Nucleic Acids Research, Vol 24, Issue 16 3149-3157, Copyright © 1996 by Oxford University Press
YC Wang and MT Hsu
To study the mechanism by which ultraviolet (UV) light inhibits DNA
replication, we examined the effects of UV 254 nm irradiation on the
replication of simian virus 40 (SV40) DNA and SV40-based plasmid in monkey
cells. The study was designed to determine the relative contributions made
by inhibition of replication initiation and chain elongation to the
immediate inhibition of DNA replication following UV irradiation. We used
two-dimensional neutral-alkaline electrophoresis to examine the behaviour
of replication intermediates unambiguously. Kinetic analysis using this
technique showed that initiation of replication started to decline at 15
min post-irradiation. When the pulse label incorporated in SV40 replication
intermediates before irradiation was chased for 1 h, most of the label was
found in mature Form I and II molecules. This indicated that replication
elongation took place on damaged template. We also used a transfection
technique to show that heavily irradiated plasmids replicated efficiently
in unirradiated transfected cells. By the transfection technique, we
observed that UV irradiation of host cells dose-dependently inhibited
replication of transfected non-irradiated plasmids, suggesting that the
inhibition of DNA replication is due to a global change in cellular
physiology induced by UV. This change was also apparent from poor staining
of the chromatin by fluorescent-DNA-binding dyes immediately after UV
irradiation of intact cells. We conclude that a significant fraction of
chain elongation proceeds on damaged templates and DNA replication during
the acute response of cells irradiated with UV is mainly controlled by the
inhibition of replication initiation.
ARTICLES
Inhibition of initiation of simian virus 40 DNA replication during acute response of cells irradiated by ultraviolet light
Institute of Toxicology, Chung Shan Medical and Dental College, Taichung, Taiwan, Republic of China.
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