Translation stalling induced mitochondrial entrapment of ribosomal quality control related proteins offers cancer cell vulnerability.

Ribosome-associated quality control (RQC) monitors ribosomes for aberrant translation. While the role of RQC in neurodegenerative disease is beginning to be appreciated, its involvement in cancer is understudied. Here, we show a positive correlation between RQC proteins ABCE1 and ZNF598 and high-grade muscle-invasive bladder cancer. Translational stalling by the inhibitor emetine (EME) leads to increased mitochondrial localization of RQC factors including ABCE1, ZNF598, and NEMF, which are continuously imported into mitochondria facilitated by increased mitochondrial membrane potential caused by EME. This reduces the availability of these factors in the cytosol, compromising the effectiveness of RQC in handling stalled ribosomes in the cytosol and those associated with the mitochondrial outer membrane (MOM). Imported RQC factors form aggregates inside the mitochondria in a process we term stalling-induced mitochondrial stress (SIMS). ABCE1 plays a crucial role in maintaining mitochondrial health during SIMS. Notably, cancer stem cells (CSCs) exhibit increased expression of ABCE1 and consequently are more resistant to EME-induced mitochondrial dysfunction. This points to a potential mechanism of drug resistance by CSCs. Our study highlights the significance of mitochondrial entrapment of RQC factors such as ABCE1 in determining the fate of cancer cells versus CSCs. Targeting ABCE1 or other RQC factors in translational inhibition cancer therapy may help overcome drug resistance.

Ribosome stalling during c-myc translation presents actionable cancer cell vulnerability.

Myc is a major driver of tumor initiation, progression, and maintenance. Up-regulation of Myc protein level rather than acquisition of neomorphic properties appears to underlie most Myc-driven cancers. Cellular mechanisms governing Myc expression remain incompletely defined. In this study, we show that ribosome-associated quality control (RQC) plays a critical role in maintaining Myc protein level. Ribosomes stall during the synthesis of the N-terminal portion of cMyc, generating aberrant cMyc species and necessitating deployment of the early RQC factor ZNF598 to handle translational stress and restore cMyc translation. ZNF598 expression is up-regulated in human glioblastoma (GBM), and its expression positively correlates with that of cMyc. ZNF598 knockdown inhibits human GBM neurosphere formation in cell culture and Myc-dependent tumor growth in vivo in Drosophila. Intriguingly, the SARS-COV-2-encoded translational regulator Nsp1 impinges on ZNF598 to restrain cMyc translation and consequently cMyc-dependent cancer growth. Remarkably, Nsp1 exhibits synthetic toxicity with the translation and RQC-related factor ATP-binding cassette subfamily E member 1, which, despite its normally positive correlation with cMyc in cancer cells, is co-opted by Nsp1 to down-regulate cMyc and inhibit tumor growth. Ribosome stalling during c-myc translation thus offers actionable cancer cell vulnerability.

Run, Ribosome, Run: From Compromised Translation to Human Health.

Significance: Translation is an essential cellular process, and diverse signaling pathways have evolved to deal with problems arising during translation. Erroneous stalls and unresolved ribosome collisions are implicated in many pathologies, including neurodegeneration and metabolic dysregulation. Recent Advances: Many proteins involved in detection and clearance of stalled and collided ribosomes have been identified and studied in detail. Ribosome profiling techniques have revealed extensive and nonprogrammed ribosome stalling and leaky translation into the 3' untranslated regions of mRNAs. Impairment of protein synthesis has been linked to aging in yeast and mice. Critical Issues: Ribosomes act as sensors of cellular states, but the molecular mechanisms, as well as physiological relevance, remain understudied. Most of our current knowledge stems from work in yeast and simple multicellular organisms such as Caenorhabditis elegans, while we are only beginning to comprehend the role of ribosome surveillance in higher organisms. As an example, the ribotoxic stress response, a pathway responding to global translational stress, has been studied mostly in response to small translation inhibitors and ribotoxins, and has only recently been explored in physiological settings. This review focuses on ribosome-surveillance pathways and their importance for cell and tissue homeostasis upon naturally occurring insults such as oxidative stress, nutrient deprivation, and viral infections. Future Directions: A better insight into the physiological roles of ribosome-surveillance pathways and their crosstalk could lead to an improved understanding of human pathologies and aging. Antioxid. Redox Signal. 39, 336-350.

Translational regulation by ribosome-associated quality control in neurodegenerative disease, cancer, and viral infection.

