UC San Diego

Every step in the existence of a messenger mRNA is carefully regulated. I explored the impact of a feature present in the great majority of eukaryotic mRNAs: the poly(A) tail. Poly(A) tails are important elements in mRNA maturation, translation and stability, but only recently the tail lengths of mRNAs have been revealed on a transcriptome wide scale. I have adapted the poly(A) sequencing method TAIL-seq and developed aTAIL-seq (adapted TAIL-seq). Chapter 4 contains detailed protocols of aTAIL-seq and other methods of poly(A) analysis.

In this work, I have applied aTAIL-seq to measure the poly(A) tails of Caenorhabditis elegans. The nematode C. elegans is an model organism that has been used to identify multiple conserved biological processes, including the discovery of microRNAs (reviewed in Chapter 2).

In Chapter 3, I explore the relationship between the poly(A) tail, translation and stability in C. elegans and other eukaryotes. Traditionally, long tails have been thought to enhance translation and stability of mRNAs. However, we found that the most abundant types of mRNAs, such as those encoding ribosomal proteins, have the shortest tail lengths, while the least abundant, such as mRNAs for transcription factors, have the longest tails. This difference is related to translation efficiency, as genes enriched for optimal codons and ribosomal occupancy have the shortest median tails. These results suggest that translation promotes poly(A) tail shortening, an idea supported by our observation that non-coding RNAs carry long poly(A) tails. We find that, in general, the most abundant and well-translated mRNAs have the shortest median poly(A) tail lengths. While this study seems to contradict the dogma that deadenylation is associated with translational inhibition and mRNA decay, it actually points to a mechanism where well expressed mRNAs undergo pruning of their poly(A) tails to lengths that accommodate 1-2 poly(A) binding proteins (PABPs). This may be an optimal size for PABP to engage in protective and translational functions. Overall, our findings suggest that translation regulates pruning of poly(A) tails, either by actively promoting their shortening or by stabilizing the short-tailed mRNAs. This work changes our understanding of the regulation of poly(A) tail length and gene expression.