INTERACTION BETWEEN THE N-TERMINUS OF HUMAN TOPOISOMERASE-I AND SV40 LARGE T-ANTIGEN

We have attempted to identify human topoisomerase I-binding proteins i n order to gain information regarding the cellular roles of this prote in and the cytotoxic mechanisms of the anticancer drug camptothecin, w hich specifically targets topoisomerase I. In the course of this work we identified an interaction between the N-terminus of human topoisome rase I and the SV40 T antigen that is detectable in vitro using both a ffinity chromatography and co-immunoprecipitation. Additional results indicate that this interaction does not require intermediary DNA or st oichiometric quantities of other proteins. Furthermore, the interactio n is detectable in vivo using a yeast two-hybrid assay. Two binding si tes for T antigen are apparent on the topoisomerase I protein: one con sisting of amino acids 1-139, the other present in the 383-765 region of the protein. Interestingly, nucleolin, which binds the 166-210 regi on of topoisomerase I, is able to bind an N-terminal fragment of topoi somerase I concurrently with T antigen. Taken together with our prior identification of nucleolin as a topoisomerase I-binding protein, the current results suggest that helicase-binding is a major role of the N -terminus of human topoisomerase I and that the resultant helicase-top oisomerase complex may function as a eukaryotic gyrase.