Summary
Covalently closed circular mitochondrial DNA (mt-DNA) from mouse fibroblasts (L-cells) forms a separate band from nicked circular or linear DNA molecules in ethidium bromide-cesium chloride buoyant density gradient analysis. The conversion of covalently closed circular mt-DNA to nicked circular or linear DNA may serve as a sensitive method of detecting DNA strand breaks induced by bleomycin (BLM). At a very high concentration BLM induces the conversion of closed circular mt-DNA to nicked circular or linear DNA, however even after 24 h of exposure to BLM at a high concentration a considerable amount of closed circular mt-DNA molecules remains intact.
Zusammenfassung
Mitochondriale Desoxyribonucleinsäure (mt-DNS) aus Mäusefibroblasten (L-Zellen) tritt vorwiegend in Form von ringförmigen, doppelsträngingen durch kovalente Bindung geschlossenen Molekülen auf, welche in der Ethidium-bromid-Cäsiumchlorid-Dichtegradientenanalyse eine von doppelsträngig linearer oder teilweise unterbrochener ringförmiger mt-DNS getrennte Bande bilden. Die Umwandlung der völlig geschlossenen ringförmigen mt-DNS in die teilweise oder ganz geöffnete Form ergibt eine sehr empfindliche Methode zum Nachweis von DNS-Strangbrüchen unter der Einwirkung von Bleomycin (BLM). BLM führt in hoher Dosierung (200 μg ml) zur Umwandlung von kovalent geschlossener ringförmiger mt-DNS in teilweise geöffnete oder lineare DNS Moleküle, jedoch bleibt selbst nach 24 stündiger BLM Einwirkung ein beträchtlicher Anteil an doppelsträngigen, kovalent geschlossenen, ringförmigen mt-DNS Molekülen intakt.
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References
Barranco,S.C, Humphrey,R.M.: The effects of bleomycin on survival and cell progression in Chinese hamster cells in vitro. Cancer Res. 31, 1218–1223 (1971)
Blum,R.H., Carter,S.K., Agre,K.: A clinical review of bleomycin — A new antineoplastic agent. Cancer 31, 903–913 (1973)
Byfield,J.E., Lee,Y.C., Tu,L., Kulhanian,F.: Molecular interactions of the combined effects of bleomycin and X-rays on mammalian cell survival. Cancer Res. 36, 1138–1143 (1976)
Clayton,D.A., Doda,J.N., Friedberg,E.C.: The absence of a pyrimidine dimer repair mechanism in mammalian mitochondria. Proc. nat. Acad. Sci. (Wash.) 71, 2777–2781 (1974)
Hittelman,W.N., Rao,P.N.: Bleomycin-induced damage in prematurely condensed chromosomes and its relationship to cell cycle progression in CHO cells. Cancer Res. 34, 3433–3439 (1974)
Katsumata,T., Watanabe,M., Takabe,Y., Terasima,T., Umezawa,H.: Cinemicrographic analysis of death in synchronously growing mouse l cells after exposure to bleomycin. Gann 64, 71–77 (1973)
Koch,J.: Introduction of “nicks” and “chops” into human mitochondrial DNA in vivo and in vitro. Eur. J. Biochem. 33, 98–103 (1973)
Madreiter,H., Mittermayer,C., Osieka,R.: 3H-thymidine incorporation into mitochondria of synchronized mouse fibroblasts. Beitr. Path. 145, 249–255 (1972)
Miyaki,M., Ono,T.: Inhibition of ligase reaction by bleomycin. J. Antibiot. (Tokyo) 24, 587–592 (1971)
Müller,W.E.G., Yamazaki,Z., Breter,H.J., Zahn,R.K.: Action of bleomycin on DNA and RNA. Eur. J. Biochem. 31, 518–525 (1972)
Pasupathy,K., Netrawali,M.S., Pradhan,D.S., Sreenivasan,A.: Repair of radiation-induced strand scissions in nuclear and mitochondrial DNAs in Tetrahymena pyriformis. Radiat. Res. 66, 147–157 (1976)
Potter,S.S., Newbold,J.E., Hutchison III,C.A., Edgell,M.H.: Specific cleavage analysis of mammalian mitochondrial DNA. Proc. nat. Acad. Sci. (Wash.) 72, 4496–4500 (1975)
Radloff,R., Bauer,W., Vinograd,J.: A dye-buoyant-density method for detection and isolation of closed circular duplex DNA: the closed circular DNA in HeLa cells. Proc. nat. Acad. Sci. (Wash.) 57, 1514–1521 (1967)
Robberson,D.L., Clayton,D.A.: Replication of mitochondrial DNA in mouse l cells and their thymidine kinase=derivatives: displacement replication on a covalently-closed circular template. Proc. nat. Acad. Sci. (Wash.) 69, 3810–3814 (1972)
Shirakawa,I., Azegami,M., Ishii,S., Umezawa,H.: Reaction of bleomycin with DNA strand scission of DNA in the absence of sulfhydryl or peroxide compounds. J. Antibiot. (Tokyo) 24, 761–766 (1971)
Twentyman,P.R., Bleehen,N.M.: The sensitivity of cells in exponential and stationary phases of growth to bleomycin and to 1,3-bis(2-chloroethyl)-1-nitrosourea. Br. J. Cancer 28, 500–507 (1973)
Watanabe,N., Takabe,Y., Katsumata,T., Terasima,T.: Effects of bleomycin on progression through the cell cycle of mouse l cells. Cancer Res. 34, 878–881 (1974)
Zöllner,E.J., Weinblum,D., Obermeier,J., Zahn,R.K.: Influence of UV irradiation, 4-nitroquinoline-1-oxide, methylmethanesulfonate, and bleomycin on the activity of an alkaline deoxyribonuclease from human lymphocytes. Exp. Cell Res. 99, 185–189 (1976)
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Supported by grant II B 8 — FA 5654 from the Landesamt für Forschung, Ministerium für Wissenschaft und Forschung des Landes Nordrhein-Westfalen
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Osieka, R., Madreiter, H. & Schmidt, C.G. The effect of bleomycin on mitochondrial DNA. Z. Krebsforsch. 88, 11–15 (1976). https://doi.org/10.1007/BF00284742
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DOI: https://doi.org/10.1007/BF00284742