DNA topoisomerase II cleavage of telomeres in vitro and in vivo.
In this work, we have analyzed the reactivity of DNA topoisomerase II with telomeric DNA both in vitro and in vivo. Topoisomerase II cleavage reactions were performed on the tandem repeats of telomeric DNA. Analysis of this DNA on sequencing gels revealed that DNA topoisomerase II is catalytically active in cleaving the telomere DNA repeat. The topoisomerase II cleavage site is 5'TTAGG*G3' (cleavage site marked by the asterisk) and since telomere DNA is a tandem array of the above sequence, topoisomerase cleavage sites could exist every six base pairs. Detection of topoisomerase II cleavages was strongly dependent upon one specific topoisomerase II poison, etoposide (VP-16). A number of other topoisomerase II poisons were tested but did not stimulate cleavage activity at the telomere repeat. We have also analyzed the association of endogenous topoisomerase II with chromosomal telomeric DNA in HeLa cells. The in vivo complex of enzyme (ICE) bioassay was used to isolate topoisomerase II-DNA covalent complexes. In consistence with in vitro cleavage data, endogenous topoisomerase II-telomeric DNA complexes were detected in only etoposide-treated HeLa cells.
Yoon
H J
HJ
Department of Biology, Yonsei University, Seoul, South Korea.
Choi
I Y
IY
Kang
M R
MR
Kim
S S
SS
Muller
M T
MT
Spitzner
J R
JR
Chung
I K
IK
D004247:DNA / Q000235:genetics / Q000378:metabolism*; D004250:DNA Topoisomerases, Type II / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005047:Etoposide / Q000494:pharmacology; D006367:HeLa Cells; D006801:Humans; D012091:Repetitive Sequences, Nucleic Acid; D017422:Sequence Analysis, DNA; D016615:Telomere / Q000378:metabolism*; D059005:Topoisomerase II Inhibitors
D004791:Enzyme Inhibitors; D059005:Topoisomerase II Inhibitors; D005047:Etoposide; D004247:DNA; D004250:DNA Topoisomerases, Type II