Abstract
Eukaryotic genes generate multiple RNA transcript isoforms though alternative transcription, splicing, and polyadenylation. However, the relationship between human transcript diversity and protein production is complex as each isoform can be translated differently. We fractionated a polysome profile and reconstructed transcript isoforms from each fraction, which we term Transcript Isoforms in Polysomes sequencing (TrIP-seq). Analysis of these data revealed regulatory features that control ribosome occupancy and translational output of each transcript isoform. We extracted a panel of 5' and 3' untranslated regions that control protein production from an unrelated gene in cells over a 100-fold range. Select 5' untranslated regions exert robust translational control between cell lines, while 3' untranslated regions can confer cell type-specific expression. These results expose the large dynamic range of transcript-isoform-specific translational control, identify isoform-specific sequences that control protein output in human cells, and demonstrate that transcript isoform diversity must be considered when relating RNA and protein levels.
Highlights
Eukaryotic genes can produce a staggering diversity of messenger RNA products
We showed that the dynamic range of transcript-isoform-specific translational control spans at least two orders of magnitude in human cells (Figure 5), indicating that it is crucial to take isoform-level effects into account when assessing translation in organisms with extensive alternative transcript processing (Figure 7A)
We find that alternative 30 untranslated region (UTR) broadly influence translation, with longer isoforms of the same gene associated with lower protein production (Figure 3B)
Summary
Eukaryotic genes can produce a staggering diversity of messenger RNA products. In humans, there is a median of five annotated transcript isoforms per gene, with more than 80 annotated isoforms in some cases (Ensembl release 75). These transcript isoforms arise from the combined action of alternative transcription initiation, splicing and cleavage, and polyadenylation, which are often cell type specific (Barbosa-Morais et al, 2012; Mele et al, 2015; Merkin et al, 2012; Wang et al, 2008; Xiong et al, 2015; Zheng and Black, 2013) As each of these transcript isoforms may in principle harbor a unique set of translational control elements, gene-level expression, which averages out the translational potential of each individual transcript isoform, may not be an accurate measure of protein levels.
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