Abstract
Heterochromatic regions render the replication process particularly difficult due to the high level of chromatin compaction and the presence of repeated DNA sequences. In humans, replication through pericentromeric heterochromatin requires the binding of a complex formed by the telomeric factor TRF2 and the helicase RTEL1 in order to relieve topological barriers blocking fork progression. Since TRF2 is known to bind the Origin Replication Complex (ORC), we hypothesized that this factor could also play a role at the replication origins (ORI) of these heterochromatin regions. By performing DNA combing analysis, we found that the ORI density is higher within pericentromeric satellite DNA repeats than within bulk genomic DNA and decreased upon TRF2 downregulation. Moreover, we showed that TRF2 and ORC2 interact in pericentromeric DNA, providing a mechanism by which TRF2 is involved in ORI activity. Altogether, our findings reveal an essential role for TRF2 in pericentromeric heterochromatin replication by regulating both replication initiation and elongation.
Highlights
Genomic DNA exhibits a higher-order nuclear organization that needs to be preserved to ensure the adequate physiological functioning of the cell
TRF1 prevents ATR kinase signaling by recruiting the Bloom helicase (BLM) to facilitate lagging strand synthesis [5], while TRF2 prevents the accumulation of topological stress during replication by regulating the 50 exonuclease activity of Apollo [6] and by the recruitment of the BUB1-BUB3 complex, which in turn phosphorylates TRF1 [7]
These results show that TRF2 is required for the recruitment of Origin Replication Complex (ORC) in pericentromeric DNA
Summary
Genomic DNA exhibits a higher-order nuclear organization that needs to be preserved to ensure the adequate physiological functioning of the cell. This compartmentalized nuclear structure is associated with DNA epigenetic modification and several associated proteins that are packed into two major types, euchromatin and heterochromatin [1]. Heterochromatin regions replicate late during the S phase and this timely process is important for the fine regulation of the cell cycle homeostasis. Rif and Taz ensure the firing of late replication origins throughout the genome [2]. In Schizosaccharomyces pombe, Taz delays the firing of several late-replication origins and recruits Ccq to assemble heterochromatin [3,4]. TRF2 binds to satellite DNA II and III preferentially during the S phase and protects those regions against
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