Functionally distinct roles for eEF2K in the control of ribosome availability and p-body abundance

Smith, Patrick; Loerch, Sarah; Kunder, Nikesh; Stanowick, Alexander; Lou, Tzu-Fang; Campbell, Zachary T.

doi:10.1038/s41467-021-27160-4

Cited by 26 publications

(26 citation statements)

References 109 publications

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“…2B). The rotated ‘eEF2’ state without tRNA fitted the previous structural model very well (15, 17) (fig. S4G), although the eEF2 position was shifted by ∼1.7 nm as compared to the ‘eEF2, ap/P, E’ state (fig.…”

Section: Main Textsupporting

confidence: 63%

Translation dynamics in human cells visualized at high-resolution reveal cancer drug action

Xing

Taniguchi

Khusainov

et al. 2023

Preprint

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Ribosomes catalyze protein synthesis by cycling through various functional states. These states have been extensively characterized in vitro, yet their distribution in actively translating human cells remains elusive. Here, we optimized a cryo-electron tomography-based approach and resolved ribosome structures inside human cells with a local resolution of up to 2.5 angstroms. These structures revealed the distribution of functional states of the elongation cycle, a Z tRNA binding site and the dynamics of ribosome expansion segments. In addition, we visualized structures of Homoharringtonine, a drug for chronic myeloid leukemia treatment, within the active site of the ribosome and found that its binding reshaped the landscape of translation. Overall, our work demonstrates that structural dynamics and drug effects can be assessed at near-atomic detail within human cells.One-Sentence SummarySnapshots of ribosome dynamics at near-atomic resolution within native and drug-treated human cells are revealed.

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Section: Main Textsupporting

confidence: 63%

Translation dynamics in human cells visualized at high-resolution reveal cancer drug action

Xing

Taniguchi

Khusainov

et al. 2023

Preprint

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“…SpotitPy successfully identified the significant decrease in the number of P-bodies per cell when BMDMs were treated with rapamycin, a natural anti-fungal antibiotic known to inhibit mRNA translation and P-body formation. This decrease is also consistent with the existing literature [ 39 , 40 ] (Fig. 4 A–C).…”

Section: Resultssupporting

confidence: 93%

SpotitPy: a semi-automated tool for object-based co-localization of fluorescent labels in microscopy images

Akalestou-Clocher

Kalamara²,

Garinis³

2022

BMC Bioinformatics

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Background In fluorescence microscopy, co-localization refers to the spatial overlap between different fluorescent labels in cells. The degree of overlap between two or more channels in a microscope may reveal a physical interaction or topological functional interconnection between molecules. Recent advances in the imaging field require the development of specialized computational analysis software for the unbiased assessment of fluorescently labelled microscopy images. Results Here we present SpotitPy, a semi-automated image analysis tool for 2D object-based co-localization. SpotitPy allows the user to select fluorescent labels and perform a semi-automated and robust segmentation of the region of interest in distinct cell types. The workflow integrates advanced pre-processing manipulations for de-noising and in-depth semi-automated quantification of the co-localized fluorescent labels in two different channels. We validated SpotitPy by quantitatively assessing the presence of cytoplasmic ribonucleoprotein granules, e.g. processing (P) bodies, under conditions that challenge mRNA translation, thus highlighting SpotitPy benefits for semi-automatic, accurate analysis of large image datasets in eukaryotic cells. SpotitPy comes in a command line interface or a simple graphical user interphase and can be used as a standalone application. Conclusions Overall, we present a novel and user-friendly tool that performs a semi-automated image analysis for 2D object-based co-localization. SpotitPy can provide reproducible and robust quantifications for large datasets within a limited timeframe. The software is open-source and can be found in the GitHub project repository: (https://github.com/alexiaales/SpotitPy).

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“…6B), though the contributions to protein levels are unclear. It is also possible that, rather than a reduction in absolute ribosome content, translation slows due to an accumulation of inactive 80S monosomes, which are stable but not bound to mRNA, and have been shown to arise in several conditions [57][58][59] . However, while the decrease in translating ribosomes is a defining difference between cycling and senescent cells, quiescent cells also reduce their translation capacity and have a similar aim to survive under sub-optimal conditions indefinitely.…”

Section: Discussionmentioning

confidence: 99%

Senescence suppresses the integrated stress response and activates a stress-enhanced secretory phenotype

Payea

Dar

Anerillas

et al. 2023

Preprint

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Senescence is a state of indefinite cell cycle arrest associated with aging, cancer, and age-related diseases. Here, using label-based mass spectrometry, ribosome profiling and nanopore direct RNA sequencing, we explore the coordinated interaction of translational and transcriptional programs of human cellular senescence. We find that translational deregulation and a corresponding maladaptive integrated stress response (ISR) is a hallmark of senescence that desensitizes senescent cells to stress. We show that senescent cells maintain high levels of eIF2α phosphorylation, typical of ISR activation, but translationally repress the stress response transcription factor 4 (ATF4) by ineffective bypass of the inhibitory upstream open reading frames. Surprisingly, ATF4 translation remains inhibited even after acute proteotoxic and amino acid starvation stressors, resulting in a highly diminished stress response. Furthermore, absent a response, stress exacerbates the senescence secretory phenotype and inflammatory pathways thus acting as a possible mechanistic link to disease. Our results reveal a novel mechanism that senescent cells exploit to evade an adaptive stress response and remain viable.

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Functionally distinct roles for eEF2K in the control of ribosome availability and p-body abundance

Cited by 26 publications

References 109 publications

Translation dynamics in human cells visualized at high-resolution reveal cancer drug action

Translation dynamics in human cells visualized at high-resolution reveal cancer drug action

SpotitPy: a semi-automated tool for object-based co-localization of fluorescent labels in microscopy images

Senescence suppresses the integrated stress response and activates a stress-enhanced secretory phenotype

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