bims-tricox 2022-08-07 papers

Acta Naturae. 2022 Apr-Jun;14(2):14(2): 39-49

Eukaryotic Ribosome Biogenesis: The 60S Subunit.

A A Moraleva, A S Deryabin, Yu P Rubtsov, M P Rubtsova, O A Dontsova.

Ribosome biogenesis is consecutive coordinated maturation of ribosomal precursors in the nucleolus, nucleoplasm, and cytoplasm. The formation of mature ribosomal subunits involves hundreds of ribosomal biogenesis factors that ensure ribosomal RNA processing, tertiary structure, and interaction with ribosomal proteins. Although the main features and stages of ribosome biogenesis are conservative among different groups of eukaryotes, this process in human cells has become more complicated due to the larger size of the ribosomes and pre-ribosomes and intricate regulatory pathways affecting their assembly and function. Many of the factors involved in the biogenesis of human ribosomes have been identified using genome-wide screening based on RNA interference. A previous part of this review summarized recent data on the processing of the primary rRNA transcript and compared the maturation of the small 40S subunit in yeast and human cells. This part of the review focuses on the biogenesis of the large 60S subunit of eukaryotic ribosomes.

Keywords: biogenesis; nucleolus; ribosome; ribosomopathy

DOI: https://doi.org/10.32607/actanaturae.11541

Nat Chem. 2022 Aug 04.

The ribosome stabilizes partially folded intermediates of a nascent multi-domain protein.

Sammy H S Chan, Tomasz Włodarski, Julian O Streit, Anaïs M E Cassaignau, Lauren F Woodburn, Minkoo Ahn, Georg Johannes Freiherr von Sass, Christopher A Waudby, Nediljko Budisa, Lisa D Cabrita, John Christodoulou.

Co-translational folding is crucial to ensure the production of biologically active proteins. The ribosome can alter the folding pathways of nascent polypeptide chains, yet a structural understanding remains largely inaccessible experimentally. We have developed site-specific labelling of nascent chains to detect and measure, using 19F nuclear magnetic resonance (NMR) spectroscopy, multiple states accessed by an immunoglobulin-like domain within a tandem repeat protein during biosynthesis. By examining ribosomes arrested at different stages during translation of this common structural motif, we observe highly broadened NMR resonances attributable to two previously unidentified intermediates, which are stably populated across a wide folding transition. Using molecular dynamics simulations and corroborated by cryo-electron microscopy, we obtain models of these partially folded states, enabling experimental verification of a ribosome-binding site that contributes to their high stabilities. We thus demonstrate a mechanism by which the ribosome could thermodynamically regulate folding and other co-translational processes.

DOI: https://doi.org/10.1038/s41557-022-01004-0

Semin Cell Dev Biol. 2022 Jul 28. pii: S1084-9521(22)00237-3. [Epub ahead of print]

Ribosome homeostasis.

Michael Buszczak.

DOI: https://doi.org/10.1016/j.semcdb.2022.07.008

Methods Mol Biol. 2022 ;2516 305-316

Toeprint Assays for Detecting RNA Structure and Protein-RNA Interactions.

Helen Yakhnin, Paul Babitzke.

Toeprint assays are primer extension inhibition assays that can detect the 3' end of an RNA secondary structure, the position of a bound RNA binding protein, as well as the position of a bound 30S ribosomal subunit or a stalled ribosome. Here we describe how this assay was used to identify an RNA hairpin that sequesters a Shine-Dalgarno sequence, how the RNA-binding protein CsrA can alter RNA structure and affect 30S ribosomal subunit binding, and how the macrolide antibiotic tylosin can induce ribosome stalling.

Keywords: Primer extension inhibition; Protein–RNA interaction; RNA structure; Ribosome; Toeprint; Translational control

DOI: https://doi.org/10.1007/978-1-0716-2413-5_16

Microbiology (Reading). 2022 Aug;168(8):

Era, a GTPase-like protein of the Ras family, does not control ribosome assembly in Mycobacterium tuberculosis.

Nisheeth Agarwal, Shivani Sharma, Pramila Pal, Prem S Kaushal, Naresh Kumar.

Era GTPase is universally present in microbes including Mycobacterium tuberculosis (Mtb) complex bacteria. While Era is known to regulate ribosomal assembly in Escherichia coli and predicted to be essential for in vitro growth, its function in mycobacteria remains obscured. Herein, we show that Era ortholog in the attenuated Mtb H37Ra strain, MRA_2388 (annotated as EraMT) is a cell envelope localized protein harbouring critical GTP-binding domains, which interacts with several envelope proteins of Mtb. The purified Era from M. smegmatis (annotated as EraMS) exhibiting ~90 % sequence similarity with EraMT, exists in monomeric conformation. While it is co-purified with RNA upon overexpression in E. coli, the presence of RNA does not modulate the GTPase activity of the EraMS as against its counterpart from other organisms. CRISPRi silencing of eraMT does not show any substantial effect on the in vitro growth of Mtb H37Ra, which suggests a redundant function of Era in mycobacteria. Notably, no effect on ribosome assembly, protein synthesis or bacterial susceptibility to protein synthesis inhibitors was observed upon depletion of EraMT in Mtb H37Ra, further indicating a divergent role of Era GTPase in mycobacteria.

Keywords: Era; GTPase; Mycobacterium tuberculosis; ribosome assembly

DOI: https://doi.org/10.1099/mic.0.001200

Ageing Res Rev. 2022 Jul 28. pii: S1568-1637(22)00142-8. [Epub ahead of print]81 101700

The epitranscriptome in ageing and stress resistance: A systematic review.

Anja Wagner, Markus Schosserer.

Modifications of RNA, collectively called the "epitranscriptome", might provide novel biomarkers and innovative targets for interventions in geroscience but are just beginning to be studied in the context of ageing and stress resistance. RNA modifications modulate gene expression by affecting translation initiation and speed, miRNA binding, RNA stability, and RNA degradation. Nonetheless, the precise underlying molecular mechanisms and physiological consequences of most alterations of the epitranscriptome are still only poorly understood. We here systematically review different types of modifications of rRNA, tRNA and mRNA, the methodology to analyze them, current challenges in the field, and human disease associations. Furthermore, we compiled evidence for a connection between individual enzymes, which install RNA modifications, and lifespan in yeast, worm and fly. We also included resistance to different stressors and competitive fitness as search criteria for genes potentially relevant to ageing. Promising candidates identified by this approach include RCM1/NSUN5, RRP8, and F33A8.4/ZCCHC4 that introduce base methylations in rRNA, the methyltransferases DNMT2 and TRM9/ALKBH8, as well as factors involved in the thiolation or A to I editing in tRNA, and finally the m6A machinery for mRNA.

Keywords: Ageing; Caenorhabditis elegans; Drosophila melanogaster; Epitranscriptome; Geroscience; Lifespan; RNA modification; Saccharomyces cerevisiae

DOI: https://doi.org/10.1016/j.arr.2022.101700