COMPLEX FUNCTIONAL INTERACTIONS AT THE EARLY ENHANCER OF THE PQ STRAIN OF BK VIRUS

BK virus is a human papovavirus that latently infects a majority of th e world's population. There are more than 30 strains of the virus, mos t of which differ in the structure of the early enhancer region. The e nhancer of the progenitor strain, WW, from which the other strains can be derived, consists of four conserved DNA domains, P, Q, R; and S. R earrangement of the enhancer occurs upon passage in tissue culture and is thought to occur during virus replication. The strain under study, PQ, was selected upon passage of the Gardner strain (PPPQS) in the pe rmissive cell line, Vero. Mutational analysis of the entire enhancer r egion demonstrates the importance of five cia-acting sequences: DNA si tes B, C, and F, which have homology to the NF-1 protein binding seque nce; one purine rich motif designated A; and site D, which is similar to an SP-1 protein binding site. Two sites, B and C, appear to have a negative influence on gene activity. To study the functional interacti ons in more detail, promoter-enhancer constructions that contain diffe rent combinations of the five DNA sites linked to the chloramphenicol acetyltransferase gene were tested for early gene activity. The result s reveal that the proteins binding to the enhancer functionally cooper ate with each other. The effects of making mutations at the DNA sites are very similar to the effects of using excess enhancer DNA sequences to titrate the proteins that bind to the cis-acting DNA sites (in viv o competition). Moreover, the effects of changing the spacing between the DNA sites also demonstrate that there are cooperative interactions among the proteins that bind to the PQ strain enhancer. DNA sites B, C, and F ire clearly protected from DNase I digestion by Vero cell nuc lear proteins. In addition, mutation of each DNA site alters its sensi tivity to DNase I in the presence of Vero cell proteins. Interestingly , mutation of site B affects protein binding to site B as well as to s ites A, C, D, and F. These results suggest that cooperative functional and physical interactions occur at the early enhancer of the PQ strai n.