Chromatin Remodeling Proteins

By Tara Wirsching

What is a Chromatin Remodeling Protein?

First, chromatin is a string of nucleosomes that are DNA wound up onto histone protein octamers.
The histone octamers can be moved up and down the DNA (called sliding) to either decrease or increase
the amount of DNA between the nucleosomes. This DNA that is between nucleosomes is called linker
DNA and is the only DNA that can be accessed by other proteins and therefore can be transcribed.
However, the linker DNA is also susceptible to damage and may even be targeted by proteases. A
chromatin remodeling protein is a protein that is able to change chromatin structure by moving
nucleosomes, adding nucleosomes, or removing the histone octamer of the nucleosome to remove the
nucleosome. To be able to preform their function, Chromatin remodelers have domains that can recognize
DNA sequences. These domains are known as nuclease domains since they can read the DNA sequence.
Chromatin remodelers are also able to identify histones and non-nucleosome proteins. This can be done
through a bromodomain or other similar domains. Since chromatin remodeling take energy, chromatin
remodelers also have ATPase domains that allow them to hydrolyze ATP and use the energy from this
reaction to drive the process of chromatin remodeling[ CITATION Bec13 \l 1033 ].

Fun30
Fun30 is a chromatin remodeling protein found in yeast and with homologs in other eukaryotes.
Fun30 stands for Function Unknown now 30. From the genetic sequence, it was predicted that Fun30 has
a Snf2 domain. Due to presence of the subunit SNf2, it was predicted that Fun30 would also have an
ATPase domain and likely was able to function as a chromatin remodeling protein. Fun 30 also contains a
domain similar to the Cue domain that is known in other proteins to allow proteins to preferentially bind
to ubiquitinylated histone proteins. Salma Awad and their team conducted biochemical experiments to
learn if Fun30 was able to function as a Chromatin remodeling protein.

In regards to Fun30’s structure, it was found that Fun30 consistently weighed 250kDa, larger than
the size indicated from the SDS page and silver staining which indicated a size of 128kDa[ CITATION
Awa10 \l 1033 ]. From this information it was predicted that Fun30 exists in a monomeric form that is in
equilibrium with its dimeric form, causing it to constantly be assembled and disassembled with the
concentrations of the monomeric and dimeric form remaining constant. This was confirmed through
tagging Fun30 with a histidine (Fun30-His6). Fun30-His6, binds to Ni2+ beads while Fun30 does not.
When Fun30 and Fun30-His6 were mixed together and then mixed with Ni2+ beads, it was found that both
forms of the protein were isolated by the nickel beads. This confirmed that Fun30 directly interacts with
itself and form a homodimer.

From the research, it was found that Fun30 can bind to DNA, nucleosomes, and hydrolyze ATP.
It was also found that Fun30 is better at exchanging out histones than moving histones. In the in vivo
experiments, using E. coli bacteria it was found that Fun30 preferentially exchange histones and targets
the histone protein variant Htz1. Fun30 did not show preferential binding to ubiquitylated histones, likely
due to the differences in sequences observed in the Cue domain.
 Figure 6

This figure shows the gel images of the

experiment looking at Fun30’s and
RSC’s ability to slide nucleosomes along
a piece of DNA and their ability to
exchange nucleosomes. RSC is a known
chromatin remodeling protein and was
used as a positive control. For all
treatments A, B, and C exchange of the
nucleosomes was detected from the
proteins Fun30 and RSC. Treatments A,
B, and C test under which conditions
Fun30 can slide nucleosomes. For each
of the different treatments, different
amounts of linker DNA were present
around the nucleosome as indicated by
the top line of image. If the nucleosome
was repositioned, it would able to be
digested by enzymes and appears in the
gel in the row indicated by the “0W0
Acceptor”. In addition, bands would
appear in the row marked “repositioned
nucleosomes”. In Figure6A, both Fun30
and RSC move the nucleosome since
they appear as bands at eh 0W0 acceptor.
The lack of repositioned chromosomes
in Figure 6B shows that Fun30 requires
DNA segments on either side of the
nucleosome to reposition the
nucleosome. For all treatments, Fun30
shows less repositioning activity than
RSC, indicating that Fun30 is less
efficient at repositioning nucleosome
than exchanging histones.

Image from [CITATION Awa10 \p 9482 \l 1033 ]

Take away
The research into Fun30 adds to the growing body of information about the interactions and function
between proteins and chromatin and how DNA nucleotide sequences correlates to gene function. This
information can be used to better predict protein function of genes currently labeled hypothetical or with
unknown functions. Biochemical and biological experiments are still needed to build evidence that
support or do not support predicted functions and structures from DNA sequences, however better
predictions can speed up the process of devising experimental tests since they provide clues to what are
the possible functions and roles of the protein.

Works Cited
Awad, S., Ryan, D., Prochasson, P., Owen-Hughes, T., & Hassan, A. H. (2010). The Snf2 Homolog
Fun30 Acts as a Homodimeric ATP-dependent Chromatin-remodeling Enzyme. Journal of
Biological Chemistry, 285(13), 9477-9484. doi:DOI 10.1074/jbc.M109.082149

Becker, P. B., & Workman, J. L. (2013). Nucleosome Remodeling and Epigenetics. Cold Spring Harbor
Perspectives in Biology, 5(9). doi: 10.1101/cshperspect.a017905