Nucleosome Occupancy and DNA Sequence Content
When we examined histone occupancy with antibodies against core histone
H3 or histone H4, using genomic DNA as the reference channel, we found
a reduction of ~20% in histone occupancy in intergenic sequences relative
to genic sequences for the average gene. We were surprised to find that
differential enrichment of intergenic and genic regions also occurred
in control experiments lacking antibody. Nonetheless, approximately 40%
of yeast promoters do have lower levels of histones than their downstream
transcribed regions, even after the normalization by control experiments.
We found that reduced nucleosome occupancy of specific promoter sites
is generally associated with both transcription factor occupancy and the
presence of poly (dA:dT) sequences.
Composite profile of histone occupancy plotted against
relative poly A9 occurrence. Probes corresponding to all genes were weighted
according to the number of nearby A9 (or T9) tracts. The resulting occupancy
is relative to the maximum average occupancy.
Relative occupancy of intergenic regions bound by transcriptional
regulators (P <0.001) and containing matches to cognate consensus binding
specificities determined in Harbison et al (2004). Probes corresponding
to all promoter regions were weighted according to the number of nearby
regulator-bound binding sites. The resulting occupancy is relative to
the maximum average occupancy.
|To explore whether poly(dA:dT) sequences are
associated with reduced nucleosome occupancy, we examined the frequency
of their association with nucleosome-depleted promoter regions. We found
that the distribution of poly A9 sites (in black, above) is strongly anti-correlated
with nucleosome occupancy (in red, above), peaking within intergenic regions
and at the lowest levels throughout open reading frames. Approximately 77%
of poly A9 sites located within promoter regions coincide with sites of
minimal nucleosome occupancy. Furthermore, unlike binding sites for transcriptional
regulators, which tend to occur upstream of genic regions, poly A9 elements
occur frequently within downstream regions. This could explain why histone
minima occur frequently in intergenic regions that are downstream of two
convergently transcribed genes.
||We investigated whether the binding sites occupied by most
transcriptional regulators (Harbison et al., 2004 in black, above) are associated
with minimal nucleosome occupancy (in red, above). Of the nearly 2000 promoter
regions that are occupied at least by one transcriptional regulator at high
confidence (P value less than 0.001), 74% showed minimal nucleosome occupancy
in the vicinity of the transcription factor binding site. Furthermore, we
noted an association between promoter binding by multiple regulators and
reduced nucleosome occupancy. In agreement with a previous study (Bernstein
et al., 2004), we note that certain transcription factors (Rap1, Fhl1, Swi4)
were more likely to have strong associations with nucleosome minima (up
to a 90% overlap) than others.
These results strongly indicate that certain DNA sequences are associated
with regions of low nucleosome occupancy, although they do not show that
the presence of these sequences themselves are responsible for altered
nucleosome occupancy. In order to test this hypothesis, we created 5 strains
in which both poly(dA:dT) and transcription factor binding sites had been
removed singly or in combination. Surprisingly,
deletion of these sites failed to confer any changes on the relative occupancy
of nucleosomes, indicating that other sequences or mechanisms must contribute
to the control of nucleosome density.