Polysomes bypass a 50 nucleotide coding gap less efficiently than monosomes due to attenuation of an unstable 5’ mRNA stem loop stimulator and enhanced drop-off
Ontology highlight
ABSTRACT:
ORGANISM(S): Tequatrovirus T4 Escherichia coli
PROVIDER: GSE146240 | GEO | 2020/06/01
REPOSITORIES: GEO
Similar Datasets
Project description:Efficient translational bypassing of a 50-nt non-coding gap in a phage T4 topoisomerase subunit gene (gp60) requires several recoding signals. Here we investigate the function of the mRNA stem-loop 5' of the take-off codon, as well as the importance of ribosome loading density on the mRNA for efficient bypassing. We show that polysomes are less efficient at mediating bypassing than monosomes, both in vitro and in vivo, due to their preventing formation of a stem-loop 5' of the take-off codon and allowing greater peptidyl-tRNA drop off. A ribosome profiling analysis of phage T4-infected Escherichia coli yielded protected mRNA fragments within the normal size range derived from ribosomes stalled at the take-off codon. However, ribosomes at this position also yielded some 53-nucleotide fragments, 16 longer. These were due to protection of the nucleotides that form the 5' stem-loop. NMR shows that the 5' stem-loop is highly dynamic. The importance of different nucleotides in the 5' stem-loop is revealed by mutagenesis studies. These data highlight the significance of the 5' stem-loop for the 50-nt bypassing and further enhance appreciation of relevance of the extent of ribosome loading for recoding.
| S-EPMC7245268 | biostudies-literature
Project description:Polysomes bypass a 50 nucleotide coding gap less efficiently than monosomes due to attenuation of an unstable 5’ mRNA stem loop stimulator and enhanced drop-off
| PRJNA609751 | ENA
Project description:We investigated the association of cryptic unstable transcripts (CUTs) to ribosomes in Saccharomyces cerevisiae. We used 5’cap-sequence followed by sucrose gradient fractionation of polyribosome fractions after ultracentrifugation to assess the relative association of transcripts to the ribosomes. We used 5PSeq approach, which measures ribosome dynamics by sequencing the presence of co-translation mRNA degradation intermediates, to assess if cryptic transcripts are engaged in active translation. We performed 5PSeq after glucose depletion to increase the number of ribosomes stop at the 5’UTR and start codon.
2019-10-01 | GSE119134 | GEO
Project description:miRNAs moderately inhibit the translation and enhance the degradation of their target mRNAs via cognate binding sites located predominantly in the 3'-untranslated regions (UTR). Paradoxically, miRNA targets are also polysome-associated. We studied the polysome association by the comparative translationally less-active light- and more-active heavy-polysome profiling of a wild type (WT) human cell line and its isogenic mutant (MT) with a disrupted DICER1 gene and, thus, mature miRNA production. As expected, the open reading frame (ORF) length is a major determinant of light- to heavy-polysome mRNA abundance ratios, but is rendered less powerful in WT than in MT cells by miRNA-regulatory activities. We also observed that miRNAs tend to target mRNAs with longer ORFs, and that adjusting the mRNA abundance ratio with the ORF length improves its correlation with the 3'-UTR miRNA-binding-site count. In WT cells, miRNA-targeted mRNAs exhibit higher abundance in light relative to heavy polysomes, i.e., light-polysome enrichment. In MT cells, the DICER1 disruption not only significantly abrogated the light-polysome enrichment, but also narrowed the mRNA abundance ratio value range. Additionally, the abrogation of the enrichment due to the DICER1 gene disruption, i.e., the decreases of the ORF-length-adjusted mRNA abundance ratio from WT to MT cells, exhibits a nearly perfect linear correlation with the 3'-UTR binding-site count. Transcription factors and protein kinases are the top two most enriched mRNA groups. Taken together, the results provide evidence for the light-polysome enrichment of miRNA-targeted mRNAs to reconcile polysome association and moderate translation inhibition, and that ORF length is an important, though currently under-appreciated, transcriptome regulation parameter.
| S-EPMC10741098 | biostudies-literature
Project description:Historically, ribosomes have been viewed as unchanged homogeneous macromolecular machines with no intrinsic regulatory capacity for mRNA translation. However, an emerging concept is that heterogeneity of ribosomal composition exists, which can exert a regulatory function or specificity in translational control. This is supported by recent discoveries identifying compositionally distinct ‘specialized ribosomes’ that actively regulate mRNA translation. Viruses lack their own translational machinery and impose a high translational demand on the host cell during replication. Whilst viruses have evolved a variety of mechanisms to overcome this, we hypothesize that some viruses manipulate the host cell to produce specialized ribosomes to preferentially translate viral transcripts. Quantitative proteomic analysis has identified changes in the stoichiometry and composition of precursor ribosomal complexes during the switch from latent to lytic KSHV replication. Intriguingly, we demonstrate the enhanced association of ribosomal biogenesis factors BUD23 and NOC4L, and a previously uncharacterized KSHV lytic protein, ORF11, with small ribosomal subunit precursor complexes during lytic KSHV infection. Notably, BUD23 depletion resulted in significantly reduced viral gene expression and progression through the lytic cascade, culminating in a dramatic reduction of infectious virion production. Importantly, ribosome profiling demonstrated that BUD23 is essential for the reduced association of ribosomes with KSHV uORFs in late lytic genes, required for the efficient translation of the main open reading frame. Together our results provide new mechanistic insight into KSHV-mediated manipulation of cellular ribosome composition inducing a population of specialized ribosomes to facilitate efficient translation of viral mRNAs.
