The replication kinase Cdc7‐Dbf4 promotes the interaction of the p150 subunit of chromatin assembly factor 1 with proliferating cell nuclear antigen

Abstract

The coordination of chromatin assembly with DNA replication, which is essential for genomic stability, requires the combined activation of histone deposition with the firing of replication origins. We report here the direct interaction of chromatin assembly factor 1 (CAF1), a key factor involved in histone deposition, with the replication kinase Cdc7-Dbf4. We isolated a complex containing both the largest subunit of CAF1 (p150) and the Cdc7-Dbf4 kinase specifically in S phase and thus prove the existence of this interaction in vivo. We then show that the Cdc7-Dbf4 kinase efficiently phosphorylates p150. This event induces a change in p150 oligomerization state, which promotes binding to proliferating cell nuclear antigen (PCNA). Conversely, CAF1 recruitment is reduced in a PCNA/DNA loading assay using Cdc7-depleted extracts. Our data define p150 as a new target for this kinase with implications for the coordination between DNA replication and CAF1 functions.

Cdc7 Is an Active Kinase in Human Cancer Cells Undergoing Replication Stress

“…It is, however, possible that Cdc7 may regulate some subtle events that occur when replication forks are arrested, such as fine-tuning of MCM helicase activity or some protein-protein interactions at the replisome. In this respect it was recently reported that the Cdc7⅐Dbf4 kinase also phosphorylates the p150 subunit of the chromatin assembly factor 1, thus promoting its binding to proliferating cell nuclear antigen (50). It is likely that an active Cdc7 kinase may also be required to control chromatin assembly factor 1 activity and histone deposition at delayed replication forks in etoposide and HU.…”

CDC28 phosphorylates Cac1p and regulates the association of chromatin assembly factor i with chromatin

“…6 The persistence of Cac1p phosphorylation in cdc7-1 cells (Fig. 1B and D) raised the possibility that, unlike its human counterpart, the budding yeast Cac1p is not phosphorylated by DDK.…”

“…2 It is believed that CAF-I is recruited to replication forks via contacts with PCNA (Proliferating Cell Nuclear Antigen, POL30), the DNA replication sliding clamp. [3][4][5][6] Mutations in POL30 or CAC1 (which encodes the largest subunit of CAF-I) that cripple their interaction in vitro also impair the assembly of chromatin in vitro 3,7 and show gene silencing defects in vivo. [7][8][9][10] However, the mechanisms that regulate the association of CAF-I with PCNA and with chromatin are poorly understood.…”

“…12 Another in vitro experiment has shown that the phosphorylation of the largest subunit of the human CAF-I (p150) by the Cdc7-Dbf4 kinase, but not by CDK2, promoted its binding to PCNA. 6 It remains unclear if Cdc7-Dbf4 regulates the association of CAF-I with chromatin and PCNA in vivo. Therefore, the precise regulation of CAF-I by protein kinases remains poorly understood.…”

Inhibition of DNA Damage-induced Apoptosis through Cdc7-mediated Stabilization of Tob

“…This characteristic of HMGB1 and its ability to interact with DNA repair proteins and chromatin remodeling factors render the damaged DNA more accessible to the DNA repair apparatus (27,28). Cdc7 is also likely to contribute to chromatin remodeling because Cdc7 aids the recruitment of chromatin assembly factor 1 to chromatin by promoting the interaction between chromatin assembly factor 1 and proliferating cell nuclear antigen, the homotrimeric processivity factor for DNA polymerase (31). Intriguingly, we found that HMGB1 functions as an upstream regulator of Cdc7 to suppress DNA damage-induced apoptosis (Fig.…”