FIS CANNOT SUPPORT ORIC DNA-REPLICATION IN-VITRO

The effects of the histone like proteins, HU, IHF, and Fis on DNA repl ication have been examined in vitro using the oriC and pBR322 DNA repl ication systems. In the oriC system, although low concentrations of HU and IHF stimulated DNA replication, no stimulation was observed upon the addition of Fis. On the contrary, high concentrations of Fis inhib ited oriC DNA replication completely. The inhibitory effect of Fis cou ld be overcome when HU, IHF, or RNA polymerase were also present in th e reaction mixtures. Unwinding of duplex DNA at the origin correlated with the elevated level of DNA replication. In contrast, only inhibiti on, rather than stimulation, was observed when these proteins were add ed to the pBR322 DNA replication system. Thus, the observed dependence on Fis of oriC DNA replication in vivo cannot be attributed to a posi tive effect on localized DNA unwinding.