DNA-LIGASE-I IS RECRUITED TO SITES OF DNA-REPLICATION BY AN INTERACTION WITH PROLIFERATING CELL NUCLEAR ANTIGEN - IDENTIFICATION OF A COMMON TARGETING MECHANISM FOR THE ASSEMBLY OF REPLICATION FACTORIES

In mammalian cells, DNA replication occurs at discrete nuclear sites t ermed replication factories. Here me demonstrate that DNA ligase I and the large subunit of replication factor C (RF-C p140) have a homologo us sequence of similar to 20 amino acids at their N-termini that funct ions as a replication factory targeting sequence (RFTS). This motif co nsists of two boxes: box 1 contains the sequence IxxFF whereas box 2 i s rich in positively charged residues. N-terminal fragments of DNA lig ase I and the RF-C large subunit that contain the RFTS both interact w ith proliferating cell nuclear antigen (PCNA) in vitro. Moreover, the RFTS of DNA ligase I and of the RF-C large subunit is necessary and su fficient for the interaction with PCNA. Both subnuclear targeting and PCNA binding by the DNA ligase I RFTS are abolished by replacement of the adjacent phenylalanine residues within box 1. Since sequences simi lar to the RFTS/PCNA-binding moth have been identified in other DNA re plication enzymes and in p21(CIP1/WAF1), we propose that, in addition to functioning as a DNA polymerase processivity factor, PCNA plays a c entral role in the recruitment and stable association of DNA replicati on proteins at replication factories.