trans

CLASS I TRANSCRIPTION IN TRYPANOSOMATIDS 51
they exhibit structural differences, and there
is substantial evidence that life-cycle-specific
gene expression is in part regulated at the transcriptional
level. Hence, it is possible that procyclin
and (m)VSG ES promoter transcription
depends on novel transcription factors or
factor domains.
Transcription regulation
T. brucei has two diploid procyclin ESs which
are exclusively expressed in procyclic cells.
Procyclin ES promoters are active in both
bloodstream and insect forms, but posttranscriptional
processes prevent procyclin
expression in the bloodstream. On the other
hand, the procyclin ES promoter is up to tenfold
more active in procyclics than in bloodstreamform
trypanosomes. This differential activity
was observed independently of the genomic
context in which the procyclin ES promoter
was integrated. Moreover, procyclin ES promoter
transcription in a procyclic cell extract
was in comparison to VSG ES and rDNA promoter
transcription exceptional by its fourfold
higher efficiency, a distinct lag phase, a high
template DNA concentration optimum, and its
tolerance to manganese cations. Taken together,
these data suggest that procyclin ES transcription
in procyclic cells is enhanced by a life cyclespecific
component.
There are at least 27 mVSG ESs and about
20 bloodstream-form VSG ESs in a T. brucei
cell. However, the VSG coat of metacyclic and
bloodstream-form trypanosomes consists of
identical molecules which are expressed from
a single gene. Therefore, only one (m)VSG ES
is maximally expressed, while the others are
inactivated. Regulation occurs at the level of
transcription, but it is currently under debate
whether in bloodstream-form trypanosomes
inactivation is predominantly due to inefficient
transcription initiation or to transcription
attenuation. Differential expression of VSG
ESs does not depend on promoter sequences,
because replacement of the VSG ES promoter by
an rDNA promoter did not impair activation/
inactivation control of VSG ESs. Furthermore,
the detection of DNase I hypersensitive sites
in both active and inactive VSG ES promoters
indicated that auxiliary transcription factors
are assembled on all VSG ESs. These findings
suggest an epigenetic control of VSG expression
in the bloodstream. In procyclic cells,
VSG ES promoters generally direct a low level
of transcription which is terminated within
700 bp of the transcription initiation site (TIS).
Interestingly, rDNA and procyclin ES promoters
integrated into a VSG ES are fully active and
not repressed, suggesting that repression of
VSG ES transcription is dependent on the promoter
sequence. Finally, mVSG ES promoters
in their chromosomal contexts are only active
in the metacyclic stage and, according to nascent
RNA analysis, are completely inactivated
in both proyclic and bloodstream-form cells.
Promoter structures
A well characterized example of a class I gene
promoter from lower eukaryotes is the S. cerevisiae
rDNA promoter which comprises three
sequence blocks denoted as domains I, II, and
III (Figure 3.1). Domain I ranges from position
28 to position 8 (28/8) relative to the
TIS and represents the core promoter, being
defined as the minimal structure required for
accurate transcription initiation. Typically, it is
the only absolutely essential promoter element.
Domain II (76/51) and domain III
(146/91) constitute a bipartite upstream
element which, unlike the core promoter, stably
interacts with trans-acting factors. Finally,
a sequence motif was identified at position
MOLECULAR BIOLOGY