Not4 and Not5 modulate translation elongation by Rps7A ubiquitination, Rli1 moonlighting, and condensates that exclude eIF5A

Allen, George E.; Panasenko, Olesya O.; Villányi, Zoltán; Zagatti, Marina; Weiss, Benjamin; Pagliazzo, Lucile; Huch, Susanne; Polte, Christine; Zahorán, Szabolcs; Hughes, Christopher S.; Ignatova, Zoya; Collart, Martine A.

doi:10.1016/j.celrep.2021.109633

Cited by 27 publications

(80 citation statements)

References 77 publications

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“…In a recent study we showed that the dynamics of translation elongation were greatly affected in the absence of Not4 and Not5 in S.cerevisiae and we proposed that this was due to defective tethering of translating ribosomes to ribonucleoprotein (RNP) condensates, in particular ribosomes paused at non-optimal codons (17). This model predicts that ribosomes paused at non-optimal codons should be enriched in the insoluble mRNA condensates.…”

Section: Resultsmentioning

confidence: 99%

“…For this latter category the reads for the total RNA pool were less different between the first and second halves of the CDS than for the first category that showed a pattern more similar to that seen for all mRNAs. (17) showed instead a mild inverse correlation with 5'P-RDO changes upon Not5 depletion (Figure S3A). This suggests that in not5D cells limiting amounts of Not1 have a dominant impact on RDOs.…”

Section: Metagene Profiles Of Decay Intermediates Distinguish Mrnas I...mentioning

confidence: 98%

“…The copyright holder for this preprint this version posted March 14, 2022. ; https://doi.org/10.1101/2022.03.14.484207 doi: bioRxiv preprint charged tRNAs, codon bias, the amino acid composition of the nascent chain, co-translational folding, interactions of nascent chains with auxiliary factors, and by mRNA localization or mRNA partitioning into membrane-less granules (8)(9)(10)(11)(12)(13)(14). mRNA-protein condensates were first associated with translational repression (stress granules) and mRNA decay (p-bodies) (15), but recent evidence indicates active translation in stress granules (16), and positive roles of granules for translation have been proposed (12,(17)(18)(19). The development of techniques such as ribosome profiling (Ribo-Seq) (20), visualizing with codon-specific precision the position of ribosomes on mRNAs genome-wide, or the sequencing of 5'P decay intermediates (5'P-Seq) (21), revealing patterns of co-translational decay intermediates, has enabled the analysis of translation elongation dynamics with unprecedented depth and precision.…”

Section: Introductionmentioning

confidence: 99%

“…The copyright holder for this preprint this version posted March 14, 2022. ; https://doi.org/10.1101/2022.03.14.484207 doi: bioRxiv preprint exclude the translation initiation and elongation factor eIF5A (51,52). A central role of Not4 and Not5 in translation elongation dynamics is corroborated by the fact that, when deleted in cells, newly synthesized proteins massively aggregate (42,53) accompanied with a high level of abortive translation products (17).…”

Section: Introductionmentioning

confidence: 99%

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Not1 and Not4 inversely determine mRNA solubility that sets the dynamics of co-translational events

Allen

Weiss

Panasenko

et al. 2022

Preprint

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Background: The Ccr4-Not complex is most well known as the major eukaryotic deadenylase. However, several studies have uncovered roles of the complex, in particular of the Not subunits, unrelated to deadenylation and relevant for translation. In particular, the existence of Not condensates that regulate translation elongation dynamics have been reported. Typical studies that evaluate translation efficiency rely on soluble extracts obtained after disruption of cells and ribosome profiling. Yet cellular mRNAs in condensates can be actively translated and may not be present in such extracts. In this work, by analyzing soluble and insoluble mRNA decay intermediates in yeast, we determine that insoluble mRNAs are enriched for ribosomes dwelling at non-optimal codons compared to soluble mRNAs. mRNA decay is higher for soluble RNAs, but the proportion of co-translational degradation relative to the overall mRNA decay is higher for insoluble mRNAs. We show that depletion of Not1 and Not4 inversely impact mRNA solubilities and, for soluble mRNAs, ribosome dwelling according to codon optimality. Depletion of Not4 solubilizes mRNAs with lower non-optimal codon content and higher expression that are rendered insoluble by Not1 depletion. By contrast, depletion of Not1 solubilizes mitochondrial mRNAs, which are rendered insoluble upon Not4 depletion. Our results reveal that mRNA solubility defines dynamics of co-translation events and is oppositely regulated by Not1 and Not4, a mechanism that we additionally determine may already be set by Not1 promoter association in the nucleus.

