scholarly journals Pseudouridine-Free Ribosome Exhibits Distinct Inter-Subunit Movements

2021 ◽  
Author(s):  
Yu Zhao ◽  
Jay Rai ◽  
Hongguo Yu ◽  
Hong Li
Keyword(s):  
Free Ribosome

scholarly journals Polyribonucleotide synthesis by subfractions of microsomes from rat liver

1968 ◽  
Vol 109 (2) ◽  
pp. 229-238 ◽  
Author(s):  
N. M. Wilkie ◽  
R. M. S. Smellie
Keyword(s):  
Acetic Acid ◽  
Rat Liver ◽  
Transfer Rna ◽  
Light Fraction ◽  
Free Ribosome ◽  
Polyadenylic Acid ◽  
Insoluble Material ◽  
Polyuridylic Acid ◽  
Microsome Fraction

1. The microsome fraction of rat liver has been fractionated and the ability of the fractions to incorporate ribonucleotides into polyribonucleotides has been studied. Activity was found in the rough-surfaced vesicle (light) fraction and in the free-ribosome fraction and this latter activity has been examined. 2. The free-ribosome fraction contains ribosome monomers, dimers and trimers together with some higher oligomers and ferritin. In addition to catalysing the incorporation of ribonucleotides into acid-insoluble material it contains diesterase activity. It catalyses the incorporation of UMP from UTP, but not UDP, AMP from ATP and CMP from CTP into polyribonucleotide material, and for UTP the product appears to be a homopolymer not more than eight units long attached to the ends of primer polyribonucleotide strands. 3. The activity could not be removed from the free-ribosome fraction by washing or by isolation in the presence of ethylenediaminetetra-acetic acid. 4. Partially hydrolysed polyuridylic acid but not polyadenylic acid could serve as a primer for the incorporation of UMP, but some activity was always associated with an endogenous primer. 5. Analysis of RNA extracted from the free-ribosome fraction after incubation with [3H]UTP showed the presence of 28s, 18s, 5s and transfer RNA types, but no radioactivity was associated with any of these RNA fractions.

scholarly journals Contamination of free-ribosome fractions by detached previously membrane-bound ribosomes (Short Communication)

1974 ◽  
Vol 138 (2) ◽  
pp. 305-307 ◽  
Author(s):  
K. O'Toole
Keyword(s):  
Rat Liver ◽  
Membrane Fraction ◽  
Short Communication ◽  
Free Ribosome ◽  
Membrane Bound ◽  
Free Polyribosome

A rough-membrane fraction isolated from rat liver by a procedure designed to prevent membrane denaturation was subjected to the gradient treatment normally used to isolate free ribosomes. Under these conditions, at most 20% of the ribosomes were detached from membrane with less than 5% sedimenting into the free-polyribosome pellet.

scholarly journals Generation and Selection of a Panel of Pan-Filovirus Single-Chain Antibodies using Cell-Free Ribosome Display

2018 ◽  
Author(s):  
Adinarayana Kunamneni ◽  
Elizabeth C. Clarke ◽  
Chunyan Ye ◽  
Steven B. Bradfute ◽  
Ravi Durvasula
Keyword(s):  
Hemorrhagic Fever ◽  
Cross Reactivity ◽  
Free Ribosome ◽  
Rapid Diagnostics ◽  
Single Chain ◽  
Ribosome Display ◽  
Single Chain Antibodies ◽  
Single Chain Fv ◽  
New Generation ◽  
Selection Of

AbstractFiloviruses, which include ebolaviruses and marburgvirus, can cause outbreaks of highly lethal hemorrhagic fever. This disease causes significant morbidity and mortality in humans and non-human primates, with human fatality rates reaching 90% during some outbreaks. Currently, there are a lack of licensed vaccines or antivirals for these viruses. Since early symptoms of filovirus infection mimic more common diseases, there is a strong unmet public health and biodefense need for broad-spectrum filovirus rapid diagnostics. We have generated a panel of mouse single-chain Fv-antibodies (scFvs) to filovirus glycoproteins (GPs) using cell-free ribosome display and determined their cross-reactivity profiles to all known filovirus species. Two scFvs (4-2 and 22-1) were able to detect all known Ebolavirus and Marburgvirus species. This is the first report on ribosome display scFvs that can detect a broad set of filovirus GPs, which demonstrates their potential use in the development of a new generation of rapid diagnostic immunoassays.

