Project Summary/Abstract
“Establishment and visualization of nuclear architecture
at the onset of mammalian development”
Nuclear organization is a key feature of our epigenome. The way in which our DNA is folded into the cell nucleus
can affect gene expression and determine cellular identity, but it can also regulate recombination and genome
instability. Indeed, the disruption of key elements underlying the architecture of the nucleus, such as
Topologically-Associated Domain (TAD) boundaries, has been shown to be the cause of genetically inherited
diseases in some instances.
How nuclear organization is first established after fertilization in mammals is largely unknown. In particular, the
lack of approaches to observe in real time the different layers of nuclear organization has been a barrier for the
full understanding of the dynamics and the impact of nuclear organization in vivo. Most importantly, the chromatin
and structural pathways that regulate the initial folding of our genome into the nucleus as well as the impact of
the resulting genome organization for key developmental and transcriptional processes in vivo, is largely
unknown.
We have recently shown that nuclear organization, particularly the association of specific genomic regions to the
nuclear lamina, is established de novo in the newly formed embryo immediately after fertilization. Lamina-
Associated Domains (LADs) precede the consolidation of TADs and compartments, suggesting that the
association with the nuclear lamina may be the first nuclear architectural scaffold and set the framework for
nuclear architecture.
Here, to fulfill the outstanding need for methodologies to visualize specific chromosome domains, we will
establish “LivePaint”, a technology to visualize the fundamental pillars of genome organization and their
dynamics in real time (Aim 1). We propose to image the process of establishment of nuclear architecture by
applying this tool to the mouse pre-implantation embryo, a full organism amenable to imaging and in which real
time dynamics can be studied in their natural context. In parallel, we will deploy low input genomic approaches,
such as DamID to identify the regulators of LAD establishment (Aim 2). Finally, using this information, we will
perform perturbations to establish the role of LADs in regulating the establishment of nuclear architecture through
both imaging and genomics approaches (Aim 3). We will also identify the impact of the disruption to the
establishment of nuclear organization in vivo on epigenetic features, such as replication timing and gene
expression and ultimately their impact on developmental competence.
Through the perturbation and live imaging of the main determinants of nuclear organization in vivo, this proposal
will establish the functional dependencies between the different layers of nuclear organization and will reveal
their role in cellular function, gene expression and developmental progression at the beginning of mammalian
development.
The knowledge generated through these Aims will address outstanding challenges for our understanding of how
the mammalian genome folds within the nuclear space in a physiological context and will reveal molecular
regulators of this process.
