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DNA replication in mammalian cells occurs in discrete nuclear foci. Here we show that terminally dlfferentiated myotubes can be induced to reenter S phase and show the same pattern of replication foci as cycling cells. We used this cellular system to analyze the interaction of cell cycle proteins with these foci in vivo. Cyclin A and cdk2, but not cyclln Bl and cdcl, were speclflcally localized at nuclear replication foci, just like the repllcatlon protein proliferating cell nuclear antigen. A potential target of cyclin A and cdk2 Is the 34 kd subunit of replication protein A (RPA34). In contrast with the 70 kd subunit, which localizes to the foci, RPA34 was not detected at these replication sites, which may reflect a transient interaction. The specific localization of cyclln A and cdk2 at nuclear replication foci provides adirect link between cell cycle regulation and DNA replication.