Nonsense-Mediated mRNA Decay: Degradation of Defective Transcripts Is Only Part of the Story

Abstract

Nonsense-mediated mRNA decay (NMD) is a eukaryotic surveillance mechanism that monitors cytoplasmic mRNA translation and targets mRNAs undergoing premature translation termination for rapid degradation. From yeasts to humans, activation of NMD requires the function of the three conserved Upf factors: Upf1, Upf2, and Upf3. Here, we summarize the progress in our understanding of the molecular mechanisms of NMD in several model systems and discuss recent experiments that address the roles of Upf1, the principal regulator of NMD, in the initial targeting and final degradation of NMD-susceptible mRNAs. We propose a unified model for NMD in which the Upf factors provide several functions during premature termination, including the stimulation of release factor activity and the dissociation and recycling of ribosomal subunits. In this model, the ultimate degradation of the mRNA is the last step in a complex premature termination process.

Keywords: Upf proteins; quality control; ribosome dissociation; translation termination.

Figure 1

Possible mechanistic differences between normal translation termination and premature translation termination. (a) An overview of normal translation termination. Here, the release factors eRF1 and eRF3 are thought to be efficiently recruited to the A site of the terminating ribosome, thereby preventing Upf2 and Upf3 from associating with the termination complex and precluding the stable association of Upf1 with the 40S subunit. The ordered and coordinated actions of eRF1, eRF3, and ABCE1 promote efficient peptide release, after which ATP hydrolysis by ABCE1 and the combined actions of initiation factors eIF1, eIF1A, and eIF3j promote the dissociation and recycling of the ribosomal subunits. (b) An overview of premature translation termination. Owing to the lack of a 3′-UTR-based stimulatory activity, the release factors eRF1 and eRF3 are inefficiently recruited to the A site of the prematurely terminating ribosome. The delayed recruitment of the release factors or a particular structural feature of a premature termination complex triggers the binding of Upf2 and Upf3 to the terminating ribosome. The joining of Upf2 and Upf3 stabilizes Upf1’s binding to the premature terminating ribosome and, collectively, the three Upf factors control several activities of the termination process. At the early stage, they function in recruiting the release factors or stimulating release factor activities to promote peptide release. At the next stage, they promote the dissociation and recycling of ribosomal subunits. The function of the Upf factors at this stage of premature termination is similar to that of ABCE in normal termination, but mechanistic differences must exist. For example, ABCE1 activity at normal termination leads to complete dissociation of the ribosomal subunits from the mRNA, but the activity of the Upf factors at premature termination may only lead to dissociation of the 60S subunit. Abbreviations: CBP, cap-binding protein; PABPC1, poly(A)-binding protein.

Figure 1

Possible mechanistic differences between normal translation termination and premature translation termination. (a) An overview of normal translation termination. Here, the release factors eRF1 and eRF3 are thought to be efficiently recruited to the A site of the terminating ribosome, thereby preventing Upf2 and Upf3 from associating with the termination complex and precluding the stable association of Upf1 with the 40S subunit. The ordered and coordinated actions of eRF1, eRF3, and ABCE1 promote efficient peptide release, after which ATP hydrolysis by ABCE1 and the combined actions of initiation factors eIF1, eIF1A, and eIF3j promote the dissociation and recycling of the ribosomal subunits. (b) An overview of premature translation termination. Owing to the lack of a 3′-UTR-based stimulatory activity, the release factors eRF1 and eRF3 are inefficiently recruited to the A site of the prematurely terminating ribosome. The delayed recruitment of the release factors or a particular structural feature of a premature termination complex triggers the binding of Upf2 and Upf3 to the terminating ribosome. The joining of Upf2 and Upf3 stabilizes Upf1’s binding to the premature terminating ribosome and, collectively, the three Upf factors control several activities of the termination process. At the early stage, they function in recruiting the release factors or stimulating release factor activities to promote peptide release. At the next stage, they promote the dissociation and recycling of ribosomal subunits. The function of the Upf factors at this stage of premature termination is similar to that of ABCE in normal termination, but mechanistic differences must exist. For example, ABCE1 activity at normal termination leads to complete dissociation of the ribosomal subunits from the mRNA, but the activity of the Upf factors at premature termination may only lead to dissociation of the 60S subunit. Abbreviations: CBP, cap-binding protein; PABPC1, poly(A)-binding protein.

Figure 2

Possible modes of Upf1 recruitment and activation during nonsense-mediated mRNA decay (NMD). Four different modes of Upf1 recruitment and activation during NMD are described. (a) Upf1 recruitment and activation based on the EJC (exon junction complex) model. (b) Upf1 recruitment and activation based on the Upf1 3′-UTR (untranslated region) sensing and potentiation model. (c) Upf1 recruitment and activation based on the faux 3′-UTR model. (d) Upf1 recruitment and activation based on stochastic binding of Upf1 and activation by Upf2 and Upf3 binding only at a premature termination codon. Abbreviations: CBP, cap-binding protein; PABPC1, poly(A)-binding protein.

Figure 3

Major decay events in yeast and mammalian nonsense-mediated mRNA decay (NMD). (a) Major decay events in yeast NMD. At the end of the premature termination process, Upf1 is still associated with the 40S subunit, which remains attached to the mRNA. Upf1 on the 40S subunit then interacts with the Dcp1/Dcp2 decapping enzyme to trigger decapping of the mRNA. After decapping, the mRNA is digested by the Xrn1 5′-to-3′ exonuclease. (b) Major decay events in mammalian NMD. At the end of the premature termination process, phosphorylated Upf1 is still associated with the 40S subunit, which remains attached to the mRNA. Phosphorylated Upf1 on the 40S subunit then interacts with the Smg6 endonuclease and the effector Smg5-Smg7 heterodimer to trigger either endonucleolytic cleavage in the vicinity of the premature termination codon or deadenylation from the 3′ end of the mRNA. In the endonucleolytic cleavage pathway, the 5′ cleavage product is digested by the cytoplasmic exosome and the 3′ cleavage product is digested by Xrn1. In the deadenylation pathway, the Ccr4-Not deadenylase is recruited to the mRNA through an interaction between Smg7 and Pop2. After deadenylation, the mRNA is decapped by the Dcp1/Dcp2/Edc4 complex and then digested by Xrn1.