Puf1p acts in combination with other yeast Puf proteins to control mRNA stability

Abstract

The eukaryotic Puf proteins bind 3′ untranslated region (UTR) sequence elements to regulate the stability and translation of their target transcripts, and such regulatory events are critical for cell growth and development. Several global genome analyses have identified hundreds of potential mRNA targets of the Saccharomyces cerevisiae Puf proteins; however, only three mRNA targets for these proteins have been characterized thus far. After direct testing of nearly 40 candidate mRNAs, we established two of these as true mRNA targets of Puf-mediated decay in yeast, HXK1 and TIF1. In a novel finding, multiple Puf proteins, including Puf1p, regulate both of these mRNAs in combination. TIF1 mRNA decay can be stimulated individually by Puf1p and Puf5p, but the combination of both proteins is required for full regulation. This Puf-mediated decay requires the presence of two UGUA binding sites within the TIF1 3′ UTR, with one site regulated by Puf5p and the other by both Puf1p and Puf5p. Alteration of the UGUA site in the tif1 3′ UTR to more closely resemble the Puf3p binding site broadens the specificity to include regulation by Puf3p. The stability of the endogenously transcribed HXK1 mRNA, cellular levels of Hxk1 protein activity, and HXK1 3′ UTR-directed decay are affected by Puf1p and Puf5p as well as Puf4p. Together these results identify the first mRNA targets of Puf1p-mediated decay, describe similar yet distinct combinatorial control of two new target mRNAs by the yeast Puf proteins, and suggest the importance of direct testing to evaluate RNA-regulatory mechanisms.

Keywords

• Puf
• decay
• stability
• mRNA
• yeast
• 3′ UTR