New method reveals positions of gene-regulating nucleosomes
David E. Fisher, MD, PhD
Dana-Farber Cancer Institute researchers have developed a powerful
method for charting the positions of key gene-regulating molecules
called nucleosomes throughout the human genome. The mapping tool could
help uncover important clues for understanding and diagnosing cancer and
other diseases, the scientists say. Moreover, it may shed light on the
role of nucleosomes in the process of "reprogramming" an adult cell to
its original embryonic state, which is a critical operation in cloning.
David E. Fisher, MD, PhD, and his Dana-Farber colleagues describe
their findings in Nature Biotechnology, which has published the paper as
an advanced online publication on its Web site.
"This study presents the first global view of human nucleosome
positioning," said Fisher. Although analyses of this type had been
pioneered in simpler organisms such as yeast, those approaches were not
feasible when applied to the massively larger and more complex human
The new technique "provides major new clues to the locations of many
hallmark features of the human genome, such as where transcription
factors bind, where transcription begins and possibly ends, and where
there are other biologically important structural features," said
Fisher. Transcription factors are proteins that bind to particular DNA
sites in "promoter" regions of genes and turn the genes on or off.
The novel method employed gene microarray studies followed by
sophisticated computational data analysis to pin down the nucleosome
locations. The paper describes how the scientists used the technique to
locate nucleosomes in 3,692 promoters (regions of DNA that interact with
regulatory factors to control gene activity) within seven human cell
lines, including malignant melanoma.
Nucleosomes are spherical packing units for DNA. They consist of a
length of DNA wrapped around a core, like ribbon around a spool that is
made up of proteins called histones, and the nucleosomes are located
along the chromosomes like beads on a string.
Nucleosomes have multiple functions, including allowing several feet
of DNA to be packed tightly into a cell's nucleus. They also regulate
gene expression, or activity, by determining whether DNA sequences can
be accessed by transcription factors, allowing the factors to regulate
expression of a nearby gene.
It has long been assumed that these factors can't bind to a stretch
of DNA that is bound by a nucleosome; they can, however, attach to DNA
"linkers" between two nucleosomes. With their new method, Fisher's group
found that transcription factor binding indeed typically occurs in the
"linker" regions in between nucleosomes, rather than in the DNA regions
that are tied up by the nucleosome complexes. These results suggest that
nucleosome positioning controls the turning on or off of genes, and the
nucleosomes can be relocated if cellular needs change.
Fisher, who heads Dana-Farber's Melanoma Program and is a professor
of pediatrics at Harvard Medical School, conducts basic research on gene
mutations involved in the deadly skin cancer. He said the work that led
to the current paper began when his lab was studying the protein, MITF,
made by one of these genes, which influences the expression of other
"We wanted to understand how MITF regulates target genes, and
specifically where in promoter regions of those genes does MITF bind,"
Fisher said. "In the process of this work, we asked whether MITF is
binding between nucleosomes or on top of nucleosomes, and that led us to devise a method to ask where the nucleosomes are located."
The researchers used gene microarrays to which DNA associated with
single nucleosomes was added. The nucleosomal DNA was derived from
several cancer cell lines including melanoma and breast cancer, as well
as several normal human cell types. A critical component of the analysis
involved processing the data using computational algorithms devised by
Dana-Farber faculty member X. Shirley Liu, PhD, and her postdoctoral
fellow Jun S. Song, PhD, working closely with Fisher's graduate student
Fatih Ozsolak. The researchers borrowed a computational technique from
signal processing called "wavelet denoising" that revealed the striking
patterns of positioned nucleosomes.
On a graph, the data displayed as a series of peaks and troughs
corresponding to positioned nucleosomes and nucleosome-free regions,
respectively. Analysis revealed that promoters of genes that had similar
expression status (they were all "off" or "on") had related nucleosome
locations. The technique successfully pinpointed the location of some
nucleosomes previously found through other means, but the new method can
be applied to the entire genomes of human and other cells, said the
In the future, the techniques might be useful as a diagnostic tool, Fisher said.
"Gene expression is important in all diseases," he explained. "And
this method offers a new perspective for looking at how gene expression
has been altered in human diseases."
The lead authors of the report are Ozsolak and Song.
The research was supported by a grant from the National Institutes of
Health and a Claudia Adams Barr Award for Innovative Basic Cancer