scholarly journals Translation Elongation Factor 4 (LepA) Contributes to Tetracycline Susceptibility by Stalling Elongating Ribosomes

2018 ◽  
Vol 62 (8) ◽  
Author(s):  
Bin Liu ◽  
Chunlai Chen
Keyword(s):  
Protein Synthesis ◽  
Ribosome Biogenesis ◽  
Molecular Mechanisms ◽  
Elongation Factor ◽  
Ribosome Profiling ◽  
Translation Elongation Factor ◽  
Translation Elongation ◽  
Content Type ◽  
Elongation Phase ◽  
Profiling Analysis

ABSTRACTEven though elongation factor 4 (EF4) is the third most conserved protein in bacteria, its physiological functions remain largely unknown and its proposed molecular mechanisms are conflicting among previous studies. In the present study, we show that the growth of anEscherichia colistrain is more susceptible to tetracycline than its EF4 knockout strain. Consistent with previous studies, our results suggested that EF4 affects ribosome biogenesis when tetracycline is present. Through ribosome profiling analysis, we discovered that EF4 causes 1-nucleotide shifting of ribosomal footprints on mRNA when cells have been exposed to tetracycline. In addition, when tetracycline is present, EF4 inhibits the elongation of protein synthesis, which leads to the accumulation of ribosomes in the early segment of mRNA. Altogether, when cells are exposed to tetracycline, EF4 alters both ribosome biogenesis and the elongation phase of protein synthesis.

scholarly journals Translation elongation factor 2 depletion by siRNA in mouse liver leads to mTOR-independent translational upregulation of ribosomal protein genes

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Maxim V. Gerashchenko ◽  
Mikhail V. Nesterchuk ◽  
Elena M. Smekalova ◽  
Joao A. Paulo ◽  
Piotr S. Kowalski ◽  
...  
Keyword(s):  
Protein Synthesis ◽  
Mammalian Cells ◽  
Ribosomal Proteins ◽  
Mouse Liver ◽  
Negative Impact ◽  
Elongation Factor ◽  
Ribosome Profiling ◽  
Translation Elongation Factor ◽  
Translation Elongation ◽  
Elongation Factor 2

Abstract Due to breakthroughs in RNAi and genome editing methods in the past decade, it is now easier than ever to study fine details of protein synthesis in animal models. However, most of our understanding of translation comes from unicellular organisms and cultured mammalian cells. In this study, we demonstrate the feasibility of perturbing protein synthesis in a mouse liver by targeting translation elongation factor 2 (eEF2) with RNAi. We were able to achieve over 90% knockdown efficacy and maintain it for 2 weeks effectively slowing down the rate of translation elongation. As the total protein yield declined, both proteomics and ribosome profiling assays showed robust translational upregulation of ribosomal proteins relative to other proteins. Although all these genes bear the TOP regulatory motif, the branch of the mTOR pathway responsible for translation regulation was not activated. Paradoxically, coordinated translational upregulation of ribosomal proteins only occurred in the liver but not in murine cell culture. Thus, the upregulation of ribosomal transcripts likely occurred via passive mTOR-independent mechanisms. Impaired elongation sequesters ribosomes on mRNA and creates a shortage of free ribosomes. This leads to preferential translation of transcripts with high initiation rates such as ribosomal proteins. Furthermore, severe eEF2 shortage reduces the negative impact of positively charged amino acids frequent in ribosomal proteins on ribosome progression.

scholarly journals A Chemical Genomic Screen in Saccharomyces cerevisiae Reveals a Role for Diphthamidation of Translation Elongation Factor 2 in Inhibition of Protein Synthesis by Sordarin

2008 ◽  
Vol 52 (5) ◽  
pp. 1623-1629 ◽  
Author(s):  
Javier Botet ◽  
María Rodríguez-Mateos ◽  
Juan P. G. Ballesta ◽  
José Luis Revuelta ◽  
Miguel Remacha
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Protein Synthesis ◽  
Elongation Factor ◽  
Translation Elongation Factor ◽  
Cellular Mechanisms ◽  
Translation Elongation ◽  
Chemical Genomic ◽  
Elongation Factor 2 ◽  
Fungal Organisms ◽  
Inhibitor Of Protein Synthesis

ABSTRACT Sordarin and its derivatives are antifungal compounds of potential clinical interest. Despite the highly conserved nature of the fungal and mammalian protein synthesis machineries, sordarin is a selective inhibitor of protein synthesis in fungal organisms. In cells sensitive to sordarin, its mode of action is through preventing the release of translation elongation factor 2 (eEF2) during the translocation step, thus blocking protein synthesis. To further investigate the cellular components required for the effects of sordarin in fungal cells, we have used the haploid deletion collection of Saccharomyces cerevisiae to systematically identify genes whose deletion confers sensitivity or resistance to the compound. Our results indicate that genes in a number of cellular pathways previously unknown to play a role in sordarin response are involved in its growth effects on fungal cells and reveal a specific requirement for the diphthamidation pathway of cells in causing eEF2 to be sensitive to the effects of sordarin on protein synthesis. Our results underscore the importance of the powerful genomic tools developed in yeast (Saccharomyces cerevisiae) to more comprehensively understanding the cellular mechanisms involved in the response to therapeutic agents.

