mechanisms used to direct transcription initiation 
by RNA polymerase II. 
Lymphocyte Specificity of TdT 
Transcription 
Dr. Smale and his colleagues have focused on the 
TdT gene for their studies of transcriptional regula- 
tion because of its expression patterns in normal and 
leukemic cells. TdT is a template-independent DNA 
polymerase that inserts nucleotides at the D-J (D, 
diversity; J, joining) and V-DJ (V, variable) junctions 
during immunoglobulin and T cell receptor gene 
rearrangements. In normal cells, TdT expression is 
unusual because it is found in both early B and early 
T cells, suggesting that it may be regulated by tran- 
scription factors common to these two related but 
distinct lineages. Moreover, TdT is expressed at high 
levels in acute lymphocytic leukemias (ALLs) and at 
lower levels in a subset of acute myelocytic leuke- 
mias (AMLs) . TdT expression in AMLs is particularly 
intriguing because it suggests that the regulatory 
protein (s) that deregulates TdT expression (the TdT 
gene is not transcribed in normal myeloid cells) may 
also play a role in leukemogenesis. 
A major objective of Dr. Smale's laboratory has 
been to understand the important protein-DNA in- 
teractions that occur at a DNA sequence element 
called D', which is located ~60 bp upstream from 
the TdT transcription start site. This element is es- 
sential for TdT promoter activity in lymphoid cell 
lines and is capable of interacting with multiple 
DNA-binding proteins. Previously Dr. Smale identi- 
fied and purified a protein called LyF-1, which in- 
teracts with both the D' element and a second ele- 
ment in the TdT promoter and is expressed at high 
levels in lymphoid cells. LyF-1 is also bound to the 
promoters for several other lymphocyte-specific 
genes, suggesting that it may be a transcriptional 
activator for a variety of genes expressed specifi- 
cally in the B and/or T lymphocyte lineages. 
Dr. Smale's laboratory has now demonstrated that 
in addition to LyF-1, members of the ets family of 
nuclear oncoproteins bind specifically to the D' re- 
gion and overlap the LyF- 1 -binding sites. Some of 
the ets members, including ets-1 and fli-1, are ex- 
pressed predominantly in lymphoid tissues. A de- 
tailed characterization suggests that two molecules 
of LyF- 1 bind immediately adjacent to each other on 
the D' element and that the ets-binding site directly 
overlaps the more proximal of the two LyF-1 sites. 
Analysis of a series of substitution mutations sug- 
gests that both LyF- 1 and an ets protein may be es- 
sential for TdT transcriptional activation. Lending 
further support to this hypothesis is the finding that 
the binding sites for LyF- 1 and ets proteins are con- 
served through evolution, as both proteins bind to 
the human TdT promoter as well as to the murine 
promoter. Further studies are directed toward ana- 
lyzing the interplay between these two proteins and 
determining which proteins might be responsible 
for TdT deregulation in acute myelocytic leuke- 
mias. (This work was supported by a grant from the 
National Institutes of Health.) 
Transcription Initiation from Promoters 
That Lack TATA Elements 
A TATA box is a common control element found in 
the promoters for most genes that have been identi- 
fied and in virtually every promoter that has been 
analyzed in biochemical detail. This element is not 
present in the TdT promoter, nor in the promoters 
for many other lymphocyte-specific genes (e.g., X5, 
VpreB, Ick, B29, and pp52). To gain further under- 
standing of transcription initiation in the absence of 
a TATA box. Dr. Smale's laboratory has been analyz- 
ing a core element in the TdT promoter that appears 
to carry out the same functions as TATA. This ele- 
ment, called an initiator (or Inr), is distinct from 
TATA in that it overlaps the transcription start site 
rather than, like TATA, being located 30 bp up- 
stream. Previous studies by Dr. Smale demonstrated 
that the Inr acts in concert with upstream activator 
elements, in the absence of TATA, to direct high lev- 
els of accurately initiated transcription. Moreover, 
Dr. Smale and others surprisingly have found that 
transcription from promoters lacking a TATA box 
and containing an Inr element is absolutely depen- 
dent on the TATA-binding protein (TBP) . 
A number of studies in the laboratory are directed 
toward further understanding of the biological and 
biochemical characteristics of Inr elements. One of 
these studies addresses the role of TBP during tran- 
scription from TATA-less promoters. The nucleotide 
sequence 30 bp upstream from the TdT Inr was var- 
ied extensively, and each resulting promoter was 
tested for transcriptional activity and for TBP bind- 
ing. Efficient Inr-mediated transcription was found 
with most promoters, but promoter strength was 
greatly reduced by highly GC-rich sequences that 
possess the lowest affinity for TBP. 
These results suggest that TBP must interact with 
the —30 region during transcription initiation me- 
diated by an Inr and that most, but not all, sequences 
interact with an affinity sufficient for high transcrip- 
tion levels. Interestingly, the —30 region of the TdT 
promoter contains a sequence that is highly GC-rich 
and relatively poor in promoting Inr-mediated tran- 
scription. Future studies will determine whether 
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