between ORC binding and nucleosome turnover,
suggesting that turnover facilitates ORC binding.
In contrast, other chromatin features that would
be expected for open or dynamic chromatin, in-
cluding nucleosome density, mononucleosome/
oligonucleosome ratio (a measure of micrococcal
nuclease accessibility), H2Av (an H2A.Z his-
tone variant enriched in active chromatin), and
salt-soluble nucleosomes, show little if any de-
pendence on ORC abundance (Fig. 3, H to P).
Our findings support the hypothesis that repli-
cation origins are determined by chromatin, not
by sequence features (
). The better quan-
titative correspondence of ORC to CATCH-IT
data than to other chromatin measurements implies
that the ORC occupies DNA that is made acces-
sible by nucleosome turnover. In support of this
interpretation, we note that very similar corre-
spondences are seen when CATCH-IT data are
aligned with GAF sites (fig. S9) and that GAF
directs nucleosome turnover in vivo (
Our direct strategy for measuring the kinetics
of nucleosome turnover does not rely on trans-
genes or antibodies but rather uses native his-
tones and generic reagents. Thus, CATCH-IT
provides a general tool for studying activities
that influence nucleosome turnover. With use of
CATCH-IT, we found direct evidence that epige-
netic maintenance involves nucleosome turnover,
a process that erases histone modifications (
The fact that EZ is responsible for di- and tri-
methylation of H3K27, but the nucleosomes that
it modifies turn over faster than a cell cycle,
argues against proposals that histone modifica-
tions required for cellular memory themselves
transmit epigenetic information (
). Rather, by
simply increasing or decreasing accessibility of
DNA to sequence-specific binding proteins, regu-
lated nucleosome turnover may perpetuate active
or silent gene expression states and facilitate ini-
tiation of replication.
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