Dissecting the functions of SMG5, SMG7, and PNRC2 in nonsense-mediated mRNA decay of human cells

Abstract

The term "nonsense-mediated mRNA decay" (NMD) originally described the degradation of mRNAs with premature translationtermination codons (PTCs), but its meaning has recently been extended to be a translation-dependent post-transcriptional regulator of gene expression affecting 3%-10% of all mRNAs. The degradation of NMD target mRNAs involves both exonucleolytic and endonucleolytic pathways in mammalian cells. While the latter is mediated by the endonuclease SMG6, the former pathway has been reported to require a complex of SMG5-SMG7 or SMG5-PNRC2 binding to UPF1. However, the existence, dominance, and mechanistic details of these exonucleolytic pathways are divisive. Therefore, we have investigated the possible exonucleolytic modes of mRNA decay in NMD by examining the roles of UPF1, SMG5, SMG7, and PNRC2 using a combination of functional assays and interaction mapping. Confirming previous work, we detected an interaction between SMG5 and SMG7 and also a functional need for this complex in NMD. In contrast, we found no evidence for the existence of a physical or functional interaction between SMG5 and PNRC2. Instead, we show that UPF1 interacts with PNRC2 and that it triggers 5 ′ ′ ′ ′ ′-3 ′ ′ ′ ′ ′ exonucleolytic decay of reporter transcripts in tethering assays. PNRC2 interacts mainly with decapping factors and its knockdown does not affect the RNA levels of NMD reporters. We conclude that PNRC2 is probably an important mRNA decapping factor but that it does not appear to be required for NMD.

Activation and inhibition of nonsense-mediated mRNA decay controls the abundance of alternative polyadenylation products

“…Increased UPF1 occupancy on an mRNA increases the chances of UPF1 phosphorylation by SMG1, the principal UPF1 kinase (Durand et al, 2016;Kashima et al, 2006). UPF1 phosphorylation is an important signal that favors transcript decay, leading to recruitment of the SMG6 endonuclease and the SMG5/7 complex, which links NMD to decapping and deadenylation enzymes (Eberle et al, 2009;Huntzinger et al, 2008;Loh et al, 2013;Nicholson et al, 2018). Proximal pA site usage may produce short 3'UTRs immune to NMD, while distal pA sites may produce NMD-sensitive mRNAs.…”

A role for DIS3L2 over human nonsense-mediated mRNA decay targets

“…This event promotes the mRNA degradation that involves both exonucleolytic and endonucleolytic pathways in mammalian cells [17]. The different decay paths all seem to require phosphorylated UPF1, which interacts with SMG6, SMG5-SMG7 and Proline-Rich Nuclear Receptor Coactivator 2 (PNRC2) [18][19][20][21]. The first pathway relies on the recruitment of SMG5-SMG7 heterodimer or SMG5-PNCR2 that recruit the decapping enzymes (DCP1 and DCP2) and the CCR4-NOT deadenylation complex.…”

Ribosome recycling factor ABCE1 depletion inhibits nonsense-mediated mRNA decay by promoting stop codon readthrough

“…SMG1-mediated phosphorylation of UPF1, a core factor of NMD, appears to be a key event in activating NMD, because hyper-phosphorylated UPF1 subsequently serves as a binding platform for the heterodimer SMG5/SMG7 and for the endonuclease SMG6, allowing for two different decay routes (Muhlemann and Lykke-Andersen, 2010). A third decay pathway operating via SMG5 and PNRC2 has also been proposed (Cho et al, 2013) but remains controversial (Loh et al, 2013;Nicholson et al, 2018). Moreover, a recent study challenged the SURF complex-based NMD activation model by showing that UPF3B rather than UPF1 interacts with the release factors (Neu-Yilik et al…”