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Nucleic Acids Research, 1984, Vol. 12, No. 2 1077-1086
© 1984

MOLECULAR BIOLOGY

Adenovirus DNA synthesized in the presence of aphidicolin

Mieko Oguro1,+, Tadashi Yamashita2, Hiroyoshi Ariga3,§ and Hiroshi Nagano1,++,*

1Dept. Physiological Chemistry and Nutrition, Faculty of Medicine, Univ. Tokyo Hongo, Bunkyo-ku, Tokyo 113 2Dept. Microbiology, Daiichi College of Pharmaceutical Sciences Minami-ku, Fukuoka, 815 3Dept. Virology, Inst. Medical Science, Univ. Tokyo Minato-ku, Tokyo, 108, Japan

*To whom correspondence should be addressed

Received September 28, 1983. Revised November 28, 1983. Accepted November 28, 1983.

Adenovirus types 2 and 5 DNA synthesized in vivo and in vitro in the presence of aphidicolin were studied. Inhibition of adenoviral DNA synthesis by aphidicolin was only 70% even at a concentration of 30 ug/ml of aphidicolin, at which the cellular DNA synthesis was completely inhibited. When initiation of viral DNA synthesis was synchronized with hydroxyurea and labeled with [3H]thymidine for 60 mm, the viral DNA synthesized in the presence of 30 pg/ml of aphidicolin was not of full length (35 kb) but small (approximately 12 kb) by analysis of alkaline sucrose density gradient centrifugation. When initiation of the viral DNA synthesis was not synchronized, the viral DNAs ranging from full size to 12 kb were synthesized in the presence of aphidicolin, indicating that the nascent DNAs longer than about 12 kb can continue to elongate in the presence of aphidicolin. This 12 kb DNA was not derived from the degradation products of newly synthesized full size adenoviral DNA. The viral DNA synthesis was restored and the full size of adenoviral DNA was attained within 15 min following removal of aphidicolin. About 20% of the entire viral genome length from the 5' -end was not inhibited by aphidicolin, while the synthesis of interior fragments of the adenoviral DNA was markedly inhibited by aphidicolin, judging from the electrophoretic pattern on neutral agarose gel after digestion of DNA with Hind III. These results indicate that aphidicolin inhibits adenoviral DNA replication at the internal region located approximately 20–30% from both terminals.

+Present address: Department of Microbiology, Nihon University School of Medicine, Oyaguchi, Itabashi-ku, Tokyo 173, Japan

§Present address: Department of Microbiology, School of Medicine, Health Science Center, State University of New York, Stony Brook, NY 11794, USA

++Present address: Department of Biochemistry, Central Research Laboratories, Yamanouchi Pharmaceutical Co., Ltd., Azusawa, ltabashi-ku, Tokyo 174, Japan


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