scholarly journals An integrated genome-wide multi-omics analysis of gene expression dynamics in the preimplantation mouse embryo

2018 ◽  
Author(s):  
Steffen Israel ◽  
Mathias Ernst ◽  
Olympia E. Psathaki ◽  
Hannes C. A. Drexler ◽  
Ellen Casser ◽  
...  
Keyword(s):  
Mouse Embryo ◽  
Developmental Stages ◽  
Protein Complexes ◽  
Large Fraction ◽  
Transcript Level ◽  
Free Ribosome ◽  
Translational Machinery ◽  
Preimplantation Mouse ◽  
Morula Stage ◽  
The Impact

AbstractEarly mouse embryos have an atypical translational machinery comprised of cytoplasmic lattices, poorly competent for translation. Thus, the impact of transcriptomic changes on the operational levels of proteins has likely been overestimated in the past. To find out, we used liquid chromatography–tandem mass spectrometry to detect and quantify 6,550 proteins in the oocyte and in six developmental stages (from zygote to blastocyst) collected in triplicates, and we also performed mRNA sequencing.In contrast to the known split between the 2-cell and 4-cell stages at the transcript level, on the protein level the oocyte-to-embryo transition appeared to last until the morula stage. In general, protein abundance profiles were weakly correlated with those of their cognate mRNAs and we found little or no concordance between changes in protein and transcript expression relative to the oocyte at early stages. However, concordance increased towards morula and blastocyst, hinting at a more direct coupling of proteins with transcripts at these stages, in agreement with the increase in free ribosome abundance. Independent validation by immunofluorescence and qPCR confirmed the existence of genes featuring strongly positively and negatively correlated protein and transcript. Moreover, consistent coverage of most known protein complexes indicates that our dataset represents a large fraction of the expressed proteome. Finally, we identified 20 markers, including members of the endoplasmic reticulum pathway, for discriminating between early and late stages.This resource contributes towards closing the gap between the ‘predicted’ phenotype, based on mRNA, and the ‘actual’ phenotype, based on protein, of the mouse embryo.

scholarly journals Free Ribosome

2020 ◽  
Author(s):  
Keyword(s):  
Free Ribosome

scholarly journals Pseudouridine-free Ribosome Exhibits Distinct Inter-subunit Movements

2021 ◽  
Author(s):  
Yu Zhao ◽  
Jay Rai ◽  
Hong-Guo Yu ◽  
Hong Li
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cancer Predisposition ◽  
Rna Modification ◽  
Structural Basis ◽  
Free Ribosome ◽  
Electron Cryomicroscopy ◽  
Pseudouridine Synthase ◽  
Phosphate Backbone ◽  
Close Contacts

Pseudouridine, the most abundant form of RNA modification, is known to play important roles in ribosome function. Mutations in human DKC1, the pseudouridine synthase responsible for catalyzing the ribosome RNA modification, cause translation deficiencies and are associated with a complex cancer predisposition. The structural basis for how pseudouridine impacts ribosome function remains uncharacterized. Here we report electron cryomicroscopy structures of a fully modified and a pseudouridine-free ribosome from Saccharomyces cerevisiae. In the modified ribosome, the rearranged N1 atom of pseudouridine is observed to stabilize key functional motifs by establishing predominately water-mediated close contacts with the phosphate backbone. The pseudouridine-free ribosome, however, is devoid of such interactions and displays conformations reflective of abnormal inter-subunit movements. The erroneous motions of the pseudouridine-free ribosome may explain its observed deficiencies in translation.

scholarly journals Single-Dose P2 X4R Single-Chain Fragment Variable Antibody Permanently Reverses Chronic Pain in Male Mice

2021 ◽  
Vol 22 (24) ◽  
pp. 13612
Author(s):  
Karin N. Westlund ◽  
Marena A. Montera ◽  
Aleyah E. Goins ◽  
Sascha R. A. Alles ◽  
Nikita Suri ◽  
...  
Keyword(s):  
Chronic Pain ◽  
Neuropathic Pain ◽  
Single Dose ◽  
Recombinant Antibody ◽  
Feature Binding ◽  
Peptide Sequence ◽  
Free Ribosome ◽  
Male Mice ◽  
Single Chain ◽  
Chronic Neuropathic Pain

