GANP DNA primase associated with MCM3 and DNA synthesis

GC-associated DNA primase (GANP) contains two functionally important regions, the N-terminal RNA/DNA primase and C-terminal MCM3-associated/import domains, which might be involved in DNA replication. Here, we investigated the functions of these regions by transfection studies. In addition to the C-terminal MCM3-binding (Gmap80) region, the RNA/DNA primase region (Gp) possesses an additional MCM3-interaction activity that mediates its translocation between the nucleus and the cytoplasm. Gp with an N-terminal nuclear localization signal sequence (NLS) is present in the nucleus, but Gmap80 with a C-terminal NLS appeared in the cytoplasm. The whole GANP (Ganp) protein is present in both nuclear and cytoplasmic compartments, in a Crm1-dependent manner. Gp associated with MCM3 through the NLS, and MCM3 induced nuclear localization of Ganp. NLS-negative MCM3 does not have such activity. These results suggest that the NLS of MCM3 is involved in nuclear translocation of Ganp. The nuclear-cytoplasm MCM3 translocation was demonstrated in mammalian cells, and over-expression of Ganp, but not of Gp or Gmap80, facilitated this translocation of MCM3 in co-transfectants. Moreover, introduction of Ganp induced DNA synthesis in the transfectants but dominant-negative mutants did not. It is concluded that GANP is translocated from the nucleus to the cytoplasm in association with MCM3 and is involved in DNA synthesis.