2022-12-14 | GSE199095 | GEO
Project description:Drop foot is a frequent abnormality in gait after central nervous system lesions. Different treatment strategies are available to functionally restore dorsal extension during swing phase in gait. Orthoses as well as surface and implantable devices for electrical stimulation of the peroneal nerve may be used in patients who do not regain good dorsal extension. While several studies investigated the effects of implanted systems on walking speed and gait endurance, only a few studies have focussed on the system's impact on kinematics and long-term outcomes. Therefore, our aim was to further investigate the effects of the implanted system ActiGait on gait kinematics and spatiotemporal parameters for the first time with a 1-year follow-up period. 10 patients were implanted with an ActiGait stimulator, with 8 patients completing baseline and follow-up assessments. Assessments included a 10-m walking test, video-based gait analysis and a Visual Analogue Scale (VAS) for health status. At baseline, gait analysis was performed without any assistive device as well as with surface electrical stimulation. At follow-up patients walked with the ActiGait system switched off and on. The maximum dorsal extension of the ankle at initial contact increased significantly between baseline without stimulation and follow-up with ActiGait (p = 0.018). While the spatio-temporal parameters did not seem to change much with the use of ActiGait in convenient walking speed, patients did walk faster when using surface stimulation or ActiGait compared to no stimulation at the 10-m walking test at their fastest possible walking speed. Patients rated their health better at the 1-year follow-up. In summary, a global improvement in gait kinematics compared to no stimulation was observed and the long-term safety of the device could be confirmed.
| S-EPMC6469760 | biostudies-literature
Project description:mRNAs bound by ribosomes from yeast cells were analysed in order to determine the exact position of ribosomes in the presence or absence of Rio1p. Beside total Ribosome Protected Fragments (RPFs), RPFs from mRNAs protected by immature pre-40S pre-ribosomes was also analysed. The analysis showed that immature 40S ribosomes can carry out translation and their premature entry into translation is hindered by Rio1p.
2017-08-16 | GSE98333 | GEO
Project description:A foot drop stimulator (FDS) is a rehabilitation intervention that stimulates the common peroneal nerve to facilitate ankle dorsiflexion at the appropriate time during post-stroke hemiplegic gait. Time-frequency analysis (TFA) of non-stationary surface electromyograms (EMG) and spectral variables such as instantaneous mean frequency (IMNF) can provide valuable information on the long-term effects of FDS intervention in terms of changes in the motor unit (MU) recruitment during gait, secondary to improved dorsiflexion.The aim of this study was to apply a wavelet-based TFA approach to assess the changes in neuromuscular activation of the tibialis anterior (TA), soleus (SOL), and gastrocnemius (GA) muscles after utilization of an FDS during gait post-stroke.Surface EMG were collected bilaterally from the TA, SOL, and GA muscles from six participants (142.9?±?103.3?months post-stroke) while walking without the FDS at baseline and 6?months post-FDS utilization. Continuous wavelet transform was performed to get the averaged time-frequency distribution of band pass filtered (20-300?Hz) EMGs during multiple walking trials. IMNFs were computed during normalized gait and were averaged during the stance and swing phases. Percent changes in the energies associated with each frequency band of 25?Hz between 25 and 300?Hz were computed and compared between visits.Averaged time-frequency representations of the affected TA, SOL, and GA EMG show altered spectral attributes post-FDS utilization during normalized gait. The mean IMNF values for the affected TA were significantly lower than the unaffected TA at baseline (p?=?0.026) and follow-up (p?=?0.038) during normalized stance. The mean IMNF values significantly increased (p?=?0.017) for the affected GA at follow-up during normalized swing. The frequency band of 250-275?Hz significantly increased in the energies post-FDS utilization for all muscles.The application of wavelet-based TFA of EMG and outcome measures (IMNF, energy) extracted from the time-frequency distributions suggest alterations in MU recruitment strategies after the use of FDS in individuals with chronic stroke. This further establishes the efficacy of FDS as a rehabilitation intervention that may promote motor recovery in addition to treating the secondary complications of foot drop due to post-stroke hemiplegia.
| S-EPMC5581808 | biostudies-literature
Project description:Detection of cryptic unstable transcripts associated with ribosomes in yeast
| PRJNA488250 | ENA
Project description:Accurate positional information concerning ribosomes and RNA binding proteins with respect to their transcripts is important to understand the global regulatory network underlying protein and RNA fate in living cells. Most footprinting approaches generate RNA fragments bearing a phosphate or cyclic phosphate groups at their 3′ end. Unfortunately, all current protocols for library preparation rely only on the presence of a 3′ hydroxyl group. Here, we developed circAID-p-seq, a PCR-free library preparation for 3′ phospho-RNA sequencing. We applied circAID-p-seq to ribosome profiling, which produces fragments protected by ribosomes after endonuclease digestion. CircAID-p-seq, combined with the dedicated computational pipeline circAidMe, facilitates accurate, fast, highly efficient and low-cost sequencing of phospho-RNA fragments from eukaryotic cells and tissues. While assessing circAID-p-seq to portray ribosomes engaged with transcripts, we provide a versatile tool to unravel any 3′-phospho RNA molecules.
2022-03-09 | GSE174754 | GEO