Regulation of mRNA Decay by Puf Proteins is Dependent on Environmental Conditions

The nucleus of a eukaryotic cell contains the genetic code within DNA that directs growth and function of the cell. The genes contained in DNA make copies of themselves called RNAs, which are molecules able to leave the nucleus and direct protein synthesis. Over- or under-production of any one protein can cause cell malfunction and disease. Regulation of RNA lifespan is one method to ensure proper protein production. The Puf family of RNA-binding proteins regulate mRNA lifespans by controlling the rate of mRNA decay. More specifically, Puf proteins stimulate the removal of the poly(A) tails of mRNAs, which results in translational inhibition of the mRNA into protein and leads to complete mRNA degradation. In yeast, the Puf3 protein (Puf3p) regulates hundreds of mRNAs that encode proteins necessary for mitochondrial function. The ability of Puf3p to regulate mRNA decay is altered by the type of sugar source present. Puf3p is turned “on” in the presence of galactose and “off” in dextrose. The goal of this study was to analyze the effects of mutations in a component of the RNA decay machinery (Pop2p) that is required for condition-specific Puf3-mediated decay stimulation. The mutational effects were tested on both an mRNA normally targeted for degradation by Puf3p (COX17) and a control mRNA not targeted for decay (CBS1). Transcriptional shut-offs were performed to examine the decay rate of pooled mRNA following inhibition of mRNA production. When the temperature sensitive yeast cells are heatshocked, new mRNA production is turned off and the existing mRNAs begin to decay over time. If the Pop2p mutations affect the decay rate of COX17 only, a regulation mechanism specific to Puf3-mediated decay is indicated. If both COX17 and CBS1 decay rates are affected, a global RNA-decay control mechanism is indicate