Non-opioid single-chain variable fragment (scFv) small antibodies were generated as pain-reducing block of P2X4R receptor (P2X4R). A panel of scFvs targeting an extracellular peptide sequence of P2X4R was generated followed by cell-free ribosome display for recombinant antibody selection. After three rounds of bio-panning, a panel of recombinant antibodies was isolated and characterized by ELISA, cross-reactivity analysis, and immunoblotting/immunostaining. Generated scFv antibodies feature binding activity similar to monoclonal antibodies but with stronger affinity and increased tissue penetrability due to their ~30% smaller size. Two anti-P2X4R scFv clones (95, 12) with high specificity and affinity binding were selected for in vivo testing in male and female mice with trigeminal nerve chronic neuropathic pain (FRICT-ION model) persisting for several months in untreated BALBc mice. A single dose of P2X4R scFv (4 mg/kg, i.p.) successfully, completely, and permanently reversed chronic neuropathic pain-like measures in male mice only, providing retention of baseline behaviors indefinitely. Untreated mice retained hypersensitivity, and developed anxiety- and depression-like behaviors within 5 weeks. In vitro P2X4R scFv 95 treatment significantly increased the rheobase of larger-diameter (>25 µm) trigeminal ganglia (TG) neurons from FRICT-ION mice compared to controls. The data support use of engineered scFv antibodies as non-opioid biotherapeutic interventions for chronic pain.

Secondary Resistance to Lenalidomide in Del(5q) MDS Is Associated with CDC25C & PP2A Overexpression.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 292-292 ◽  
Author(s):  
Alan F List ◽  
Kathy Rocha ◽  
Ling Zhang ◽  
Rami S. Komrokji ◽  
Justine Clark ◽  
...  
Keyword(s):  
Gene Expression ◽  
Drug Resistance ◽  
Board Of Directors ◽  
Nuclear Export ◽  
Research Funding ◽  
Free Ribosome ◽  
Nuclear Staining ◽  
Advisory Committees ◽  
Secondary Resistance ◽  
Cytoplasmic Staining

Abstract Abstract 292 Background: Allelic deficiency for the RPS14 gene impairs differentiation and survival of erythroid progenitors in del(5q) MDS (Nature 2008; 451:335). Nucleolar stress arising from disruption of ribosome assembly fosters MDM2 sequestration by free ribosome components resulting in p53 stabilization and erythroid hypoplasia (Nat Cell Biol 2009; 11:501). We recently reported that reduced gene dosage of the lenalidomide (LEN) inhibitable, haplodeficient phosphatases CDC25C and PP2Acα is a key determinant of drug sensitivity in del(5q) MDS (PNAS 2009; 106: 12974). We now show that shRNA suppression of these genes to levels commensurate with haplodeficiency reinforces p53 accumulation, and that treatment with LEN promotes MDM2-mediated p53 degradation to transition del(5q) clones to G2/M arrest. We hypothesized that emergence of resistance to LEN in del(5q) MDS arises from two possible mechanisms: (1) up-regulation of haplodeficient drug targets or compensatory isotypes, or (2) inactivating mutations of the TP53 or CDC25C genes. Methods: To investigate mechanisms of LEN resistance, we studied sequential bone marrow (BM) specimens obtained at baseline (BL), response to treatment (TR) and treatment failure (TF) from 12 LEN treated patients with Low/INT-1 risk, transfusion-dependent del(5q) MDS. Eleven patients achieved clonal suppression and transfusion independence; 7 patients developed clinical drug resistance with primary clonal recovery. Immunohistochemical (IHC) staining for cdc25-C, -A and -B; PP2A–Ca and p53 were performed using a biotin-streptavidin-horseradish peroxidase method and compared to 6 age-matched controls; intensity of cytoplasmic or nuclear staining in hematopoietic elements was recorded after blinded review. DNA and RNA were extracted from cryopreserved BM mononuclear cells (BM-MNC) or fixed paraffin blocks from BM clot and biopsy sections. Expression of CDC25C splice variants was assessed by RT-PCR and total gene expression by real time (QT)-PCR. Exonic DNA encoding the catalytic [exons 8–14] and nuclear export domains [exon 11] of CDC25C and the DNA-binding domain of TP53 [exons 4–9] was sequenced for gene mutation analysis. Differences in mean values were compared by paired t-test. Results: P53 immunostaining was significantly higher in del(5q) BL specimens compared to controls ( relative expression [RE] 9.6 vs. 0.25; P =0.007). An admixture of nuclear and cytoplasmic staining for p53 and each cdc25 isotype was observed at BL that was largely restricted to erythroid precursors, whereas at TR cdc25-C and -A expression was primarily cytoplasmic, consistent with drug-induced nuclear exclusion. At TR, RE of only cdc25C (BL, 75 vs. TR, 49; P=0.05) and PP2A (29.2 vs. 12.3; P=0.025) was significantly reduced; whereas at TF cdc25C (TR, 43 vs. TF, 166; P=0.003), cdc25A (42.4 vs. 150; P=0.006), PP2A (7.3 vs. 65.6; P=0.028) and p53 (0.92 vs. 25.4; P=0.024) RE significantly increased. Nuclear localization of cdc25C and p53 but not cdc25A predominated at TF, consistent with escape from cdc25C inhibition. QT-PCR confirmed transcriptional up-regulation of CDC25C at TF with a mean 8.8-fold increase in gene expression vs. BL. DNA sequencing revealed no acquisition of somatic mutations within the CDC25C and TP53 exons studied [n=5]. Conclusions: Secondary resistance to LEN in del(5q) MDS is associated with over-expression and activation of the haplodeficient drug-inhibitable phosphatases, cdc25C and PP2A, with consequent restoration of wt-p53 activation. Absence of gene mutations within the coding exons analyzed suggests that transcriptional compensation alone is responsible for drug resistance. Novel agents targeting transcriptional repression of CDC25C may restore LEN sensitivity and merit investigation in drug resistant del(5q) MDS. Disclosures: List: Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau. Komrokji:Celgene: Research Funding, Speakers Bureau. Lancet:Celgene: Research Funding. Maciejewski:Esai: Membership on an entity's Board of Directors or advisory committees; Celgene: Speakers Bureau. Sekeres:Celgene: Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau.

