Inhibition of Nonsense-Mediated RNA Decay Activates Autophagy

Abstract

Nonsense-mediated RNA decay (NMD) is an mRNA surveillance mechanism which rapidly degrades select cytoplasmic mRNAs. We and others have shown that NMD is a dynamically regulated process inhibited by amino acid deprivation, hypoxia, and other cellular stresses commonly generated by the tumor microenvironment. This inhibition of NMD can result in the accumulation of misfolded, mutated, and aggregated proteins, but how cells adapt to these aberrant proteins is unknown. Here we demonstrate that the inhibition of NMD activates autophagy, an established protein surveillance mechanism, both in vitro and in vivo. Conversely, the hyperactivation of NMD blunts the induction of autophagy in response to a variety of cellular stresses. The regulation of autophagy by NMD is due, in part, to stabilization of the documented NMD target ATF4. NMD inhibition increases intracellular amino acids, a hallmark of autophagy, and the concomitant inhibition of autophagy and NMD, either molecularly or pharmacologically, leads to synergistic cell death. Together these studies indicate that autophagy is an adaptive response to NMD inhibition and uncover a novel relationship between an mRNA surveillance system and a protein surveillance system, with important implications for the treatment of cancer. N onsense-mediated RNA decay (NMD) is a cellular surveillance system that rapidly degrades select mRNAs. While the exact mechanism of NMD is still not completely delineated, current models suggest that during a pioneer round of translation, the translation complex pauses at premature termination codons (PTCs) upstream of exon junction complexes (EJCs). Crucial components of the NMD mechanism, including Upf1/Rent1 and Upf2/Rent2, bridge the EJC to a paused translation complex and initiate RNA degradation. In addition to PTCs located upstream of EJCs, long 3= untranslated regions and other, less-well-defined features of the processed mRNA transcript can result in mRNA degradation by NMD (reviewed in reference 1). Up to 30% of all mutations responsible for human genetic disorders, including cystic fibrosis, muscular dystrophy, and thalassemia, result in PTCs upstream of EJCs (2). NMD also degrades nonmutated mRNAs, including those important for the response to cellular stress and amino acid transport (3-5). The recent observation that NMD is inhibited by amino acid deprivation, hypoxia, the accumulation of unfolded proteins in the endoplasmic reticulum (ER), and other cellular stresses that lead to the phosphorylation of eukaryotic initiation factor 2␣ (eIF2␣) has led to a greater appreciation of the potential role that the dynamic regulation of gene expression by NMD could play in physiology and pathology (3-5).The stress-induced inhibition of NMD stabilizes the stressresponsive transcription factor ATF4, augments the ER stress response, and permits cells to grow in soft agar and as tumor explants, conditions in which cells are deprived of nutrients and oxygen (5). However, the full biological consequences of NMD regulation are unknown. These conse...

Phosphorylation of eIF2α triggered by mTORC1 inhibition and PP6C activation is required for autophagy and is aberrant in PP6C-mutated melanoma

“…We noted that multiple ATF-4 downstream transcriptional targets (ATF-3, CHOP, the CHOP target gadd34) and transcripts important for the induction of autophagy (LC3B and ATG5 (4, 6, 7)) were upregulated in rapamycin treated eIF2α wild-type cells and not in eIF2α S51A/S51A cells (Fig 2A), despite a minimal upregulation of ATF-4 protein with rapamycin treatment (Fig 2B). …”

“…The phosphorylation of eIF2α, which also occurs with amino acid deprivation as well as with endoplasmic reticulum (ER) and other cellular stresses, similarly attenuates global protein synthesis (reviewed in (3)). However, eIF2α phosphorylation also paradoxically increases the translation of select mRNAs, including the transcription factor ATF-4 which transactivates the autophagy genes LC3B and ATG5 (4-7). …”

Caspases shutdown nonsense-mediated mRNA decay during apoptosis

“…For example, apoptotic-like programed cell death does not involve caspases activation, but rather an apoptosis-inducing factor. 40 Recent studies have also suggested that inhibition of NMD can lead to autophagy; 41 however, whether NMD is inhibited during distinct autophagic inductions has yet to be determined. Cell death by necrosis is yet another mechanism that involves a family of proteases, the calpains rather than caspases.…”

Overexpression of Smooth Muscle Myosin Heavy Chain Leads to Activation of the Unfolded Protein Response and Autophagic Turnover of Thick Filament-associated Proteins in Vascular Smooth Muscle Cells

“…Therefore, the pathway linking ER stress response to autophagy in SM1 cells appears to be independent of decreased levels of mTOR signaling. It is interesting to note that a recent study found that inhibition of nonsense-mediated mRNA decay in cells activates autophagy (50). Although the cellular pathway for the increased autophagy was not investigated, protein production from the mutant transcript may have induced ER stress and autophagy in a similar manner that overexpression of myosin heavy chain does in the SM1 cells.…”