Inability of T7 RNA polymerase to
processively transcribe higher eukaryotic chromatin is interpreted as
a correlate of its reported inhibition by nucleosomes on
reconstituted templates in vitro. We used chromosomally integrated
reporter cassettes to examine features of T7 transcription in a lower
eukaryotic system. Luciferase reporters were targeted to rDNA in
transgenic Trypanosoma brucei stably expressing the phage polymerase,
Because trypanosome mRNAs are capped by RNA splicing in trans, T7
transcription could be gauged by luciferase activity. In contrast to
findings from higher eukaryotes, T7 transcription is vigorous and
processive on chromatin templates in T.brucei, surpassing levels
achieved with endogenous promoters, including those recruiting RNA
polymerase I, This may be a reflection of intrinsic differences in
chromatin structure between differently evolved eukaryotes or of an
integration site that is exceptionally permissive for T7
transcription due to a local accessible chromatin conformation. T7
transcription could be manipulated to achieve different levels of
constitutive expression, through the use of promoter mutations.
Moreover, T7 initiation could be regulated by the prokaryotic Tet
repressor and elongation halted by T7 terminator sequences. We have
exploited these features to construct a robust inducible expression
system, whose utility potentially extends to other trans-splicing
organisms.