scholarly journals DHX30 Coordinates Cytoplasmic Translation and Mitochondrial Function Contributing to Cancer Cell Survival

Cancers ◽  
2021 ◽  
Vol 13 (17) ◽  
pp. 4412
Author(s):  
Bartolomeo Bosco ◽  
Annalisa Rossi ◽  
Dario Rizzotto ◽  
Meriem Hadjer Hamadou ◽  
Alessandra Bisio ◽  
...  
Keyword(s):  
Ribosomal Proteins ◽  
Ribosome Biogenesis ◽  
Translational Efficiency ◽  
General Function ◽  
Translation Control ◽  
Mcf7 Cells ◽  
Mitochondrial Energy Metabolism ◽  
Cancer Cell Survival ◽  
The Impact ◽  
Global Translation

DHX30 was recently implicated in the translation control of mRNAs involved in p53-dependent apoptosis. Here, we show that DHX30 exhibits a more general function by integrating the activities of its cytoplasmic isoform and of the more abundant mitochondrial one. The depletion of both DHX30 isoforms in HCT116 cells leads to constitutive changes in polysome-associated mRNAs, enhancing the translation of mRNAs coding for cytoplasmic ribosomal proteins while reducing the translational efficiency of the nuclear-encoded mitoribosome mRNAs. Furthermore, the depletion of both DHX30 isoforms leads to higher global translation but slower proliferation and lower mitochondrial energy metabolism. Isoform-specific silencing supports a role for cytoplasmic DHX30 in modulating global translation. The impact on translation and proliferation was confirmed in U2OS and MCF7 cells. Exploiting RIP, eCLIP, and gene expression data, we identified fourteen mitoribosome transcripts we propose as direct DHX30 targets that can be used to explore the prognostic value of this mechanism in cancer. We propose that DHX30 contributes to cell homeostasis by coordinating ribosome biogenesis, global translation, and mitochondrial metabolism. Targeting DHX30 could, thus, expose a vulnerability in cancer cells.

scholarly journals DHX30 coordinates cytoplasmic translation and mitochondrial function contributing to cancer cell survival

2020 ◽  
Author(s):  
Bartolomeo Bosco ◽  
Annalisa Rossi ◽  
Dario Rizzotto ◽  
Sebastiano Giorgetta ◽  
Alicia Perzolli ◽  
...  
Keyword(s):  
Gene Expression ◽  
Energy Metabolism ◽  
Cancer Cells ◽  
Ribosomal Proteins ◽  
General Function ◽  
Translation Control ◽  
Mcf7 Cells ◽  
Cancer Types ◽  
The Impact ◽  
Global Translation

AbstractDHX30 was recently implicated in the translation control of mRNAs involved in p53-dependent apoptosis. Here we show that DHX30 exhibits a more general function by integrating the activities of its cytoplasmic isoform and of the more abundant mitochondrial one. The depletion of both DHX30 isoforms in HCT116 cells leads to constitutive changes in polysome-associated mRNAs, enhancing the translation of mRNAs coding for cytoplasmic ribosomal proteins while reducing the translational efficiency of the nuclear-encoded mitoribosome mRNAs. Furthermore, depletion of both DHX30 isoforms exhibits higher global translation but slower proliferation, and reduced mitochondrial energy metabolism. Isoform-specific silencing established a role for cytoplasmic DHX30 in modulating global translation. The impact on global translation and proliferation were confirmed in U2OS and MCF7 cells, although the effect of DHX30 depletion on mitochondrial gene expression was observed only in MCF7 cells. Exploiting RIP, eCLIP, and gene expression data, we identified a gene signature comprising DHX30 and fourteen mitoribosome transcripts that we candidate as direct targets: this signature shows prognostic value in several TCGA cancer types, with higher expression associated with reduced overall survival. We propose that DHX30 contributes to cell homeostasis by coordinating ribosome biogenesis, global translation, and mitochondrial metabolism. Targeting DHX30 could, thus, expose a vulnerability in cancer cells.Author summaryTranslation occurs in the cell both through cytoplasmic and mitochondrial ribosomes, respectively translating mRNAs encoded by the nuclear and the mitochondrial genome. Here we found that DHX30, an RNA-binding protein implicated in p53-dependent apoptosis, enhances the translation of mRNAs coding for cytoplasmic ribosomal proteins while reducing that of the mitoribosome mRNAs when silenced. This coordination of the cytoplasmic and mitochondrial translation machineries affected both cell proliferation and energy metabolism, suggesting an important role for this mechanism in determining the fitness of cancer cells. Indeed, the analysis of publicly available cancer datasets led us to define a 15-genes signature that is able to affect the prognosis of a subset of cancer types. In this subset, we found that higher expression of the genes composing the signature is associated with a worse prognosis. We thus propose DHX30 as a potential vulnerability in cancer cells, that could be targeted to develop novel therapeutic strategies.

