Overlapping sense and antisense transcription units in Trypanosoma brucei

Procyclins are the major surface glycoproteins of insect-form Trypanosoma b rucei. The procyclin expression sites are polycistronic and are transcribed by an alpha -amanitin-resistant polymerase, probably RNA polymerase I (Pol I). The expression sites are flanked by transcription units that are sensi tive to alpha -amanitin, which is a hallmark of pol II-driven transcription . We have analysed a region of 9.5 kb connecting the EP/PAG2 expression sit e with the downstream transcription unit. The procyclin expression site is longer than was previously realized and contains an additional gene, procyc lin-associated gene 4 (PAG4), and a region of unknown function, the T regio n, that gives rise to trans-spliced, polyadenylated RNAs containing small o pen reading frames (ORFs), Two new genes, GU1 and GU2, were identified in t he downstream transcription unit on the opposite strand. Unexpectedly, the 3' untranslated region of GU2 and the complementary T transcripts overlap b y several hundred base pairs. Replacement of GU2 by a unique tag confirmed that sense and antisense transcription occurred from a single chromosomal l ocus. Overlapping transcription is stage specific and may extend greater th an or equal to 10 kb in insect-form trypanosomes, The nucleotide compositio n of the T brucei genome Is such that antisense ORFs occur frequently, If s table mRNAs can be derived from both strands, the coding potential of the g enome may be substantially larger than has previously been suspected.