IDENTIFICATION OF REGIONS WITHIN THE 4 SMALL SUBUNITS OF HUMAN REPLICATION FACTOR-C REQUIRED FOR COMPLEX-FORMATION AND DNA-REPLICATION

Replication factor C (RFC) and proliferating cell nuclear antigen (PCN A) are processivity factors for eukaryotic DNA polymerases delta and e psilon. RFC binds to a DNA primer end and loads PCNA onto DNA in an AT P-dependent reaction. The five RFC subunits p140, p40, p38, p37, and p 36, all of which are required to form the active RFC complex, share re gions of high homology including the defined RFC boxes II-VIII. RFC bo xes III and V constitute a putative ATP binding site, whereas the func tion of the other conserved boxes is unknown. To study the individual subunits in the RFC complex and the role of the RFC boxes, deletion mu tations were created in all subunits. Sequences close to the C terminu s of each of the small subunits are required for formation of the five subunit complex. A N-terminal region of the small subunits, containin g the RFC homology box II, plays a critical role in the function of th ese subunits, deletion of which reduces but does not abolish RFC activ ity in loading PCNA onto DNA and in supporting an RFC-dependent replic ation reaction. The N termini of p37 and p40, although highly homologo us, are not interchangeable, suggesting unique functions for the indiv idual subunits.