Ribonucleic acid degrading activity associated with ribosomes from spinach leaf chloroplasts

1972 ◽  
Vol 50 (4) ◽  
pp. 691-695 ◽  
Author(s):  
Richard C. Howe ◽  
Donald J. Ursino
Keyword(s):  
Ribonucleic Acid ◽  
Component Part ◽  
Free Ribosome ◽  
Spinach Leaf ◽  
Dissociated State ◽  
Spinach Leaves

Ribosomes isolated from the chloroplasts of spinach leaves show the presence of RNA-degrading activity. The activity is optimal at pH 6.0 and at temperatures between 37 and 40 °C. The activity is cation-independent, exhibits exonuclease properties, and is considerably more pronounced under conditions in which the ribosome particles are in the dissociated state. The results are interpreted as showing that the RNA-degrading activity is a component part of the free ribosome population in the chloroplast.

scholarly journals Identification and functional characterization of a fish-specific tlr19 in common carp (Cyprinus carpio L.) that recruits TRIF as an adaptor and induces ifn expression during the immune response

2021 ◽  
Vol 52 (1) ◽  
Author(s):  
Shijuan Shan ◽  
Rongrong Liu ◽  
Hanxiao Feng ◽  
Fei Meng ◽  
Muhanmmad Aizaz ◽  
...  
Keyword(s):  
Common Carp ◽  
Cyprinus Carpio ◽  
Critical Role ◽  
Functional Characterization ◽  
Head Kidney ◽  
Poly I:C ◽  
Future Research ◽  
Free Ribosome ◽  
Early Endosomes ◽  
Localization Analysis

AbstractToll-like receptor 19 (Tlr19) is a fish-specific TLR that plays a critical role in innate immunity. In the present study, we aimed to identify tlr19 from common carp (Cyprinus carpio L.) and explored its expression profile, localization, adaptor, and signaling pathways. A novel tlr19 cDNA sequence (Cctlr19) was identified in common carp. Phylogenetic analysis revealed that CcTlr19 was most closely related to Danio rerio Tlr19. Subcellular localization analysis indicates that CcTlr19 was synthesized in the free ribosome and then transported to early endosomes. Cctlr19 was constitutively expressed in all the examined tissues, with the highest expression in the brain. After poly(I:C) and Aeromonas hydrophila injection, the expression of Cctlr19 was significantly upregulated in immune-related organs. In addition, the expression of Cctlr19 was upregulated in head kidney leukocytes (HKL) upon stimulation with different ligands. Immunofluorescence and luciferase analyses indicate that CcTlr19 recruited TRIF as an adaptor. Furthermore, CcTlr19 can activate the expression of ifn-1 and viperin. Taken together, these findings lay the foundation for future research to investigate the mechanisms underlying fish tlr19.

Sign in / Sign up

Export Citation Format

Share Document