scholarly journals MAF1 is a Chronic Repressor of RNA Polymerase III Transcription in the Mouse

2019 ◽  
Author(s):  
Nicolas Bonhoure ◽  
Viviane Praz ◽  
Robyn D. Moir ◽  
Gilles Willemin ◽  
François Mange ◽  
...  
Keyword(s):  
Energy Expenditure ◽  
Rna Polymerase ◽  
Ribosomal Proteins ◽  
Rna Polymerase Iii ◽  
Mrna Levels ◽  
Wild Type ◽  
Pol Iii ◽  
The Impact ◽  
Global Translation ◽  
Modest Effect

AbstractMaf1-/- mice are lean, obesity-resistant and metabolically inefficient. Their increased energy expenditure is thought to be driven by a futile RNA cycle that reprograms metabolism to meet an increased demand for nucleotides stemming from the deregulation of RNA polymerase (pol) III transcription. Metabolic changes consistent with this model have been reported in both fasted and refed mice, however the impact of the fasting-refeeding-cycle on pol III function has not been examined. Here we show that changes in pol III occupancy in the liver of fasted versus refed wild-type mice are largely confined to low and intermediate occupancy genes; high occupancy genes are unchanged. However, in Maf1-/- mice, pol III occupancy of the vast majority of active loci in liver and the levels of specific precursor tRNAs in this tissue and other organs are higher than wild-type in both fasted and refed conditions. Thus, MAF1 functions as a chronic repressor of active pol III loci and can modulate transcription under different conditions. Our findings support the futile RNA cycle hypothesis, elaborate the mechanism of pol III repression by MAF1 and demonstrate a modest effect of MAF1 on global translation via reduced mRNA levels and translation efficiencies for several ribosomal proteins.

scholarly journals The Impact of the Stringent Response on TRAFAC GTPases and Prokaryotic Ribosome Assembly

Cells ◽  
2019 ◽  
Vol 8 (11) ◽  
pp. 1313 ◽  
Author(s):  
Bennison ◽  
Irving ◽  
Corrigan
Keyword(s):  
Ribosomal Proteins ◽  
Cellular Response ◽  
Ribosome Biogenesis ◽  
Stringent Response ◽  
Protein Translation ◽  
Bound State ◽  
Ribosome Assembly ◽  
Rrna Transcription ◽  
Stress Sensing ◽  
The Impact

Many facets of ribosome biogenesis and function, including ribosomal RNA (rRNA) transcription, 70S assembly and protein translation, are negatively impacted upon induction of a nutrient stress-sensing signalling pathway termed the stringent response. This stress response is mediated by the alarmones guanosine tetra- and penta-phosphate ((p)ppGpp), the accumulation of which leads to a massive cellular response that slows growth and aids survival. The 70S bacterial ribosome is an intricate structure, with assembly both complex and highly modular. Presiding over the assembly process is a group of P-loop GTPases within the TRAFAC (Translation Factor Association) superclass that are crucial for correct positioning of both early and late stage ribosomal proteins (r-proteins) onto the rRNA. Often described as ‘molecular switches’, members of this GTPase superfamily readily bind and hydrolyse GTP to GDP in a cyclic manner that alters the propensity of the GTPase to carry out a function. TRAFAC GTPases are considered to act as checkpoints to ribosome assembly, involved in binding to immature sections in the GTP-bound state, preventing further r-protein association until maturation is complete. Here we review our current understanding of the impact of the stringent response and (p)ppGpp production on ribosome maturation in prokaryotic cells, focusing on the inhibition of (p)ppGpp on GTPase-mediated subunit assembly, but also touching upon the inhibition of rRNA transcription and protein translation.

