Inhibition of DNA topoisomerase I activity by 2',5'-oligoadenylates and mismatched double-stranded RNA in uninfected and HIV-1-infected H9 cells.
2',5'-Oligoadenylates (2-5As) inhibit the type I DNA topoisomerase activity both in uninfected and HIV-1-infected human T cell line H9 as well as the purified enzyme (calf thymus). Topoisomerase I activity was determined by measuring the relaxation of negatively supercoiled pBR322 DNA. Inhibition of topoisomerase I by 2-5A depends on the chain length of the oligomer and the presence of 5'-phosphate. The 5'-triphosphate of the 2-5A hexamer was most active (almost total inhibition of DNA relaxation at 10 microM concentration); the 2-5A core trimer (at 100 microM) displayed no significant effect. In crosslinking and immunoprecipitation experiments we present evidence that 2-5A (32P-labelled 2-5A derivative, ppp(A2'p)3 A[32P]pCp) is able to bind to nuclear topoisomerase I. The mismatched dsRNA, poly(I).poly(C12U) (Ampligen), exhibited a strong anti-HIV-1 activity. However, our data show that this antiviral effect is not related to topoisomerase I inhibition. On the other hand, we did observe the production of longer oligomers of 2-5A in cells treated with poly(I).poly(C12U). It remains speculative, whether the in vivo effect could be catalyzed by this activity of poly(I).poly(C12U). In addition we could show that 2-5A also inhibits topoisomerase I activity associated with isolated HIV-1 particles.
Schröder
H C
HC
Abteilung Angewandte Molekularbiologie, Johannes Gutenberg-Universität, Mainz, Germany.
Kelve
M
M
Schäcke
H
H
Pfleiderer
W
W
Charubala
R
R
Suhadolnik
R J
RJ
Müller
W E
WE
D015088:2',5'-Oligoadenylate Synthetase / Q000378:metabolism; D000227:Adenine Nucleotides / Q000096:biosynthesis / Q000378:metabolism / Q000494:pharmacology*; D000818:Animals; D000998:Antiviral Agents / Q000494:pharmacology; D002417:Cattle; D002460:Cell Line; D004264:DNA Topoisomerases, Type I / Q000378:metabolism; D015658:HIV Infections / Q000188:drug therapy / Q000201:enzymology*; D015497:HIV-1 / Q000187:drug effects / Q000201:enzymology; D006801:Humans; D009843:Oligoribonucleotides / Q000096:biosynthesis / Q000378:metabolism / Q000494:pharmacology*; D011070:Poly I-C / Q000494:pharmacology*; D011072:Poly U / Q000494:pharmacology*; D012330:RNA, Double-Stranded / Q000494:pharmacology; D059004:Topoisomerase I Inhibitors
D000227:Adenine Nucleotides; D000998:Antiviral Agents; D009843:Oligoribonucleotides; D012330:RNA, Double-Stranded; D059004:Topoisomerase I Inhibitors; D011072:Poly U; C023505:2',5'-oligoadenylate; C047490:poly(I).poly(c12,U); D015088:2',5'-Oligoadenylate Synthetase; D004264:DNA Topoisomerases, Type I; D011070:Poly I-C