TOPOISOMERASE-I ENHANCES TFIID-TFIIA COMPLEX ASSEMBLY DURING ACTIVATION OF TRANSCRIPTION

The mechanism of coactivation by DNA topoisomerase I(topo I) was exami ned in a highly defined in vitro transcription system containing Pol I I and purified factors. Both stimulation of the basal reaction and coa ctivation occurred dependent on TAF(II)s. Activation was first observe d at the TFIID-TFIIA stage of initiation and maximal activation requir ed the concomitant presence of TFIID, TFIIA, topo I, and activator. El ectrophoretic mobility shift assay demonstrated a dramatic enhancement in the formation of the TFIID-TFIIA complex by topo I and activator, dependent on the TAF(II)s. DNase I footprinting confirmed this recruit ment. A catalytically inactive topo I, which coactivated transcription , similarly stimulated the rapid formation of the TFIID-TFIIA complex in the presence of activator. A camptothecin-mediated DNA cleavage ass ay demonstrated the recruitment of topo I to the template by TFIID. To po I likely functions during activation by enhancing the formation of an active TFIID-TFIIA complex on the promoter.