Abstract
Every kinetoplastid mRNA receives a common, conserved leader sequence via the process of trans-splicing. In Leishmania tarentolae the precursor spliced leader RNA is 96 nucleotides, with a 39-nucleotide exon that is 7meG-capped and methylated on the first 4 nucleotides. trans-Splicing was inferred from the presence of tagged leader in the high molecular weight RNA population and confirmed for accuracy by cDNA cloning. Linker scan substitutions within the exon between positions 10 and 39 did not affect trans-splicing. The trans-splicing efficiency for three of the scan exons was proportional to the tagged:wild type ratio in the spliced leader precursor population. Two scan leader RNAs that were efficiently spliced showed reduced methylation. Longer exons showed reduced splicing, whereas 10- or 20-base pair deletions abolished splicing. These results indicate that size, but not content, of the exon is a constraint on the splicing process. These results, in combination with previous data eliminating a role in transcription initiation, suggest that translation may be the selective pressure on the leader content.
Highlights
From the ‡Department of Microbiology and Immunology, UCLA School of Medicine, Los Angeles, California 90095 and the ¶Department of Medicine, West Los Angeles Veterans Affairs Medical Center, Los Angeles, California 90073
In the nematode Ascaris, part of the 22-nt spliced leader (SL) sequence functions as a promoter [6]; a major role for the SL in transcription initiation has been eliminated in Leishmania tarentolae and Trypanosoma brucei, which have upstream promoters [7, 8]
The predicted size differences because of the presence of intron tag (IT) sequences or exon mutations are indicated (Fig. 1A) and shown in total RNA from transfectants electrophoresed through an acrylamide-urea gel (Fig. 1B)
Summary
From the ‡Department of Microbiology and Immunology, UCLA School of Medicine, Los Angeles, California 90095 and the ¶Department of Medicine, West Los Angeles Veterans Affairs Medical Center, Los Angeles, California 90073. The predicted size differences because of the presence of IT sequences or exon mutations are indicated (Fig. 1A) and shown in total RNA from transfectants electrophoresed through an acrylamide-urea gel (Fig. 1B).
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
