Distinct and evolutionary conserved structural features of the human nuclear exosome complex
- Max Planck Institute of Biochemistry, Germany
Abstract
The nuclear RNA exosome complex mediates the processing of structured RNAs and the decay of aberrant non-coding RNAs, an important function particularly in human cells. Most mechanistic studies to date have focused on the yeast system. Here, we reconstituted and studied the properties of a recombinant 14-subunit human nuclear exosome complex. In biochemical assays, the human exosome embeds a longer RNA channel than its yeast counterpart. The 3.8 Å resolution cryo-EM structure of the core complex bound to a single-stranded RNA reveals that the RNA channel path is formed by two distinct features of the hDIS3 exoribonuclease: an open conformation and a domain organization more similar to bacterial RNase II than to yeast Rrp44. The cryo-EM structure of the holo-complex shows how obligate nuclear cofactors position the hMTR4 helicase at the entrance of the core complex, suggesting a striking structural conservation from lower to higher eukaryotes.
Data availability
The cryo-EM density maps are deposited in the Electron Microscopy Data Bank under accession numbers EMD-0127 and EMD-0128. The atomic model is deposited in the Protein Data Bank (PDB) under accession number 6H25.
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Human nuclear RNA exosome EXO-14 complex (cryo-EM density map)Publicly available at the Electron Microscopy Data Bank (accession no: EMD-0127).
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Cryo-EM density mapPublicly available at the Electron Microscopy Data Bank (accession no: EMD-0128).
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Human nuclear RNA exosome EXO-10-MPP6 complex (atomic model)Publicly available at the RCSB Protein Data Bank (accession no: 6H25).
Article and author information
Author details
Funding
European Molecular Biology Organization (ALTF 1008-2015)
- Piotr Gerlach
European Commission (ERC-2016-ADG 740329 EXORICO)
- Elena Conti
Deutsche Forschungsgemeinschaft (SFB646)
- Elena Conti
Louis-Jeantet Foundation
- Elena Conti
Max-Planck-Gesellschaft
- Elena Conti
Deutsche Forschungsgemeinschaft (SFB1035)
- Elena Conti
Deutsche Forschungsgemeinschaft (GRK1721)
- Elena Conti
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Copyright
© 2018, Gerlach et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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Distinct and evolutionary conserved structural features of the human nuclear exosome complex
eLife 7:e38686.
https://doi.org/10.7554/eLife.38686
Further reading
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Glutamine synthetases (GS) are central enzymes essential for the nitrogen metabolism across all domains of life. Consequently, they have been extensively studied for more than half a century. Based on the ATP-dependent ammonium assimilation generating glutamine, GS expression and activity are strictly regulated in all organisms. In the methanogenic archaeon Methanosarcina mazei, it has been shown that the metabolite 2-oxoglutarate (2-OG) directly induces the GS activity. Besides, modulation of the activity by interaction with small proteins (GlnK1 and sP26) has been reported. Here, we show that the strong activation of M. mazei GS (GlnA1) by 2-OG is based on the 2-OG dependent dodecamer assembly of GlnA1 by using mass photometry (MP) and single particle cryo-electron microscopy (cryo-EM) analysis of purified strep-tagged GlnA1. The dodecamer assembly from dimers occurred without any detectable intermediate oligomeric state and was not affected in the presence of GlnK1. The 2.39 Å cryo-EM structure of the dodecameric complex in the presence of 12.5 mM 2-OG demonstrated that 2-OG is binding between two monomers. Thereby, 2-OG appears to induce the dodecameric assembly in a cooperative way. Furthermore, the active site is primed by an allosteric interaction cascade caused by 2-OG-binding towards an adaption of an open active state conformation. In the presence of additional glutamine, strong feedback inhibition of GS activity was observed. Since glutamine dependent disassembly of the dodecamer was excluded by MP, feedback inhibition most likely relies on the binding of glutamine to the catalytic site. Based on our findings, we propose that under nitrogen limitation the induction of M. mazei GS into a catalytically active dodecamer is not affected by GlnK1 and crucially depends on the presence of 2-OG.
