Investigating how RIF1 coordinates the timing of replication and the spatial distribution of late-replicating genomic regions
Item statusRestricted Access
Embargo end date23/02/2023
In eukaryotes, the genome is organised into domains which are replicated at different times during S-phase. The order of replication of these domains, known as the “replication-timing (RT) program”, is transmitted from one cell cycle to the next. It is cell-type specific and changes during cell fate transition. The RT program is closely related to the 3-dimensional (3D) nuclear organisation. The co-variation of the RT program and nuclear architecture during cell fate commitment and the coincidence of their re-establishment in early G1 phase have promoted the hypothesis that nuclear organisation might contribute to the establishment of the RT program, or vice versa. During my PhD, I have contributed to our recent published work showing that RIF1 serves as a hub to regulate both the RT program and nuclear architecture via, at least partially, RIF1-PP1 interaction. I have then focused on a particular aspect of RIF1-dependent nuclear organisation, namely understanding how and why RIF1, assembled in large domains called RIF1-associated domains (RADs), ensures the peripheral localisation of late-replicating genomic regions. We have identified two different types of RADs: RADs with strong association to LAMIN-B1(RADs-LBhigh) and RADs less frequently associated with LAMIN-B1(RADs-LBlow). While the former are more resistant to the loss of RIF1, the RADs-LBlow are more sensitive to RIF1 depletion, indicated by both advanced RT and internalisation within the nucleus. This suggests that RIF1 is involved in anchoring RADs-LBlow to the nuclear periphery. To test whether peripheral localisation could determine late RT, here, I present the establishment of tethering platforms to manipulate nuclear localisation, followed by the investigation of RT. The preliminary result suggests that forced nuclear peripheral position is capable of dictating late RT. Moreover, my work also provides preliminary data suggesting that RIF1 is also involved in regulating cell cycle dynamics of H3S10ph, dependent on RIF1-PP1 interaction, and that RIF1 depletion also causes reduction of HP1g association to some of the RADs. This leads to the hypothesis that both H3S10ph and HP1g could contribute to RIF1’s function in regulating nuclear organisation. Thus, this study provides further molecular understanding of the mechanisms that underly the connection between the RT program and nuclear organisation through RIF1.