The DNA replication timing program and 3D nuclear organisation

In eukaryotic cells, DNA replication is initiated at multiple sites along the genome called origins of replication. Activation of replication origins is temporally and spatially regulated where some origins are fired at the beginning while others later during S-phase2. The temporal control of origin activation corresponds to a spatial distribution of the origins within the nucleus, with the early firing located in the nuclear centre and the origins activated later at the periphery3. The replication-timing program determines the order of origin activation and is set up over large chromosomal regions in early G1-phase, at a point termed the timing decision point (TDP). As the cell enters S-phase, this temporal program is executed locally at origins of replication at the time of their activation. In yeast it has been shown that the initiation of origin firing relies on the recruitment of replication factors, some of which exists in limiting amounts (e.g. Cdc45). The affinity of each origin competing for those same factors determines the probability, and thus the order of firing.

RADs and LADs

The Buonomo laboratory recently showed that Rif1 coats large chromosomal regions that coincide with late replicating domains9. In addition, these Rif1 associated domains (RADs) largely overlap with the Lamina associated domains (LADs) and between them, Rif1 and Lamin B1 coat 73% of all the late replicating regions9. Nuclear lamins, subdivided into A-type and B-type, and are essential components of the nuclear architecture and are important for development and differentiation. The LADs can further be divided into constitutive LADs (cLADs) and facultative LADs (fLADs). Whereas the cLADs show a high frequency binding to the lamina with a low cell-to-cell variation, the fLADs have a lower frequency of lamina binding with a high cell-to-cell variation. In addition, the cLADs show lamina association independent on cell type while the fLADs show a variability from cell type to cell type.

X chromosome inactivation as a model for studying 3D chromatin organization

The Buonomo laboratory have recently discovered that loss of Rif1 leads to lethality in female mouse embryos as a result of defective random X chromosome inactivation (unpublished). X chromosome inactivation (XCI) is the mechanism by which female mammalian cells achieve dosage compensation of X-linked gene expression. This process occurs early in murine development in two waves, first an imprinted XCI which leads to inactivation of the paternal inherited X chromosome, the inactive X is activated again, to then go through a second wave of XCI. This second wave is random with respect to the parental origin of the X chromosome.