COMPETITION FOR DNA-BINDING SITES BETWEEN THE SHORT AND LONG FORMS OFE2 DIMERS UNDERLIES REPRESSION IN BOVINE PAPILLOMAVIRUS TYPE-1 DNA-REPLICATION CONTROL

Papillomaviruses establish a long-term latency in vivo by maintaining their genomes as nuclear plasmids in proliferating cells. Bovine papil lomavirus type 1 encodes two proteins required for viral DNA replicati on: the helicase El and the positive regulator E2. The homodimeric E2 is known to cooperatively bind to DNA with El to form a preinitiation complex at the origin of DNA replication. The virus also codes for two short forms of E2 that can repress viral functions when overexpressed , and at least one copy of the repressor is required for stable plasmi d maintenance in transformed cells. Employing a tetracycline-regulated system to control El and E2 production from integrated loci, we show that the short form of E2 negatively regulates DNA replication. We als o found that the short form could repress replication in a cell-free r eplication system and that the repression requires the DNA binding dom ain of the protein. In contrast, heterodimers of the short and long fo rms were activators and, by footprint analysis, were shown to be as po tent as homodimeric E2 in loading El to its cognate site. DNA binding studies show that when El levels are low and are dependent upon E2 for occupancy of the origin site, the repressor can block EI-DNA interact ions, We conclude that DNA replication modulation results from competi tion between the different forms of E2 for DNA binding. Given that het erodimers are active and that the repressor form of E2 shows little co operativity with El for DNA binding, this protein is a weak repressor.