Early rRNA processing is a stress-dependent regulatory event whose inhibition maintains nucleolar integrity

Szaflarski, Witold; Leśniczak-Staszak, Marta; Sowiński, Mateusz; Ojha, Sandeep; Aulas, Anaïs; Dave, Dhwani; Malla, Sulochan; Anderson, Paul; Иванов, П. А.; Lyons, Shawn M.

doi:10.1093/nar/gkab1231

Cited by 37 publications

(27 citation statements)

References 89 publications

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“…Interestingly, a recent study demonstrated defects in 45S rRNA processing upon sodium arsenite exposure (40). We speculate that the accumulation of rRNA in cytoplasmic granules distinct from stress granules and P-bodies may be related to the elimination of misprocessed forms generated during oxidative stress.…”

Section: Discussionmentioning

confidence: 71%

“…Further, we find that granule formation is sensitive to the RNA Pol I inhibitor ActD but largely insensitive to the RNA Pol II inhibitor α-amanitin, suggesting that granules contain rRNA. Interestingly, a recent study demonstrated defects in 45S rRNA processing upon sodium arsenite exposure(40). We speculate that the accumulation of rRNA in cytoplasmic granules distinct from stress granules and P-bodies may be related to the elimination of misprocessed forms generated during oxidative stress.…”

Section: Discussionmentioning

confidence: 99%

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Live-cell RNA imaging with metabolically incorporated fluorescent nucleosides

Wang

Shalamberidze

Arguello

et al. 2022

Preprint

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Fluorescence imaging is a powerful method for probing macromolecular dynamics in biological systems, however approaches for cellular RNA imaging are limited to the investigation of individual RNA constructs or bulk RNA labeling methods compatible primarily with fixed samples. Here, we develop a platform for fluorescence imaging of bulk RNA dynamics in living cells. We show that fluorescent bicyclic and tricyclic cytidine analogues can be metabolically incorporated into cellular RNA by overexpression of uridine-cytidine kinase 2 (UCK2). In particular, metabolic feeding with the tricyclic cytidine-derived nucleoside tC combined with quantitative confocal imaging enables the investigation of RNA synthesis, degradation, and trafficking at single-cell resolution. We apply our imaging modality to study RNA metabolism and localization during the oxidative stress response and find that bulk RNA turnover is greatly accelerated upon NaAsO2 treatment. Further, we identify cytoplasmic RNA granules containing RNA transcripts generated during oxidative stress that are distinct from canonical stress granules and P-bodies and co-localize with the RNA helicase DDX6. Taken together, our work provides a powerful approach for live-cell RNA imaging and reveals how cells reshape RNA transcriptome dynamics in response to oxidative stress.

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Section: Discussionmentioning

confidence: 71%

Section: Discussionmentioning

confidence: 99%

Live-cell RNA imaging with metabolically incorporated fluorescent nucleosides

Wang

Shalamberidze

Arguello

et al. 2022

Preprint

View full text Add to dashboard Cite

show abstract

“…Interestingly, a recent study demonstrated defects in 45S rRNA processing upon sodium arsenite exposure. 29 We speculate that the accumulation of rRNA in cytoplasmic granules distinct from stress granules and P-bodies may be related to the elimination of misprocessed forms generated during oxidative stress. The accumulation of DDX6 in these structures is intriguing as DDX6 has been implicated in P-body assembly, mRNA decay, and translational repression 26,30 but to the best of our knowledge is not known to interact with rRNA.…”

Section: ■ Discussionmentioning

confidence: 88%

Live-Cell RNA Imaging with Metabolically Incorporated Fluorescent Nucleosides

et al. 2022

View full text Add to dashboard Cite

Fluorescence imaging is a powerful method for probing macromolecular dynamics in biological systems; however, approaches for cellular RNA imaging are limited to the investigation of individual RNA constructs or bulk RNA labeling methods compatible primarily with fixed samples. Here, we develop a platform for fluorescence imaging of bulk RNA dynamics in living cells. We show that fluorescent bicyclic and tricyclic cytidine analogues can be metabolically incorporated into cellular RNA by overexpression of uridine−cytidine kinase 2. In particular, metabolic feeding with the tricyclic cytidine-derived nucleoside tC combined with confocal imaging enables the investigation of RNA synthesis, degradation, and trafficking at single-cell resolution. We apply our imaging modality to study RNA metabolism and localization during the oxidative stress response and find that bulk RNA turnover is greatly accelerated upon NaAsO 2 treatment. Furthermore, we identify cytoplasmic RNA granules containing RNA transcripts generated during oxidative stress that are distinct from canonical stress granules and P-bodies and co-localize with the RNA helicase DDX6. Taken together, our work provides a powerful approach for live-cell RNA imaging and reveals how cells reshape RNA transcriptome dynamics in response to oxidative stress.

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“…The nucleolus possesses numerous functions, including ribosome biogenesis, nuclear organization, regulation of global gene expression, and energy metabolism (Cerqueira and Lemos, 2019). It also responds to multiple stresses, such as hypoxia, pH fluctuations, redox stress, DNA damage, or proteasome inhibition, and acts as a protein quality control center that can mitigate heat shock-induced proteotoxicity (Mekhail et al, 2005; Latonen et al, 2011; Audas et al, 2012; Yang et al, 2016; Lindström et al, 2018; Alberti and Carra, 2019; Azkanaz et al, 2019; Frottin et al, 2019; Mediani et al, 2019; Szaflarski et al, 2022). In our studies, we focused on the latter function.…”

Section: Discussionmentioning

confidence: 99%

CHIP ubiquitin ligase is involved in the nucleolar stress management

Piechota

Biriczova

Kowalski

et al. 2022

Preprint

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The nucleolus is a dynamic nuclear biomolecular condensate involved in cellular stress response. Under proteotoxic stress, the nucleolus can store damaged proteins for refolding or degradation. HSP70 chaperone is a well-documented player in the recovery process of proteins accumulated in the nucleolus after heat shock. However, little is known about the involvement of the ubiquitin-proteasome system in the turnover of its nucleolar clients. Here we show that HSP70, independently of its ATPase activity, promotes migration of the CHIP (carboxyl terminus of HSC70-interacting protein) ubiquitin ligase into the granular component of the nucleolus, specifically after heat stress. We show that while in the nucleolus, CHIP retains mobility that depends on its ubiquitination activity. Furthermore, after prolonged exposure to heat stress, CHIP self-organizes into large, intra-nucleolar droplet-like structures whose size is determined by CHIP ubiquitination capacity. Using a heat-sensitive nucleolar protein luciferase, we show that excess CHIP impairs its regeneration, probably through deregulation of HSP70. Our results demonstrate a novel role for CHIP in managing nucleolar proteostasis in response to stress.

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Early rRNA processing is a stress-dependent regulatory event whose inhibition maintains nucleolar integrity

Cited by 37 publications

References 89 publications

Live-cell RNA imaging with metabolically incorporated fluorescent nucleosides

Live-cell RNA imaging with metabolically incorporated fluorescent nucleosides

Live-Cell RNA Imaging with Metabolically Incorporated Fluorescent Nucleosides

CHIP ubiquitin ligase is involved in the nucleolar stress management

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