Footprints of a trypanosomatid RNA world: pre-small subunit rRNA processing by spliced leader addition trans-splicing.
The addition of a capped mini-exon [spliced leader (SL)] through trans-splicing is essential for the maturation of RNA polymerase (pol) II-transcribed polycistronic pre-mRNAs in all members of the Trypanosomatidae family. This process is an inter-molecular splicing reaction that follows the same basic rules of cis-splicing reactions. In this study, we demonstrated that mini-exons were added to precursor ribosomal RNA (pre-rRNA) are transcribed by RNA pol I, including the 5' external transcribed spacer (ETS) region. Additionally, we detected the SL-5'ETS molecule using three distinct methods and located the acceptor site between two known 5'ETS rRNA processing sites (A' and A1) in four different trypanosomatids. Moreover, we detected a polyadenylated 5'ETS upstream of the trans-splicing acceptor site, which also occurs in pre-mRNA trans-splicing. After treatment with an indirect trans-splicing inhibitor (sinefungin), we observed SL-5'ETS decay. However, treatment with 5-fluorouracil (a precursor of RNA synthesis that inhibits the degradation of pre-rRNA) led to the accumulation of SL-5'ETS, suggesting that the molecule may play a role in rRNA degradation. The detection of trans-splicing in these molecules may indicate broad RNA-joining properties, regardless of the polymerase used for transcription.
Laboratório de Genética, Instituto Butantan, São Paulo, SP, Brazil.
Mario Gustavo
MG
Mayer
Marcos Gonzaga dos
MG
Santos
Maria Fernanda Laranjeira da
MF
Silva
Lucile Maria
LM
Floeter-Winter
D005091:Exons / Q000235:genetics; D007894:Leishmania mexicana / Q000235:genetics*; D009690:Nucleic Acid Conformation; D012322:RNA Precursors / Q000235:genetics*; D020038:RNA, Spliced Leader / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D020040:Trans-Splicing / Q000235:genetics*
D012322:RNA Precursors; D020038:RNA, Spliced Leader