de/volltextserver/30776/1/Stefano_Secchia_Thesis_Final.pdf
identifier: DOI:10.11588/heidok.00030776
identifier: urn:nbn:de:bsz:16-heidok-307768
identifier:   Secchia, Stefano  (2022) Single-cell dissection of regulatory landscapes during embryogenesis.  [Dissertation]     
relation: https://archiv.ub.uni-heidelberg.de/volltextserver/30776/
rights: info:eu-repo/semantics/openAccess
rights: http://archiv.ub.uni-heidelberg.de/volltextserver/help/license_urhg.html
language: eng