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Section: Resultsmentioning

confidence: 99%

Section: Metagene Profiles Of Decay Intermediates Distinguish Mrnas I...mentioning

confidence: 98%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Not1 and Not4 inversely determine mRNA solubility that sets the dynamics of co-translational events

Allen

Weiss

Panasenko

et al. 2022

Preprint

Self Cite

View full text Add to dashboard Cite

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“…An important target for general translation coordination is eS7, that in yeast is monoubiquitinated by the E3 ligase Not4 1 2 9 , a component of the Ccr4-Not complex and master regulator of gene expression 26 . eS7 ubiquitination by Not4 occurs constitutively after canonical translation initiation and it was demonstrated to be required for mRNA homeostasis by serving as a prerequisite of Ccr4-Not's role in degradation of non-optimal mRNAs 1,27 . Monoubiquitination of eS7 is also responsible for translational regulation under ER stress condition 28 .…”

Section: Introductionmentioning

confidence: 99%

Recognition and deubiquitination of free 40S for translational reset by Otu2

Ikeuchi

Ivić

Cheng

et al. 2021

Preprint

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In actively translating 80S ribosomes the ribosomal protein eS7 of the 40S subunit is monoubiquitinated by the E3 ligase Not41,2 and deubiquitinated by the deubiquitination enzyme Otu2 upon ribosomal subunit recycling3. Despite its importance for general efficiency of translation the exact role and structural basis for this specific translational reset are only poorly understood. Here we present biochemical and structural data showing that Otu2 can engage the recycled 40S subunit together with the recycling factors ABCE1 and Tma64 immediately after 60S dissociation for mRNA recycling, and that it dissociates before 48S initiation complex formation. A combined structural analysis of Otu2 and Otu2-40S complexes by X-ray crystallography, AlphaFold2 prediction4 and cryo-EM revealed how Otu2 can specifically be recruited to the 40S, but not to the 80S ribosome, for removal of the eS7-bound ubiquitin moiety. Here, interactions of the largely helical N-terminal domain of Otu2 to sites that are masked and therefore inaccessible in the 80S ribosome are of crucial importance. Collectively, we provide the structural basis for the Otu2 driven deubiquitination step providing a first mechanistic understanding of this translational reset step during ribosome recycling/(re)initiation.

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Rebirth of the translational machinery: The importance of recycling ribosomes

Young

Guydosh

2022

BioEssays

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Translation of the genetic code occurs in a cycle where ribosomes engage mRNAs, synthesize protein, and then disengage in order to repeat the process again. The final part of this process-ribosome recycling, where ribosomes dissociate from mRNAsinvolves a complex molecular choreography of specific protein factors to remove the large and small subunits of the ribosome in a coordinated fashion. Errors in this process can lead to the accumulation of ribosomes at stop codons or translation of downstream open reading frames (ORFs). Ribosome recycling is also critical when a ribosome stalls during the elongation phase of translation and must be rescued to allow continued translation of the mRNA. Here we discuss the molecular interactions that drive ribosome recycling, and their regulation in the cell. We also examine the consequences of inefficient recycling with regards to disease, and its functional roles in synthesis of novel peptides, regulation of gene expression, and control of mRNA-associated proteins. Alterations in ribosome recycling efficiency have the potential to impact many cellular functions but additional work is needed to understand how this regulatory power is utilized.

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Not4 and Not5 modulate translation elongation by Rps7A ubiquitination, Rli1 moonlighting, and condensates that exclude eIF5A

Cited by 27 publications

References 77 publications

Not1 and Not4 inversely determine mRNA solubility that sets the dynamics of co-translational events

Not1 and Not4 inversely determine mRNA solubility that sets the dynamics of co-translational events

Recognition and deubiquitination of free 40S for translational reset by Otu2

Rebirth of the translational machinery: The importance of recycling ribosomes

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