initiator element. They described a novel activity 
that was present in the HeLa TFIID fraction but dis- 
tinct from TBP and the Sp 1 coactivator and that was 
only required for activation of a TATA-less promoter 
by Sp 1 . In the presence of promoter-bound Sp 1 , this 
so-called tethering factor apparently functioned as a 
substitute for the TATA box in transcription assays in 
vitro. These observations led to the proposal that 
Spl plays an essential role in assembling the basal 
initiation factors, possibly by anchoring TBP to the 
TATA-less template via the tethering factor. 
Purification of the TAF/TBP Complex 
Recently substantial progress has been made in 
characterizing the nature and complexity of coacti- 
vators in both human and Drosophila cells. Chro- 
matographic studies indicate that the endogenous 
TFIID consists of a multisubunit complex contain- 
ing the TBP, coactivators, and other associated fac- 
tors. A fraction containing the coactivator activity 
was separated from the endogenous TBP after 
disrupting the tightly associated complex with urea. 
The two separated components, when added to- 
gether with the Gal4-Pro activator, restore activated 
levels of transcription. Immunoaffinity purification 
of the TFIID complex identifies several polypep- 
tides specifically associated with the endogenous 
TBP, some or all of which function as coactivators 
when reconstituted with activators. The isolated 
coactivators also mediate activation by a chimeric 
glutamine-rich activator derived from Spl, but not 
the Gal4-VPl6 activator, suggesting distinct factor 
requirements for different types of transcriptional 
regulators. 
Using a combination of high-resolution chroma- 
tography and antibody affinity chromatography. Dr. 
Tjian and his group recently succeeded in purifying 
to homogeneity several of the TAFs. The isolated 
polypeptides were subjected to proteolysis, the 
amino acid sequences of purified peptides were de- 
termined, and DNA probes derived from the protein 
sequences were generated. In addition, monoclonal 
antibodies directed against individual TAFs were 
isolated and used to probe Xgtl 1 cDNA libraries. By 
means of these two strategies, six of the TAFs have 
recently been cloned. Their structure and function 
are being studied. 
Characterization of the RNA Polymerase I 
Factor SLl Revealed Unexpected Unifying 
Mechanisms for Transcription in Eukaryotes 
The study of transcriptional initiation unraveled 
elegant but complex sets of biochemical interac- 
tions among sequence-specific DNA-binding pro- 
teins, promoter/enhancer elements, and the basal 
transcriptional apparatus. However, the molecular 
interactions that took place between the DNA- 
binding factors and components of the basal appara- 
tus that included RNA polymerase and a variety of 
accessory transcription factors remained elusive. 
Transcription by RNA pol I olfered some unique ad- 
vantages in studying the mechanism of promoter rec- 
ognition and activation. 
In particular, Dr. Tjian and his colleagues ascer- 
tained that only one type of promoter in each spe- 
cies but at least two transcription factors — the pro- 
moter selectivity factor (SLl) and upstream binding 
factor (UBF) — are necessary to direct accurate and 
promoter-specific transcription of rRNA genes in an- 
imal cells. UBF was the only RNA pol I transcription 
factor that was necessary for initial promoter bind- 
ing. The second essential factor, SLl, did not bind 
specifically to the human promoter by itself. How- 
ever, when both UBF and SLl were present, a strong 
cooperative DNA-binding complex with an ex- 
tended DNA-binding region was formed at the hu- 
man rRNA promoter that was critical for transcrip- 
tional initiation. 
The SLl -UBF complex was reminiscent of the situ- 
ation that occurs between site-specific upstream en- 
hancer factors and potential interactions with com- 
ponents of the basal RNA pol II transcriptional 
machinery. Was there, perhaps, a common mecha- 
nistic link between the RNA pol I initiation factor 
SLl and components of the initiation complex uti- 
lized by RNA pol II? 
It was previously known that TBP and multiple 
TAFs are required for regulated transcriptional 
initiation by RNA pol II. This year Dr. Tjian and his 
group reported the biochemical properties of the 
RNA pol I promoter selectivity factor, SLl, and its 
relationship to TBP. Column chromatography and 
glycerol gradient sedimentation indicated that a 
subpopulation of TBP copurified with SLl activity. 
Antibodies directed against TBP efficiently depleted 
SLl transcriptional activity, which was restored 
with the SLl fraction but not purified TBP. Thus TBP 
was necessary but not sufficient to complement SLl 
activity. 
Analysis of purified SLl revealed a complex con- 
taining TBP and three distinct TAFs. Purified TAFs 
reconstituted with recombinant TBP complemented 
SLl activity, and this demonstrated that TBP plus 
novel associated factors are integral components of 
SLl . These findings suggest that TBP might be a uni- 
versal transcription factor and that the TBP-TAF ar- 
rangement provides a unifying mechanism for pro- 
moter recognition in animal cells. 
GENETICS 277 
