Identifying the right stop: determining how the surveillance complex recognizes and degrades an aberrant mRNA

Abstract

The nonsense-mediated mRNA decay (NMD) pathway functions by checking whether translation termination has occurred prematurely and subsequently degrading the aberrant mRNAs. In Saccharomyces cerevisiae, it has been proposed that a surveillance complex scans 3Ј of the premature termination codon and searches for the downstream element (DSE), whose recognition by the complex identifies the transcript as aberrant and promotes its rapid decay. The results presented here suggest that translation termination is important for assembly of the surveillance complex. Neither the activity of the initiation ternary complex after premature translation termination has occurred nor the elongation phase of translation are essential for the activity of the NMD pathway. Once assembled, the surveillance complex is active for searching and recognizing a DSE for~200 nt 3Ј of the stop codon. We have also identified a stabilizer sequence (STE) in the GCN4 leader region that inactivates the NMD pathway. Inactivation of the NMD pathway, as a consequence of either the DSE being too far from a stop codon or the presence of the STE, can be circumvented by inserting sequences containing a new translation initiation/termination cycle immediately 5Ј of the DSE. Further, the results indicate that the STE functions in the context of the GCN4 transcript to inactivate the NMD pathway.

Regulation of Glucose-Dependent Gene Expression by the RNA Helicase Dbp2 in Saccharomyces cerevisiae

“…Consistent with prior data stating that Dbp2 is exceptionally stable with an estimated half-life of 250 min (Laxman et al 2010), we did not observe an appreciable decrease within the 1-hr time frame of our analysis. This was not due to an incomplete translational block, as the levels of another RNA helicase, Upf1, was degraded with a half-life within the range of other studies (Figure 2A, red;Ruiz-Echevarria et al 1998). Furthermore, the stability of Dbp2 did not change upon removal of glucose within 1 hr ( Figure 2B).…”

Cloning and Characterization of Cyclooxygenase-1b (Putative Cyclooxygenase-3) in Rat

“…An mRNA is targeted by nonsensemediated decay by the presence of a cis-acting downstream element appropriately located within 200 nucleotides downstream of the stop codon (Ruiz-Echevarria and Peltz, 1996;Ruiz-Echevarria et al, 1998). Conversely, mRNA containing premature stop codons can avoid nonsense-mediated decay if they possess a cis-acting stabilizer element (Ruiz-Echevarria et al, 2001).…”

Evidence against a direct role for the Upf proteins in frameshifting or nonsense codon readthrough

“…The nonsense-mediated decay (NMD) pathway functions to recognize and rapidly degrade transcripts containing premature termination codons (PTC) in their open reading frames (ORF) (Hilleren and Parker 1999;Gonzalez et al 2001;Wagner and LykkeAndersen 2002). Because faithful recognition of a PTC is essential for triggering the rapid removal of such mRNAs, this "mRNA surveillance" pathway represents a nexus between the cell's machinery for mRNA turnover and translational fidelity (Ruiz-Echevarria et al 1998a). …”