Translational control at the initiation, elongation, and termination steps exerts immediate effects on the rate as well as the spatiotemporal dynamics of new protein synthesis, shaping the composition of the proteome. Translational control is particularly important for cells under stress as during viral infection or in disease conditions such as cancer and neurodegenerative diseases. Much has been learned about the control mechanisms acting at the translational initiation step under normal or pathological conditions. However, problems during the elongation or termination steps of translation can lead to ribosome stalling and ribosome collision, which will trigger ribosome-associated quality control (RQC) mechanism. Inadequate RQC may lead to the accumulation of faulty translation products that perturb protein homeostasis (proteostasis). Proteostasis signifies a cellular state in which the synthesis, folding, and degradation of proteins are maintained at a homeostatic state such that an intact proteome is preserved. Cellular capacity to preserve proteostasis declines with age, which is thought to contribute to age-related diseases. Proteostasis failure manifested as formation of aberrant protein aggregates, epitomized by the amyloid plaques in Alzheimer's disease (AD), is a defining feature of neurodegenerative diseases. The root cause of the proteostasis failure and protein aggregation is still enigmatic. Here I will review recent studies supporting that faulty translation products resulting from inadequate RQC of translational stalling and ribosome collision during the translation of problematic mRNAs can be the root cause of proteostasis failure and may represent novel therapeutic targets for neurodegenerative diseases. I will also review evidence that translation regulation by RQC is operative in cancer cells and during viral infection. Better understanding of RQC mechanism may lead to novel therapeutic strategies against neurodegenerative diseases, cancer, and viral infections, including the ongoing COVID-19 pandemic.

mRNA and tRNA modification states influence ribosome speed and frame maintenance during poly(lysine) peptide synthesis.

Ribosome speed is dictated by multiple factors including substrate availability, cellular conditions, and product (peptide) formation. Translation slows during the synthesis of cationic peptide sequences, potentially influencing the expression of thousands of proteins. Available evidence suggests that ionic interactions between positively charged nascent peptides and the negatively charged ribosome exit tunnel impede translation. However, this hypothesis was difficult to test directly because of inability to decouple the contributions of amino acid charge from mRNA sequence and tRNA identity/abundance in cells. Furthermore, it is unclear if other components of the translation system central to ribosome function (e.g., RNA modification) influence the speed and accuracy of positively charged peptide synthesis. In this study, we used a fully reconstituted Escherichia coli translation system to evaluate the effects of peptide charge, mRNA sequence, and RNA modification status on the translation of lysine-rich peptides. Comparison of translation reactions on poly(lysine)-encoding mRNAs conducted with either Lys-tRNALys or Val-tRNALys reveals that that amino acid charge, while important, only partially accounts for slowed translation on these transcripts. We further find that in addition to peptide charge, mRNA sequence and both tRNA and mRNA modification status influence the rates of amino acid addition and the ribosome's ability to maintain frame (instead of entering the -2, -1, and +1 frames) during poly(lysine) peptide synthesis. Our observations lead us to expand the model for explaining how the ribosome slows during poly(lysine) peptide synthesis and suggest that posttranscriptional RNA modifications can provide cells a mechanism to precisely control ribosome movements along an mRNA.

Beilunmycin, a new virginiamycins antibiotic from mangrove-derived Streptomyces sp. 2BBP-J2 and the antibacterial activity by inhibiting protein translation.

One new virginiamycin derivative, 'beilunmycin' (1), and three known virginiamycin antibiotics, 16-hydroxy-virginiamycin M1 (2), virginiamycin M2 (3), and virginiamycin M1 (4), were isolated from the culture of a mangrove-derived endophytic Streptomyces sp. 2BBP-J2. The structures were characterized on the basis of their spectroscopic data, and the absolute configuration of 1 was established by ECD calculations. Compounds 1-4 exhibited antibacterial activities against Gram-positive bacteria, with MIC values in the range of 0.5-16 µg/ml. All the compounds demonstrated strong protein translation-stalling activity, with minimal concentrations detected with pDualrep2 in the range of 1.9-5.9 nmol.

Conserved Upstream Open Reading Frame Nascent Peptides That Control Translation.

Cells utilize transcriptional and posttranscriptional mechanisms to alter gene expression in response to environmental cues. Gene-specific controls, including changing the translation of specific messenger RNAs (mRNAs), provide a rapid means to respond precisely to different conditions. Upstream open reading frames (uORFs) are known to control the translation of mRNAs. Recent studies in bacteria and eukaryotes have revealed the functions of evolutionarily conserved uORF-encoded peptides. Some of these uORF-encoded nascent peptides enable responses to specific metabolites to modulate the translation of their mRNAs by stalling ribosomes and through ribosome stalling may also modulate the level of their mRNAs. In this review, we highlight several examples of conserved uORF nascent peptides that stall ribosomes to regulate gene expression in response to specific metabolites in bacteria, fungi, mammals, and plants.