Evidence of discontinuous transcription in the trypanosomatid Crithidia fasciculata.

Abstract

In an effort to exploit the advantages of Crithidia fasciculata for detailed analysis of the mechanisms of discontinuous transcription in the trypanosomatid family, we have cloned, sequenced, and characterized the mini-exon gene repeat in Crithidia and mapped the termini of its primary transcript. We find that Crithidia contains approximately 500 mini-exon genes, present almost exclusively as tandemly repeated arrays on a single chromosome. Transcripts derived from these genes are approximately 90 bases in length with heterogeneity at both the 5' and 3' ends. Primer extension experiments reveal a putative splicing intermediate. Specific inhibition of in vitro translation of Crithidia mRNAs by an oligonucleotide complementary to the mini-exon suggests that all Crithidia mRNAs contain the mini-exon at their 5' termini. Comparison of mini-exon gene sequences from various trypanosomatids reveals several regions of conservation that imply functional constraints on the transcription of mini-exon genes and the processing of their transcripts.