Analysis of RNA decay for alternatively spliced transcripts by RNA tagging

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Alternative splicing has been proposed as a possible mechanism to increase the number of proteins significantly compared to the number of genes. However, it has been under discussion whether it actually results in functional protein products. Alternatively, it may provide a post-transcriptional mechanism for gene expression regulation or result from noise in the splicing machinery. In this project, we aim to investigate alternative splicing using a newly developed method for directly tagging newly transcribed RNA and determining RNA half-lives with high precision. Using RNA tagging and novel exon array and RNA sequencing technology, RNA transcription and decay rates can be determined for individual exons and alternative splicing events. For this purpose, we will develop methods for determining halflives of alternatively spliced transcripts and identifying significant differences between splicing isoforms. In this way, we aim to define a predictive model to distinguish functional splicing products and characterize the regulation of the corresponding protein products. Furthermore, methods will be developed to identify novel alternative splicing products from measurements of RNA de novo synthesis and to analyze the mechanisms determining RNA decay. Finally, the comparison of different RNA quantification techniques will provide a comprehensive assessment of the performance of each method and increase the confidence of the final results.