Evidence for the participation of topoisomerases I and II in cadmium-induced metallothionein expression in Chinese hamster ovary cells.
CHO-Cdr20 cells are 10-20 times more resistant to killing by cadmium than the parental CHO cells. Resistance has been linked to amplification of the metallothionein genes MT-I and MT-II and their coordinate induction by cadmium and other toxic metals. We studied the roles of the nuclear enzymes topoisomerase I and topoisomerase II in Cd-induced expression of MT-II. Camptothecin-induced DNA strand breakage, mediated by topoisomerase I in cells, increased by approximately 20% when the resistant cells were incubated first with 50 microM Cd and then with camptothecin. Short DNA fragments were enriched in MT-II-hybridizing sequences, indicating that topoisomerase I-associated breakage was directed in part toward the location of induced gene activity. Ten microM camptothecin inhibited Cd-induced accumulation of MT-II mRNA as well as induced and uninduced RNA synthesis in the resistant cells. These data are consistent with the notion that topoisomerase I participates in most or all forms of RNA synthesis. Topoisomerase II inhibitors which trap cleavable complexes (amsacrine, VM-26, VP-16) increased DNA strand breakage at very high concentrations (50-100 microM); the increased breakage appeared to be concentrated near the MT-II gene. This class of inhibitor did not block the accumulation of MT-II message. Novobiocin, a second type of topoisomerase II inhibitor blocked transcription at 300 microM. Merbarone, a novel, third type of topoisomerase II inhibitor, blocked MT-II transcription at 50-100 microM. The latter two inhibited total RNA synthesis in induced, but not uninduced cells. Thus, it is possible that topoisomerase II plays more than one role in transcription and that more than one form of this enzyme is involved.
Mattern
M R
MR
Department of Molecular Pharmacology, Smith Kline & French Laboratories, Research & Development, King of Prussia, PA 19406-0939.
Tan
K B
KB
Zimmerman
J P
JP
Mong
S M
SM
Bartus
J O
JO
Hofmann
G A
GA
Drake
F H
FH
Johnson
R K
RK
Crooke
S T
ST
Mirabelli
C K
CK
D000818:Animals; D002104:Cadmium / Q000633:toxicity*; D019256:Cadmium Chloride; D002478:Cells, Cultured; D006224:Cricetinae; D003412:Cricetulus; D004249:DNA Damage; D015342:DNA Probes; D004264:DNA Topoisomerases, Type I / Q000235:genetics / Q000502:physiology*; D004250:DNA Topoisomerases, Type II / Q000235:genetics / Q000502:physiology*; D004277:DNA, Single-Stranded / Q000187:drug effects / Q000235:genetics; D004351:Drug Resistance / Q000235:genetics / Q000502:physiology; D005260:Female; D005786:Gene Expression Regulation / Q000187:drug effects* / Q000502:physiology; D015151:Immunoblotting; D008668:Metallothionein / Q000096:biosynthesis / Q000187:drug effects* / Q000235:genetics; D009693:Nucleic Acid Hybridization / Q000187:drug effects; D010053:Ovary / Q000187:drug effects* / Q000378:metabolism; D012313:RNA / Q000096:biosynthesis / Q000187:drug effects / Q000235:genetics; D059004:Topoisomerase I Inhibitors; D059005:Topoisomerase II Inhibitors; D014158:Transcription, Genetic / Q000187:drug effects / Q000502:physiology
D015342:DNA Probes; D004277:DNA, Single-Stranded; D059004:Topoisomerase I Inhibitors; D059005:Topoisomerase II Inhibitors; D002104:Cadmium; D012313:RNA; D008668:Metallothionein; D004264:DNA Topoisomerases, Type I; D004250:DNA Topoisomerases, Type II; D019256:Cadmium Chloride