Oh, Behave! SASsy protein team's paradoxical effects on gene silencing still confound researchers (Chromatin structure/Gene regulation)
Authors: Davenport R. John 
Published: August 2003

A fickle protein continues to taunt researchers with its paradoxical behavior, but some newly discovered molecular associations are beginning to shed light on its activities. Subtle changes in the way DNA and proteins pack together can turn off genes and prevent DNA from breaking and rejoining in new combinations. This so-called gene silencing process plays a role in regulating aging: High doses of the silencing protein SIR2 extend the life-spans of yeast and nematodes (see
Kaeberlein
Perspective). According to new work, several proteins involved in silencing act in concert and perhaps shut down newly copied DNA. The results reveal previously unknown molecular contacts, but the mechanisms by which the proteins influence silencing--and the reasons they turn genes on in some situations and off in others--are still unclear.

Considerable evidence suggests that the removal of acetyl groups from histones--the protein scaffolds that wind DNA into a compact form known as chromatin--instigates silencing of genes near the chemical modifications. Removal of any one of three yeast genes known as
SAS2
,
SAS4
, and
SAS5
can restore defective silencing at a particular chromosomal location, known as HMR. This observation suggests that the normal versions of the SAS proteins inhibit silencing. Consistent with that role, the SAS2 protein resembles acetyltransferases, enzymes that attach acetyl groups to protein side chains. But in other genomic regions, a nonfunctional version of SAS2 further reduces silencing, hinting that at those sites addition of acetyl groups might hush DNA. Acetyltransferases frequently work as part of molecular teams, so identifying SAS2's co-conspirators could illuminate how SAS2 both enhances and hinders silencing.

In the new work, two groups independently sought protein partners that bind to SAS2. Using antibodies that recognize this protein, they pulled it from a mix of yeast cell contents. But it did not emerge alone: SAS4 and SAS5 accompanied SAS2, presumably because they had attached themselves to it. The result supports previous genetic experiments suggesting that the three genes act in the same pathway.

Further studies by the two teams hint that the SAS conglomeration sticks to other proteins that assist new DNA in twisting properly around histones. Osada and colleagues discovered that a protein called Asf1 also joins the SAS party; Meijsing and Ehrenhofer-Murray showed that the CacI protein attaches to the SAS trio as well. Previous work indicates that Asf1 and CacI help histones load onto freshly copied DNA to restore chromatin structure. Moreover, defects in either protein impair silencing. The new results suggest that DNA-histone assembly machinery brings the SAS conglomeration to newly replicated DNA, where SAS2 perhaps modifies histones to influence silencing. This idea is consistent with previous results indicating that silencing states must be reset after DNA is duplicated and repacked around histones.

Additional results suggest that SAS2 acetylates a specific amino acid constituent of a histone protein known as H4. Meijsing and Ehrenhofer-Murray found that a mutation in H4 that locks it into a deacetylated state produces changes in silencing similar to those exhibited by SAS2 mutations: The alteration restores defective silencing at HMR but reduces it in other areas of the genome. No one has demonstrated SAS2's enzymatic activity directly, however; SAS2 might modify an as-yet-unidentified protein, which in turn modulates silencing.

The SAS amalgam hinders silencing at a region implicated in aging. Meijsing and Ehrenhofer-Murray showed that removing any one of the three SAS proteins results in strong silencing of ribosomal DNA, the chromosomal site most important for SIR2's influence on aging--and SIR2 is required for this effect. Whether the SAS complex also antagonizes SIR2's ability to extend life-span is not known.

SAS functions have been but a gleam in a biochemist's eye: Researchers have known that the SAS genes influence silencing, but the mechanism has remained mysterious. The new identification of molecular connections made by SAS proteins will help scientists understand how acetylation contributes to silencing. Such insight could reveal the machinations behind SAS's two-faced behavior.

--R. John Davenport

S. Osada, A. Sutton, N. Muster, C. E. Brown, J. R. Yates III, R. Sternglanz, J. L. Workman, The yeast SAS (something about silencing) protein complex contains a MYST-type putative acetyltransferase and functions with chromatin assembly factor ASF1.
Genes Dev.
15
, 3155-3168 (2001).
[Abstract]
[Full Text]

S. H. Meijsing and A. E. Ehrenhofer-Murray, The silencing complex SAS-I links histone acetylation to the assembly of repressed chromatin by CAF-I and Asf1 in
Saccharomyces cerevisiae.
Genes Dev.
15
, 3169-3182 (2001).
[Abstract]
[Full Text]