Stimulated activity of human topoisomerases IIalpha and IIbeta on RNA-containing substrates.
Eukaryotic topoisomerase II is a dimeric nuclear enzyme essential for DNA metabolism and chromosome dynamics. Central to the activities of the enzyme is its ability to introduce transient double-stranded breaks in the DNA helix, where the two subunits of the enzyme become covalently attached to the generated 5'-ends through phosphotyrosine linkages. Here, we demonstrate that human topoisomerases IIalpha and IIbeta are able to cleave ribonucleotide-containing substrates. With suicide substrates, which are partially double-stranded molecules containing a 5'-recessed strand, cleavage of both strands was stimulated approximately 8-fold when a ribonucleotide rather than a deoxyribonucleotide was present at the scissile phosphodiester of the recessed strand. The existence of a ribonucleotide at the same position in a normal duplex substrate also enhanced topoisomerase II-mediated cleavage, although to a lesser extent. The enzyme covalently linked to the 5'-ribonucleotide in the cleavage complex efficiently performed ligation, and ligation occurred equally well to acceptor molecules terminated by either a 3'-ribo- or deoxyribonucleotide. Besides the enhanced topoisomerase II-mediated cleavage of ribonucleotide-containing substrates, cleavage of such substrates could be further stimulated by ATP or antitumor drugs. In conclusion, the observed in vitro activities of the human topoisomerase II isoforms indicate that the enzymes can operate on RNA or RNA-containing substrates and thus might possess an intrinsic RNA topoisomerase activity, as has previously been demonstrated for Escherichia coli topoisomerase III.
Wang
Y
Y
Department of Molecular and Structural Biology, University of Aarhus, 8000 Arhus C, Denmark.
Knudsen
B R
BR
Bjergbaek
L
L
Westergaard
O
O
Andersen
A H
AH
D000255:Adenosine Triphosphate / Q000494:pharmacology; D000951:Antigens, Neoplasm; D000970:Antineoplastic Agents / Q000494:pharmacology; D004247:DNA / Q000378:metabolism*; D004250:DNA Topoisomerases, Type II / Q000378:metabolism*; D004268:DNA-Binding Proteins; D005057:Eukaryotic Cells / Q000201:enzymology; D006801:Humans; D007527:Isoenzymes / Q000378:metabolism*; D007535:Isomerases / Q000378:metabolism; D008957:Models, Genetic; D010755:Organophosphates / Q000378:metabolism; D011485:Protein Binding; D012313:RNA / Q000378:metabolism*; D012265:Ribonucleotides / Q000378:metabolism*; D013379:Substrate Specificity
D000951:Antigens, Neoplasm; D000970:Antineoplastic Agents; D004268:DNA-Binding Proteins; D007527:Isoenzymes; D010755:Organophosphates; D012265:Ribonucleotides; D012313:RNA; D000255:Adenosine Triphosphate; D004247:DNA; D007535:Isomerases; D004250:DNA Topoisomerases, Type II