Ways to Kill the Messenger: Unraveling the Regulation of Cytoplasmic mRNA Decay

The regulation of mRNA levels is a key aspect of gene expression control and is determined by the processes of transcription and mRNA decay. Changes in these two processes can cause widespread changes in gene expression. Nevertheless, how some mRNAs are specifically targeted for degradation, particularly under cellular stress, is still unclear. Furthermore, How nuclear mRNA processing events dictate cytoplasmic mRNA fate is also an area with many unanswered questions. Here I present several studies that specifically take a closer look at the various mechanisms behind degradation of mRNAs in the cytoplasm. Chapter 2 describes how mutations in 3’ end processing and nuclear export factors (THO/TREX-2) causes aberrant mRNA-protein (mRNP) foci to form in the cytoplasm. These granules (termed TT foci) are composed of components found within stress granules (SGs) but are less dynamic in nature, as suggested by their resistance to cycloheximide-induced disassembly. TT foci are cleared by autophagy, suggesting a potentially novel mechanism of quality control to degrade aberrant mRNAs that escape nuclear degradation pathways and enter the cytoplasm. Next, chapter 3 looks at how common mechanisms used for determining mRNA half-life cause changes in major signaling pathways, specifically looking at the TORC1, Hog1, and PKC pathways. Chemical inhibitors such as 1,10 phenanthroline and thiolutin show the strongest effects and inhibits TORC1 and PKC signaling but activates Hog1 signaling. 4-thiouracil (4tU), which is used to metabolically label mRNAs and assess mRNA decay or transcription rates, causes an increase in Processing bodies (PBs). Given that signaling pathways can regulate mRNA decay, our findings suggest more research is needed in yeast to determine an easy-to-use system to probe the contributions of various signaling pathways on mRNA decay without generating experimental artifacts. Lastly, Chapter 4 is an ongoing study that examines at the role of TORC1 signaling in regulating decay of Ribosome Biogenesis (Ribi) mRNAs through the mRNA binding protein Puf4. However, due to our findings in Chapter 3, untangling the exact contribution of TORC1 on mRNA stability has been challenging. However, we present data indicating that both Puf4 and TORC1 can facilitate decay of the Ribi mRNA NSR1, and that Puf4 binding to NSR1’s 3’UTR is required for this regulation. However, Puf4 binding to NSR1 mRNA does not change in the presence or absence of TORC1 signaling. This suggests altered interactions of Puf4 with mRNA decay factors may be crucial and may be modified by TORC1 signaling, an area of immediate future interest.Release after 02/23/202