Figure 3
From: Coupling histone homeostasis to centromere integrity via the ubiquitin-proteasome system
Molecular behaviour of Ams2 throughout S phase. (A) Chromatin fractionation assay. Wild-type cells were arrested with HU (arrest) and released into the HU-free fresh medium for 15 min (release). Whole-cell extracts (W) were spun and separated into soluble supernatant (S) and chromatin-bound pellet (P). Western blotting was performed with anti-Ams2 antibody. (B) A speculative model for spatial and temporal regulation of Ams2 during S and G2 phase. Early replication origins are activated in the presence of HU (fire), whereas the firing of late origins is blocked. During this period, DDK has high kinase activity, and thereby Ams2 proteins become phosphorylated, although Ams2 is stable. When cells are released from HU arrest, phosphorylated Ams2 is still detected only on chromatin, in which the chromatin-free phosphorylated fraction may be degraded via SCFPof3. As DNA replication progresses further and completes (G2), SCFPof3 may be able to interact with chromatin-bound phosphorylated Ams2. (see text for details)