A monocistronic transcript for a trypanosome variant surface glycoprotein.

Abstract

Many protein-encoding genes of African trypanosomes are transcribed as large polycistronic pre-mRNAs that are processed into individual mRNAs containing a 5' spliced leader and 3' poly(A). The 45-to 60-kb pre-mRNAs encoding some variant surface glycoproteins (VSGs) contain as many as eight unrelated coding regions. Here we identify the promoter for a metacyclic VSG gene that is expressed without duplication in a bloodstream trypanosome clone. This 70-bp promoter is located 2 kb upstream of the telomere-linked VSG gene and directs the synthesis of a monocistronic VSG pre-mRNA lacking the 5' spliced leader. Its sequence only slightly resembles those of other known trypanosome promoters, but it does cross-hybridize with several related sequences elsewhere in the genome. These results suggest that a new class of trypanosome promoters has been found, whose function is to initiate monocistronic transcription of those VSG genes normally expressed during the metacyclic stage.

African Trypanosomes Have Differentially Expressed Genes Encoding Homologues of the Leishmania GP63 Surface Protease

“…Trypanosomes-All experiments were conducted with material isolated from the MVAT4 bloodstream clone of the Trypanosoma brucei rhodesiense WRATat serodeme, or from procyclic organisms derived from an MVAT5 bloodstream clone of the same serodeme (32,33). These bloodstream and procyclic organisms were maintained in rats or grown in culture, respectively, as described previously (32)(33)(34).…”

“…These bloodstream and procyclic organisms were maintained in rats or grown in culture, respectively, as described previously (32)(33)(34).…”

“…Radioactive signals were quantitated by electronic autoradiography using a Packard instant imager (Packard, Meriden, CT). A bacteriophage ZAP library of cDNAs, and a bacteriophage FIX library of genomic DNA fragments, both derived from T. brucei rhodesiense bloodstream clone MVAT4 (32), were screened with an African trypanosome GSS (31) using conventional procedures (40). The cDNA inserts were rescued in the plasmid pBluescript.…”

“…Leishmania and Crithidia GP63s-The African trypanosome GSS possessing similarity to GP63 genes (31) was used as a probe to screen cDNA and genomic DNA libraries derived from T. brucei rhodesiense bloodstream clone MVAT4 (32). Positive clones were identified in both libraries, and several clones from each were selected for further characterization.…”

“…Transcription of the GP63 Genes in T. brucei-Nuclear run-on transcription experiments have been used to demonstrate that the active telomere-linked VSG gene is transcribed in bloodstream trypanosomes, but not in procyclic trypanosomes (29,32). In contrast, many tandemly repeated genes in Trypanosomatids, including GP63 genes in Leishmania, are transcribed at all stages of the organisms' life cycles, and their steady state mRNA levels are determined by post-transcriptional regulatory events (reviewed in Refs.…”