W013 Unproductive Alternative Splicing of Plant Circadian Genes

Date: Sunday, January 15, 2012
Time: 3:50 PM
Room: Royal Palm Salon 1-2
Sergei Filichkin , Oregon State University, Corvallis, OR
Massively parallel transcriptome sequencing (RNA-seq) has revealed that the repertoire and extent of the alternative splicing (AS) in eukaryotes were considerably underestimated. Increasing evidence also suggests that substantial proportions of pre-mRNAs undergo unproductive alternative splicing resulting in incorporation of in-frame premature termination codons (PTCs). The destiny and potential functions of the mRNAs harboring PTCs remain poorly understood. Plant circadian genes present a special case study of unproductive splicing because their transcripts oscillate daily with the sharp changes in the abundance of the mRNAs.  We conducted a systematic survey of alternative splicing events among Arabidopsis circadian clock genes. Using RNA-seq data we identified a set of the circadian clock genes with potentially novel alternative splicing patterns. Further validation uncovered a group of circadian genes with single and multiple alternative splice isoforms. Predicted AS events were confirmed using reverse transcription polymerase chain reaction (RT-PCR), quantitative RT-PCR, and/or direct Sanger sequencing. Many of the alternatively spliced events were represented by the intron retention or by the selection of the alternative splice acceptor site. These types of events generally produced splice isoforms harboring in-frame PTCs. In several circadian genes the PTC+ transcripts accumulated at substantial levels. The ratios of the unproductive isoform to its full-length protein coding mRNA (pc-mRNA) could shift sharply under the specific environmental stress conditions. Unproductive splicing may play a general role in homeostatic regulation of the pc-mRNAs. Concomitantly, the circadian nonsense transcripts could play a role in post-transcriptional regulation of the cyclical abundance of their respective pc-mRNAs.