High-throughput RNA-Seq in Leishmania reveals unexpected patterns of polyadenylation in RNA molecules

High-throughput next generation RNA sequencing (RNA-Seq) has revealed new aspects of transcriptomes that have not been observed or were poorly described using small-scale sequencing. Using Leishmania as a model in a typical RNA-Seq transcriptome analysis by Roche 454 pyrosequencing, we found non-canonical polyadenylation of rRNA and mRNA. Leishmania RNAs are transcribed as long polycistronic pre-RNAs, and mRNAs are usually polyadenylated at the 3´-ends. Analysis of the RNA-Seq transcriptome data revealed the presence of poly(A)-rich tails at the end of the rRNA subunits. This phenomenon could explain the abundant amounts of rRNA that we obtained in mRNA samples after purification by oligo (dT). Surprisingly, we found poly(A)-rich tails in the middle of predicted transcripts (rRNA and mRNA), interrupting the messages. We validated these findings by conventional Reverse Transcription-PCR and Sanger sequencing using 28S beta rRNA, 18S rRNA, 60S ribosomal protein L5 mRNA and eiF5a2 translation factor mRNA. The internal polyadenylation RNA truncation phenomenon is possibly related to a novel post-transcriptional control of gene expression in Leishmania and highlights the importance of evaluating this RNA modification in high-throughput transcriptome analyses in other species.