scholarly journals In vivo characterization of the role of tissue‐specific translation elongation factor 1 A 2 in protein synthesis reveals insights into muscle atrophy

FEBS Journal ◽  
2013 ◽  
Vol 280 (24) ◽  
pp. 6528-6540 ◽  
Author(s):  
Jennifer Doig ◽  
Lowri A. Griffiths ◽  
David Peberdy ◽  
Permphan Dharmasaroja ◽  
Maria Vera ◽  
...  
Keyword(s):  
Protein Synthesis ◽  
Muscle Atrophy ◽  
Elongation Factor ◽  
Translation Elongation Factor ◽  
Translation Elongation ◽  
In Vivo Characterization ◽  
Elongation Factor 1

scholarly journals Overexpression of Translation Elongation Factor 1A Affects the Organization and Function of the Actin Cytoskeleton in Yeast

Genetics ◽  
2001 ◽  
Vol 157 (4) ◽  
pp. 1425-1436
Author(s):  
Raj Munshi ◽  
Kimberly A Kandl ◽  
Anne Carr-Schmid ◽  
Johanna L Whitacre ◽  
Alison E M Adams ◽  
...  
Keyword(s):  
Protein Synthesis ◽  
Elongation Factor ◽  
Nucleotide Exchange ◽  
Translation Elongation Factor ◽  
Translation Elongation ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor ◽  
And Function

Abstract The translation elongation factor 1 complex (eEF1) plays a central role in protein synthesis, delivering aminoacyl-tRNAs to the elongating ribosome. The eEF1A subunit, a classic G-protein, also performs roles aside from protein synthesis. The overexpression of either eEF1A or eEF1Bα, the catalytic subunit of the guanine nucleotide exchange factor, in Saccharomyces cerevisiae results in effects on cell growth. Here we demonstrate that overexpression of either factor does not affect the levels of the other subunit or the rate or accuracy of protein synthesis. Instead, the major effects in vivo appear to be at the level of cell morphology and budding. eEF1A overexpression results in dosage-dependent reduced budding and altered actin distribution and cellular morphology. In addition, the effects of excess eEF1A in actin mutant strains show synthetic growth defects, establishing a genetic connection between the two proteins. As the ability of eEF1A to bind and bundle actin is conserved in yeast, these results link the established ability of eEF1A to bind and bundle actin in vitro with nontranslational roles for the protein in vivo.

scholarly journals Autophagy-competent mitochondrial translation elongation factor TUFM inhibits caspase-8-mediated apoptosis

2021 ◽  
Author(s):  
Chang-Yong Choi ◽  
Mai Tram Vo ◽  
John Nicholas ◽  
Young Bong Choi
Keyword(s):  
Molecular Mechanisms ◽  
Mitochondrial Dynamics ◽  
Elongation Factor ◽  
Productive Infection ◽  
Caspase 8 ◽  
Translation Elongation Factor ◽  
Mitochondrial Translation ◽  
Translation Elongation ◽  
Cell Functions ◽  
Induced Apoptosis

AbstractMitochondria support multiple cell functions, but an accumulation of dysfunctional or excessive mitochondria is detrimental to cells. We previously demonstrated that a defect in the autophagic removal of mitochondria, termed mitophagy, leads to the acceleration of apoptosis induced by herpesvirus productive infection. However, the exact molecular mechanisms underlying activation of mitophagy and regulation of apoptosis remain poorly understood despite the identification of various mitophagy-associated proteins. Here, we report that the mitochondrial translation elongation factor Tu, a mitophagy-associated protein encoded by the TUFM gene, locates in part on the outer membrane of mitochondria (OMM) where it acts as an inhibitor of altered mitochondria-induced apoptosis through its autophagic function. Inducible depletion of TUFM potentiated caspase-8-mediated apoptosis in virus-infected cells with accumulation of altered mitochondria. In addition, TUFM depletion promoted caspase-8 activation induced by treatment with TNF-related apoptosis-inducing ligand in cancer cells, potentially via dysregulation of mitochondrial dynamics and mitophagy. Importantly, we revealed the existence of and structural requirements for autophagy-competent TUFM on the OMM; the GxxxG motif within the N-terminal mitochondrial targeting sequences of TUFM was required for self-dimerization and mitophagy. Furthermore, we found that autophagy-competent TUFM was subject to ubiquitin-proteasome-mediated degradation but stabilized upon mitophagy or autophagy activation. Moreover, overexpression of autophagy-competent TUFM could inhibit caspase-8 activation. These studies extend our knowledge of mitophagy regulation of apoptosis and could provide a novel strategic basis for targeted therapy of cancer and viral diseases.

