Understanding of a
Chromosomal Domain in
Published, JBC Papers in Press, December 28, 2004,
Catherine A. Fox‡ and Kristopher H. McConnell
From the Department of Biomolecular Chemistry,
University of Wisconsin Medical School,
Madison, Wisconsin 53706
A few discrete portions of the Saccharomyces cerevisiae genome,
such as the chromosome ends within 5–10 kilobase pairs (kb) of
telomeres and the silent mating-type regions named HMR and
HML, are defined as transcriptionally silent because they cause
repression of an RNA polymerase II gene placed within them
(reviewed comprehensively in Ref. 1). S. cerevisiae silencing is a
form of transcriptional repression that is long range, occurring over
distances larger than a single gene. Silencing occurs in only certain
chromosomal regions and depends more on a gene’s position in the
genome than its promoter sequence. Another defining characteris-
tic is a stable inheritance pattern during cell division, indicating
that the repressive state is duplicated along with the underlying
DNA sequences during DNA replication. These features make
S. cerevisiae transcriptional silencing functionally akin to position-
effect-variegation in Drosophila melanogaster and X-chromosome in-
activation in female mammals (for recent reviews see Refs. 2 and 3).
It is now established that these functionally similar chromo-
somal phenomena share a requirement for a distinct chromatin
structure. The precise molecules involved in forming repressive
chromatin vary between organisms and even different chromo-
somal regions within an organism (for examples see review in Ref.
4). However all silent chromosomal domains contain particular
post-translational modifications of nucleosomes, relatively ordered
nucleosome positioning and specialized chromatin-binding proteins
that distinguish them from actively transcribed chromosomal do-
mains (reviewed in Ref. 5).
The transcriptionally silent HMR locus in S. cerevisiae serves as
an important model for the structure, assembly, and function of a
repressive chromatin domain. HMR and HML (HM loci; silent mat-
ing-type loci) are two regions of ?3 kb each that reside near opposite
ends of chromosome III in the S. cerevisiae genome (Fig. 1A) (6). Each
contains a transcriptionally repressed (i.e. silent) copy of one of the
two types of mating-type genes that control the sexual cycle of S. cer-
evisiae. The silent HM regions serve as a reservoir of mating-type
genes for the transcriptionally active mating-type locus, MAT, during
a process called mating-type switching (reviewed in Ref. 7).
Here we focus on how HMR (Fig. 1B), which shares fundamental
molecular requirements for silencing with HML, assembles into a
silent chromatin domain. Over the past 10 years the primarily
genetic focus of this field has shifted toward more molecular anal-
yses, and several remaining questions will benefit from biochemi-
cal approaches. It is now common to present transcriptional silenc-
ing in terms of a molecular model that we have adapted for this
discussion of HMR (8–10) (Figs. 2–4).
HMR Contains a DNA Element Necessary for the Stability
and Specificity of Silent Chromatin
The HMR-E silencer is a small ?150-bp DNA element required
for silent chromatin to form at HMR (Fig. 2). HMR-E contains
binding sites for three different sequence-specific DNA-binding
proteins, the origin recognition complex (ORC),1Rap1p, and Abf1p
(only ORC and Rap1p are shown in Fig. 2). These silencer-binding
proteins bound to the silencer DNA provide a unique protein-DNA
interaction surface (the silencer protein complex, SPC, in Fig. 3)
that is recognized by four specialized chromatin-binding proteins,
the Sir (silent information regulator) proteins.
All three silencer-binding proteins have additional non-silencing
functions when bound elsewhere in the genome. The ORC is a
6-subunit heteromeric protein complex that binds to each of the
?400-replication origins distributed throughout the yeast genome
and is essential for the initiation of DNA replication (reviewed in
Ref. 11). ORC functions in silencing only when bound to the HM
silencers. Rap1p has many functions in the genome including tran-
scription activation (12). Abf1p also functions in transcription ac-
tivation and, at one origin, in replication initiation (13, 14). The Sir
proteins are recruited into a stable silent chromatin structure at
only a few discrete domains in the genome such as HMR. An
important question is how the multifunctional silencer-binding
proteins work together in the more exclusive process of silencing.
Part of the answer comes from the distinct arrangement of
silencer-binding proteins at HMR-E. The close juxtaposition of
ORC and Rap1p binding sites, for example, is unique to silencers.
Considering only two of the Sir proteins, Sir1p and Sir4p, illus-
trates how this juxtaposition contributes to the stability and spec-
ificity of HMR silencing. The Sir4p physically contacts Rap1p (15),
whereas the Sir1p contacts ORC (16, 17). In addition, Sir1p and
Sir4p physically interact (16, 18). The unique arrangement of ORC
and Rap1p at the silencer allows these individually weak interac-
tions to occur simultaneously, contributing to a stable complex.
Several other Sir-Sir and Sir protein-SPC interactions work simi-
larly to contribute to HMR-E silencer function (reviewed in Ref. 19)
(Fig. 2). Indeed, the final complex is so stable that multiple inter-
action defects must be created in vivo simultaneously, through
combinations of specific mutations, to cause a detectable HMR
Distinct interactions between the silencer-binding proteins and
the HMR-E silencer-DNA may also contribute to silencer function.
For example, the interaction of ORC with its binding site at the
HMR-E silencer is substantially stronger than its interaction with
many of its binding sites present in non-silencer DNA replication
origins (20). This strong interaction, which is greater than needed
for occupancy of ORC at replication origins, contributes to HMR-E
silencer function, raising the possibility that a silencer-specific
ORC-DNA interaction enhances the binding of Sir1p to ORC.
A mechanistic understanding of how individual silencer-binding
protein-DNA interactions at the HMR-E silencer control the bind-
ing of Sir proteins requires further investigation. For example, the
ORC-silencer DNA interaction could enhance Sir1p binding either
through promoting a unique conformation of ORC or by presenting
silencer DNA for direct contacts by Sir1p. The other silencer-
binding proteins may be influenced by their interactions with their
silencer binding site. Indeed, Rap1p function may be influenced by
the nature of its binding site (21, 22). It is unclear how Abf1p
functions at the silencer because no distinct Abf1p-Sir interactions
have been published although there are specific silencing-defective
alleles of ABF1 (19). Finally, the SPC functions with a distinct
orientation, nucleating silent chromatin more efficiently in one
direction (Fig. 3), but the mechanisms controlling this directional-
ity are unknown. Structural studies of the SPC should reveal
* This minireview will be reprinted in the 2005 Minireview Compendium,
which will be available in January, 2006.
‡ To whom correspondence should be addressed: Dept. of Biomolecular
Chemistry, 587 MSC, 1300 University Ave., University of Wisconsin Medical
School, Madison, WI 53706-1532. Tel.: 608-262-9370; E-mail: firstname.lastname@example.org.
1The abbreviations used are: ORC, origin recognition complex; SPC, si-
lencer protein complex; ChIP, chromatin immunoprecipitation assay.
THE JOURNAL OF BIOLOGICAL CHEMISTRY
Vol. 280, No. 10, Issue of March 11, pp. 8629–8632, 2005
© 2005 by The American Society for Biochemistry and Molecular Biology, Inc.
Printed in U.S.A.
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