Sequence Elements in Yeast Introns Regulate Translation to Make Functional Proteins

Abstract

RNA splicing is an important mode of eukaryotic gene regulation. By the process of splicing, introns are excised from the pre‐mRNA and exons are ligated to make a mature message, which is then translated to make protein. In eukaryotes, introns play a key role in alternative splicing whereby alternative exons are ligated to make different mRNAs from the same pre‐RNA transcript. These mRNAs can, in turn, be translated to make different protein isoforms and increase protein diversity.Here, we explore a unique role for inefficiently‐spliced introns. Previously, we showed that an intron‐retained isoform of GCR1 encodes a protein which is initiated from an in‐frame intronic AUG. We have identified regulatory sequences upstream of the AUG that control translation from the intronic translation start site. Furthermore, we show that Rps25, a small ribosomal protein subunit, regulates utilization of the intronic AUG to facilitate the expression of the protein and control the relative levels of the two Gcr1p isoforms—one from the spliced RNA and one from the unspliced RNA. In light of this result, we next analyzed the introns of all known intron‐containing genes to assess whether other introns contain an AUG that is in‐frame with the same stop codon as the spliced isoform. We have identified and tested several such genes to determine whether the intronic AUGs direct protein expression and find that, indeed, they do. Moreover, when cells deleted of these genes express only the intron‐retained isoform, this restores viability to sick cells, suggesting that the proteins produced by intron retention are functional. These studies demonstrate a unique role for introns in regulating alternative isoform production to generate functionally important protein diversity.Support or Funding InformationThis work was supported by the National Science Foundation (BIO/MCB 1518316 to T.L.J) and National Institute of General Medical Sciences (GM‐085474 to T.L.J)