trans

CLASS II TRANSCRIPTION OF PROTEIN CODING GENES 57
hydrogenosomal -succinyl CoA synthetase
gene revealed in the upstream promoter region
two important and closely spaced elements
centered at positions 91 and 75, while mutation
of sequences between 49 and the initiator
element did not greatly influence transient
reporter gene expression. Besides two 10 bp
sequence blocks at positions 54 and 194
which exhibited moderate transcriptional
effects, deletional analysis indicated the presence
of additional positive and negative regulatory
elements further upstream.
As mentioned above (-Amanitin-resistant
transcription), the RNA pol II largest subunit
of T. vaginalis has been characterized and
found to be divergent at the -amanitin binding
site. In addition, it has an unusual C-terminal
domain (CTD) because it lacks the essential
heptapeptide repeats of higher eukaryotes but
is relatively rich in serine and proline residues.
This unusual CTD structure is characteristic
for lower protists and is also found in G. lamblia
and trypanosomatids.
Trypanosomatidae
In trypanosomatids, gene expression is mainly
controlled post-transcriptionally, because the
polycistronic mode of transcription does not
leave much scope for regulation at the level of
transcription initiation. Interestingly, transcription
initiation by RNA pol II at polycistronic
transcription units appears to have a low specificity.
In several trypansomatid species it has
been observed that genes can be expressed
from episomal, promoterless vectors as long
as sequence determinants for RNA processing
are present. Nevertheless, in T. brucei, two putative
promoters have been reported upstream
of the actin gene cluster and the HSP70 locus.
However, a TIS was not unambiguously identified
in these studies and the sequences did not
promote reporter gene expression in a different
study. Hence, it appears that trypansomatid
RNA pol II is recruited to polycistronic transcription
units in an unprecedented, yet to
be determined way. The low initiation specificity
of trypanosomatid RNA pol II is also
reflected in trypanosomatid genomic databases
which lack sequences homologous to
basal class II transcription factors of other
eukaroytes. These factors, in contrast to their
class I counterparts, are highly conserved
among eukaroytes and homologs have been
identified in archaebacteria. Hence, in the
course of evolution, trypanosomatids may have
lost these factors and the ability to specifically
initiate mRNA synthesis. The RNA pol II largest
subunit has been cloned and sequenced in
several trypanosomatid species and is the only
component of the RNA pol II transcription
machinery characterized thus far. Eukaryotes
typically possess a single RNA largest subunit
gene. Therefore, it came as a surprise that
trypanosomes which undergo antigenic variation
possess two copies of this gene encoding
slightly different polypeptides. The possibility
that one of these enzymes transcribes VSG and
procyclin ESs, however, is unlikely, because
the amino acid changes are not located in
the -amanitin binding region and deletion
of both alleles of one gene did not affect cell
viability.
Entamoeba histolytica
Protein coding genes in E. histolytica are transcribed
monocistronically and possess complex
promoter structures indicating that gene
expression in this organism is mainly controlled
at the level of transcription initiation. The most
detailed analyses have been carried out with the
hgl5 gene which encodes a lectin heavy subunit.
The hgl5 promoter has a tripartite core structure
and five distinct upstream regulatory elements.
The core promoter consists of a TATA-like
MOLECULAR BIOLOGY