Selective degradation of reverse gyrase and DNA fragmentation induced by alkylating agent in the archaeon Sulfolobus solfataricus.
Reverse gyrase is a peculiar DNA topoisomerase, specific of hyperthermophilic Archaea and Bacteria, which has the unique ability of introducing positive supercoiling into DNA molecules. Although the function of the enzyme has not been established directly, it has been suggested to be involved in DNA protection and repair. We show here that the enzyme is degraded after treatment of Sulfolobus solfataricus cells with the alkylating agent MMS. MMS-induced reverse gyrase degradation is highly specific, since (i) neither hydroxyurea (HU) nor puromycin have a similar effect, and (ii) topoisomerase VI and two chromatin components are not degraded. Reverse gyrase degradation does not depend on protein synthesis. Experiments in vitro show that direct exposure of cell extracts to MMS does not induce reverse gyrase degradation; instead, extracts from MMS-treated cells contain some factor(s) able to degrade the enzyme in extracts from control cells. In vitro, degradation is blocked by incubation with divalent metal chelators, suggesting that reverse gyrase is selectively degraded by a metal-dependent protease in MMS-treated cells. In addition, we find a striking concurrence of extensive genomic DNA degradation and reverse gyrase loss in MMS-treated cells. These results support the hypothesis that reverse gyrase plays an essential role in DNA thermoprotection and repair in hyperthermophilic organisms.
Valenti
Anna
A
Institute of Protein Biochemistry, Consiglio Nazionale delle Ricerche, Via P. Castellino 111, 80131 Naples, Italy.
Napoli
Alessandra
A
Ferrara
Maria Carmina
MC
Nadal
Marc
M
Rossi
Mosè
M
Ciaramella
Maria
M
D000477:Alkylating Agents / Q000633:toxicity*; D019843:Archaeal Proteins / Q000378:metabolism; D053938:DNA Fragmentation; D004264:DNA Topoisomerases, Type I / Q000378:metabolism*; D019641:DNA, Archaeal / Q000737:chemistry; D006918:Hydroxyurea / Q000633:toxicity; D045726:Metalloproteases / Q000378:metabolism; D008741:Methyl Methanesulfonate / Q000633:toxicity*; D048229:Sulfolobus solfataricus / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics
D000477:Alkylating Agents; D019843:Archaeal Proteins; D019641:DNA, Archaeal; D008741:Methyl Methanesulfonate; D045726:Metalloproteases; C041813:DNA reverse gyrase; D004264:DNA Topoisomerases, Type I; D006918:Hydroxyurea