Transcription of long hypothetical orfs in Trypanosoma cruzi: The epimastigote stage uses trans-splicing sites that generate short 5′ UTRs

Abstract

We mapped the 5' UTR for five long hypothetical orfs from Trypanosoma cruzi; each one having a length of more than 10,000 bp. Our aim was to verify the constraints to the length of the 5' UTR and to identify the sites of alternative trans-splicing in the epimastigote stage of three T. cruzi strains. We used reverse transcription PCR to amplify the 5' UTR and demonstrated the transcription of all selected genes as well as additional trans-splicing sites in two of these genes. We observed that the length of the 5' UTR in these genes has a limit, in contrast to previous reports that indicated a trend for longer genes to display a proportionally long 5' UTR. The maximum length of the 5' UTR for the long genes analyzed in the present work is approximately 3% of the orf and, on average, is 1% of the orf length. The poly-pyrimidine tracts used as trans-splicing signal are in the range of 17-53 bases within a distance of 6-59 nt to first spliced-leader acceptor site. T. cruzi populations may use both signals differentially. We conclude that the limit for the 5' UTR length in long genes is determined primarily by the distance to neighboring genes.

Transcriptomic analysis reveals metabolic switches and surface remodeling as key processes for stage transition inTrypanosoma cruzi

“…Visual inspection of the density (sequencing depth) of mapped reads suggests that several reads mapped on UTRs regions, predominantly on 3′UTR. It should be mentioned that T. cruzi UTR lengths vary according to the gene size, and have been estimated from experimentally mapped genes to range from 10-400 bp for 5′UTR and 17-2800 bp for 3′UTR (Brandao & Jiang, 2009), being 3′UTR 2-3 times longer than its corresponding 5′UTR (Ziccardi & Brandao, 2011). This result reinforces previous findings showing the relevance of 3′UTRs in the regulation of gene expression (Coughlin et al, 2000; Da Silva, Bartholomeu & Teixeira, 2006; Di Noia et al, 2000; Nozaki & Cross, 1995; Weston, La Flamme & Van Voorhis, 1999).…”

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“…As shown in this table around 60% of the reads map into the reference genome, which represents a significant proportion taking into account the still incomplete state of the Dm28c genome assembly. experimentally mapped genes to range from 10-400 bp for 5'UTR and 17-2800 bp for 3'UTR (Brandao & Jiang 2009), being 3'UTR 2-3 times longer than its corresponding 5'UTR (Ziccardi & Brandao 2011). This result reinforces previous findings showing the relevance of 3' UTRs in the regulation of gene expression (Coughlin et al 2000;da Silva et al 2006;Di Noia et al 2000;Nozaki & Cross 1995;Weston et al 1999).…”