Roles of the mammalian target of rapamycin, mTOR, in controlling ribosome biogenesis and protein synthesis

2012 ◽  
Vol 40 (1) ◽  
pp. 168-172 ◽  
Author(s):  
Valentina Iadevaia ◽  
Yilin Huo ◽  
Ze Zhang ◽  
Leonard J. Foster ◽  
Christopher G. Proud
Keyword(s):  
Protein Synthesis ◽  
Ribosomal Proteins ◽  
Ribosome Biogenesis ◽  
Kinase Inhibitors ◽  
Mammalian Target Of Rapamycin ◽  
Target Of Rapamycin ◽  
Energy Status ◽  
Stable Isotope Labelling ◽  
Mtor Kinase ◽  
The Impact

mTORC1 (mammalian target of rapamycin complex 1) is controlled by diverse signals (e.g. hormones, growth factors, nutrients and cellular energy status) and regulates a range of processes including anabolic metabolism, cell growth and cell division. We have studied the impact of inhibiting mTOR on protein synthesis in human cells. Partial inhibition of mTORC1 by rapamycin has only a limited impact on protein synthesis, but inhibiting mTOR kinase activity causes much greater inhibition of protein synthesis. Using a pulsed stable-isotope-labelling technique, we show that the rapamycin and mTOR (mammalian target of rapamycin) kinase inhibitors have differential effects on the synthesis of specific proteins. In particular, the synthesis of proteins encoded by mRNAs that have a 5′-terminal pyrimidine tract is strongly inhibited by mTOR kinase inhibitors. Many of these mRNAs encode ribosomal proteins. mTORC1 also promotes the synthesis of rRNA, although the mechanisms involved remain to be clarified. We found that mTORC1 also regulates the processing of the precursors of rRNA. mTORC1 thus co-ordinates several steps in ribosome biogenesis.

scholarly journals A translation control module coordinates germline stem cell differentiation with ribosome biogenesis during Drosophila oogenesis

2021 ◽  
Author(s):  
Elliot T Martin ◽  
Patrick Blatt ◽  
Elaine Ngyuen ◽  
Roni Lahr ◽  
Sangeetha Selvam ◽  
...  
Keyword(s):  
Stem Cell ◽  
Cell Differentiation ◽  
Ribosomal Proteins ◽  
Ribosome Biogenesis ◽  
Stem Cell Differentiation ◽  
Germline Stem Cell ◽  
Translation Control ◽  
Rna Helicases ◽  
Drosophila Oogenesis ◽  
Cycle Arrest

Ribosomal defects perturb stem cell differentiation, causing diseases called ribosomopathies. How ribosome levels control stem cell differentiation is not fully known. Here, we discovered three RNA helicases are required for ribosome biogenesis and for Drosophila oogenesis. Loss of these helicases, which we named Aramis, Athos and Porthos, lead to aberrant stabilization of p53, cell cycle arrest and stalled GSC differentiation. Unexpectedly, Aramis is required for efficient translation of a cohort of mRNAs containing a 5′-Terminal-Oligo-Pyrimidine (TOP)-motif, including mRNAs that encode ribosomal proteins and a conserved p53 inhibitor, Novel Nucleolar protein 1 (Non1). The TOP-motif co-regulates the translation of growth-related mRNAs in mammals. As in mammals, the La-related protein co-regulates the translation of TOP-motif containing RNAs during Drosophila oogenesis. Thus, a previously unappreciated TOP-motif in Drosophila responds to reduced ribosome biogenesis to co-regulate the translation of ribosomal proteins and a p53 repressor, thus coupling ribosome biogenesis to GSC differentiation.

TARGETING CYCLIN B1 WITH ANTISENSE OLIGONUCLETOTIDES- COATED SUPERPARAMAGNETIC IRON OXIDE NANOPARTICLES

2018 ◽  
Vol 6 (10) ◽  
Author(s):  
Hosam Zaghloul ◽  
Doaa A. Shahin ◽  
Ibrahim El- Dosoky ◽  
Mahmoud E. El-awady ◽  
Fardous F. El-Senduny ◽  
...  
Keyword(s):  
Iron Oxide ◽  
Iron Oxide Nanoparticles ◽  
Cellular Uptake ◽  
Superparamagnetic Iron Oxide ◽  
Cyclin B1 ◽  
Superparamagnetic Iron Oxide Nanoparticles ◽  
Oxide Nanoparticles ◽  
Mcf7 Cells ◽  
M Phase ◽  
The Impact

