Abstract
We determined the effects of chromatin structure on template accessibility to replication factors and used three different templates as substrates for simian virus 40 (SV40) DNA replication in vitro: native and salt-treated SV40 minichromosomes and protein-free SV40 DNA. Native minichromosomes contain histone H1 and numerous nonhistone proteins in addition to the core histones, whereas salt-treated minichromosomes carry essentially only core histones. We reasoned that the less densely packed salt-treated minichromosomes should be more effective replication templates due to their more extended configuration. However, contrary to this expectation, we found that native minichromosomes replicated with significantly higher efficiency than salt-treated minichromosomes, while protein-free DNA was most active as a replication template. The higher replication efficiency of native minichromosomes was due to two activities bound to the chromatin, which were identified as DNA topoisomerases I and II. By using chromatin substrates of different general configurations, we also showed that the overall chromatin structure determines accessibility to topoisomerases I and II and thereby the efficiency of replicative chain elongation.
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