scholarly journals MicroRNA-33a-5p Modulates Japanese Encephalitis Virus Replication by Targeting Eukaryotic Translation Elongation Factor 1A1

2016 ◽  
Vol 90 (7) ◽  
pp. 3722-3734 ◽  
Author(s):  
Zheng Chen ◽  
Jing Ye ◽  
Usama Ashraf ◽  
Yunchuan Li ◽  
Siqi Wei ◽  
...  
Keyword(s):  
Viral Replication ◽  
Molecular Mechanisms ◽  
Elongation Factor ◽  
Negative Regulator ◽  
Translation Elongation Factor ◽  
Cellular Mirnas ◽  
Translation Elongation ◽  
Eukaryotic Translation ◽  
Acute Encephalitis ◽  
Replicase Complex

ABSTRACTJapanese encephalitis virus (JEV) is a typical mosquito-borne flavivirus responsible for acute encephalitis and meningitis in humans. However, the molecular mechanism for JEV pathogenesis is still unclear. MicroRNAs (miRNAs) are small noncoding RNAs that act as gene regulators. They are directly or indirectly involved in many cellular functions owing to their ability to target mRNAs for degradation or translational repression. However, how cellular miRNAs are regulated and their functions during JEV infection are largely unknown. In the present study, we found that JEV infection downregulated the expression of endogenous cellular miR-33a-5p. Notably, artificially transfecting with miR-33a-5p mimics led to a significant decrease in viral replication, suggesting that miR-33a-5p acts as a negative regulator of JEV replication. A dual-luciferase reporter assay identified eukaryotic translation elongation factor 1A1 (EEF1A1) as one of the miR-33a-5p target genes. Our study further demonstrated thatEEF1A1can interact with the JEV proteins NS3 and NS5 in replicase complex. Through this interaction,EEF1A1can stabilize the components of viral replicase complex and thus facilitates viral replication during JEV infection. Taken together, these results suggest that miR-33a-5p is downregulated during JEV infection, which contributes to viral replication by increasing the intracellular level ofEEF1A1, an interaction partner of JEV NS3 and NS5. This study provides a better understanding of the molecular mechanisms of JEV pathogenesis.IMPORTANCEMiRNAs are critical regulators of gene expression that utilize sequence complementarity to bind to and modulate the stability or translation efficiency of target mRNAs. Accumulating data suggest that miRNAs regulate a wide variety of molecular mechanisms in the host cells during viral infections. JEV, a neurotropic flavivirus, is one of the major causes of acute encephalitis in humans worldwide. The roles of cellular miRNAs during JEV infections are widely unexplored. The present study explores a novel role of miR-33a-5p as a negative regulator of JEV replication. We foundEEF1A1as a direct target of miR-33a-5p. We also demonstrated thatEEF1A1interacts with and stabilize the components of JEV replicase complex, which positively regulates JEV replication. These findings suggest a new insight into the molecular mechanism of JEV pathogenesis and provide a possible therapeutic entry point for viral encephalitis.

scholarly journals Comparison of Genomes of Brucella melitensis M28 and the B. melitensis M5-90 Derivative Vaccine Strain Highlights the Translation Elongation Factor Tu Genetuf2as an Attenuation-Related Gene

2013 ◽  
Vol 81 (8) ◽  
pp. 2812-2818 ◽  
Author(s):  
Fangkun Wang ◽  
Zujian Qiao ◽  
Sen Hu ◽  
Wenxing Liu ◽  
Huajun Zheng ◽  
...  
Keyword(s):  
Brucella Melitensis ◽  
Virulent Strain ◽  
Elongation Factor ◽  
Nucleotide Polymorphisms ◽  
Translation Elongation Factor ◽  
Translation Elongation ◽  
Content Type ◽  
Virulent Strains ◽  
Virulence Attenuation

ABSTRACTBrucella melitensiscauses brucellosis, a disease affecting sheep, cattle, and sometimes humans. AttenuatedB. melitensisstrain M5-90, derived from virulent strain M28, is widely used as a live vaccine in ruminants in China. Genetic differences between the strains may cast light on the mechanism of attenuation. We recently reported the complete genomic sequences of M28 and M5-90. Genome organization is highly conserved between these isolates, and also with virulent strains 16 M and ATCC 23457. Analysis revealed 23 open reading frames (ORFs) with consistent differences between M5-90 and the virulent strains. Notably, thetuf2gene encoding translation elongation factor EF-Tu from M5-90 contained 50 single nucleotide polymorphisms (SNPs) and 9 gaps (indels) compared totuf2of M28 or of the other virulent strains. There were no changes intuf1. To evaluate the potential role of EF-Tu in pathogenesis,tuf1andtuf2mutants of M28 and an M5-90 strain harboring wild-typetuf2were constructed, and their virulence/attenuation was evaluatedin vivo. We report that thetuf2gene plays an important role in the attenuation of M5-90 virulence.

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