Antisense oligonucleotides (ASO) represent an attractive trend as specific targeting molecules but sustain poor cellular uptake meanwhile superparamagnetic iron oxide nanoparticles (SPIONs) offer stability of ASO and improved cellular uptake. In the present work we aimed to functionalize SPIONs with ASO targeting the mRNA of Cyclin B1 which represents a potential cancer target and to explore its anticancer activity. For that purpose, four different SPIONs-ASO conjugates, S-M (1–4), were designated depending on the sequence of ASO and constructed by crosslinking carboxylated SPIONs to amino labeled ASO. The impact of S-M (1–4) on the level of Cyclin B1, cell cycle, ROS and viability of the cells were assessed by flowcytometry. The results showed that S-M3 and S-M4 reduced the level of Cyclin B1 by 35 and 36%, respectively. As a consequence to downregulation of Cyclin B1, MCF7 cells were shown to be arrested at G2/M phase (60.7%). S-M (1–4) led to the induction of ROS formation in comparison to the untreated control cells. Furthermore, S-M (1–4) resulted in an increase in dead cells compared to the untreated cells and SPIONs-treated cells. In conclusion, targeting Cyclin B1 with ASO-coated SPIONs may represent a specific biocompatible anticancer strategy.

scholarly journals Upregulation of 5′-terminal oligopyrimidine mRNA translation upon loss of the ARF tumor suppressor

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Kyle A. Cottrell ◽  
Ryan C. Chiou ◽  
Jason D. Weber
Keyword(s):  
Protein Synthesis ◽  
Tumor Cells ◽  
Ribosomal Proteins ◽  
Human Cancer ◽  
Mrna Translation ◽  
Ribosome Profiling ◽  
Translational Efficiency ◽  
Gene Encoding ◽  
Terminal Oligopyrimidine ◽  
Ribosome Export

AbstractTumor cells require nominal increases in protein synthesis in order to maintain high proliferation rates. As such, tumor cells must acquire enhanced ribosome production. How the numerous mutations in tumor cells ultimately achieve this aberrant production is largely unknown. The gene encoding ARF is the most commonly deleted gene in human cancer. ARF plays a significant role in regulating ribosomal RNA synthesis and processing, ribosome export into the cytoplasm, and global protein synthesis. Utilizing ribosome profiling, we show that ARF is a major suppressor of 5′-terminal oligopyrimidine mRNA translation. Genes with increased translational efficiency following loss of ARF include many ribosomal proteins and translation factors. Knockout of p53 largely phenocopies ARF loss, with increased protein synthesis and expression of 5′-TOP encoded proteins. The 5′-TOP regulators eIF4G1 and LARP1 are upregulated in Arf- and p53-null cells.

scholarly journals Structural basis of GTPase-mediated mitochondrial ribosome biogenesis and recycling

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Hauke S. Hillen ◽  
Elena Lavdovskaia ◽  
Franziska Nadler ◽  
Elisa Hanitsch ◽  
Andreas Linden ◽  
...  
Keyword(s):  
Ribosomal Proteins ◽  
Ribosome Biogenesis ◽  
Ribosomal Subunit ◽  
Structural Basis ◽  
Peptidyl Transferase ◽  
Mitochondrial Ribosomes ◽  
Mitochondrial Ribosome ◽  
In Vitro Reconstitution

AbstractRibosome biogenesis requires auxiliary factors to promote folding and assembly of ribosomal proteins and RNA. Particularly, maturation of the peptidyl transferase center (PTC) is mediated by conserved GTPases, but the molecular basis is poorly understood. Here, we define the mechanism of GTPase-driven maturation of the human mitochondrial large ribosomal subunit (mtLSU) using endogenous complex purification, in vitro reconstitution and cryo-EM. Structures of transient native mtLSU assembly intermediates that accumulate in GTPBP6-deficient cells reveal how the biogenesis factors GTPBP5, MTERF4 and NSUN4 facilitate PTC folding. Addition of recombinant GTPBP6 reconstitutes late mtLSU biogenesis in vitro and shows that GTPBP6 triggers a molecular switch and progression to a near-mature PTC state. Additionally, cryo-EM analysis of GTPBP6-treated mature mitochondrial ribosomes reveals the structural basis for the dual-role of GTPBP6 in ribosome biogenesis and recycling. Together, these results provide a framework for understanding step-wise PTC folding as a critical conserved quality control checkpoint.

scholarly journals miR-16-5p Promotes Erythroid Maturation of Erythroleukemia Cells by Regulating Ribosome Biogenesis

