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. (C) 2003 Published by Elsevier B.V.