The trypanosome spliced leader small RNA gene family: stage-specific modification of one of several similar dispersed genes

Abstract

Diverse mRNAs of Trypanosoma brucei possess the same 5' terminal 35 nucleotides, termed the spliced leader (SL), which appears to be derived from a separate 135 nucleotide transcript. This small SL RNA is encoded within a 1.4 kb unit of DNA which is tandemly reiterated in the genome. In addition, there are at least 4 orphon elements containing SL sequences dispersed from the tandem array. Here we show that during the trypanosome life cycle one of the SL orphons undergoes a stage-specific modification that prevents cleavage of an EcoRV site and we further demonstrate that although only one orphon is modified, three of the SL orphons are flanked by very similar sequences. Each of these contains SL reiteration units including the non-transcribed spacer DNA, suggesting that they did not originate through an RNA intermediate. In addition no evidence of direct repeats at the junction of 1.4 kb and non-1.4 kb DNA was observed. Finally, a phylogenetic survey indicates that while many trypanosomatid species possess similarly organized SL-like sequences, only the SL orphons of closely related subspecies of the T. brucei - T. evansi complex share similar flanking regions.