2021 ◽  
Vol 14 (2) ◽  
pp. 137
Author(s):  
Christos I. Papagiannopoulos ◽  
Nikoleta F. Theodoroula ◽  
Ioannis S. Vizirianakis
Keyword(s):  
Ribosomal Proteins ◽  
Ribosome Biogenesis ◽  
Cultured Cells ◽  
Erythroid Differentiation ◽  
Underlying Mechanism ◽  
Loss Of Function ◽  
Functional Studies ◽  
Differentiated Cells ◽  
Erythroleukemia Cells ◽  
Murine Erythroleukemia

miRNAs constitute a class of non-coding RNA that act as powerful epigenetic regulators in animal and plant cells. In order to identify putative tumor-suppressor miRNAs we profiled the expression of various miRNAs during differentiation of erythroleukemia cells. RNA was purified before and after differentiation induction and subjected to quantitative RT-PCR. The majority of the miRNAs tested were found upregulated in differentiated cells with miR-16-5p showing the most significant increase. Functional studies using gain- and loss-of-function constructs proposed that miR-16-5p has a role in promoting the erythroid differentiation program of murine erythroleukemia (MEL) cells. In order to identify the underlying mechanism of action, we utilized bioinformatic in-silico platforms that incorporate predictions for the genes targeted by miR-16-5p. Interestingly, ribosome constituents, as well as ribosome biogenesis factors, were overrepresented among the miR-16-5p predicted gene targets. Accordingly, biochemical experiments showed that, indeed, miR-16-5p could modulate the levels of independent ribosomal proteins, and the overall ribosomal levels in cultured cells. In conclusion, miR-16-5p is identified as a differentiation-promoting agent in erythroleukemia cells, demonstrating antiproliferative activity, likely as a result of its ability to target the ribosomal machinery and restore any imbalanced activity imposed by the malignancy and the blockade of differentiation.

scholarly journals Genotype–phenotype correlations and novel molecular insights into the DHX30-associated neurodevelopmental disorders

2021 ◽  
Vol 13 (1) ◽  
Author(s):  
Ilaria Mannucci ◽  
Nghi D. P. Dang ◽  
Hannes Huber ◽  
Jaclyn B. Murry ◽  
Jeff Abramson ◽  
...  
Keyword(s):  
De Novo ◽  
Clinical Course ◽  
Neurodevelopmental Disorder ◽  
Global Developmental Delay ◽  
Helicase Activity ◽  
Speech Impairment ◽  
Human Phenotype ◽  
Missense Variants ◽  
The Impact ◽  
Global Translation

Abstract Background We aimed to define the clinical and variant spectrum and to provide novel molecular insights into the DHX30-associated neurodevelopmental disorder. Methods Clinical and genetic data from affected individuals were collected through Facebook-based family support group, GeneMatcher, and our network of collaborators. We investigated the impact of novel missense variants with respect to ATPase and helicase activity, stress granule (SG) formation, global translation, and their effect on embryonic development in zebrafish. SG formation was additionally analyzed in CRISPR/Cas9-mediated DHX30-deficient HEK293T and zebrafish models, along with in vivo behavioral assays. Results We identified 25 previously unreported individuals, ten of whom carry novel variants, two of which are recurrent, and provide evidence of gonadal mosaicism in one family. All 19 individuals harboring heterozygous missense variants within helicase core motifs (HCMs) have global developmental delay, intellectual disability, severe speech impairment, and gait abnormalities. These variants impair the ATPase and helicase activity of DHX30, trigger SG formation, interfere with global translation, and cause developmental defects in a zebrafish model. Notably, 4 individuals harboring heterozygous variants resulting either in haploinsufficiency or truncated proteins presented with a milder clinical course, similar to an individual harboring a de novo mosaic HCM missense variant. Functionally, we established DHX30 as an ATP-dependent RNA helicase and as an evolutionary conserved factor in SG assembly. Based on the clinical course, the variant location, and type we establish two distinct clinical subtypes. DHX30 loss-of-function variants cause a milder phenotype whereas a severe phenotype is caused by HCM missense variants that, in addition to the loss of ATPase and helicase activity, lead to a detrimental gain-of-function with respect to SG formation. Behavioral characterization of dhx30-deficient zebrafish revealed altered sleep-wake activity and social interaction, partially resembling the human phenotype. Conclusions Our study highlights the usefulness of social media to define novel Mendelian disorders and exemplifies how functional analyses accompanied by clinical and genetic findings can define clinically distinct subtypes for ultra-rare disorders. Such approaches require close interdisciplinary collaboration between families/legal representatives of the affected individuals, clinicians, molecular genetics diagnostic laboratories, and research laboratories.

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