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Turnbull, Kathryn Jane
Publications (2 of 2) Show all publications
Kasari, V., Pochopien, A. A., Margus, T., Murina, V., Turnbull, K. J., Zhou, Y., . . . Hauryliuk, V. (2019). A role for the Saccharomyces cerevisiae ABCF protein New1 in translation termination/recycling. Nucleic Acids Research, 47(16), 8807-8820
Open this publication in new window or tab >>A role for the Saccharomyces cerevisiae ABCF protein New1 in translation termination/recycling
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2019 (English)In: Nucleic Acids Research, ISSN 0305-1048, E-ISSN 1362-4962, Vol. 47, no 16, p. 8807-8820Article in journal (Refereed) Published
Abstract [en]

Translation is controlled by numerous accessory proteins and translation factors. In the yeast Saccharomyces cerevisiae, translation elongation requires an essential elongation factor, the ABCF ATPase eEF3. A closely related protein, New1, is encoded by a non-essential gene with cold sensitivity and ribosome assembly defect knock-out phenotypes. Since the exact molecular function of New1 is unknown, it is unclear if the ribosome assembly defect is direct, i.e. New1 is a bona fide assembly factor, or indirect, for instance due to a defect in protein synthesis. To investigate this, we employed yeast genetics, cryo-electron microscopy (cryo-EM) and ribosome profiling (Ribo-Seq) to interrogate the molecular function of New1. Overexpression of New1 rescues the inviability of a yeast strain lacking the otherwise strictly essential translation factor eEF3. The structure of the ATPase-deficient (EQ2) New1 mutant locked on the 80S ribosome reveals that New1 binds analogously to the ribosome as eEF3. Finally, Ribo-Seq analysis revealed that loss of New1 leads to ribosome queuing upstream of 3′-terminal lysine and arginine codons, including those genes encoding proteins of the cytoplasmic translational machinery. Our results suggest that New1 is a translation factor that fine-tunes the efficiency of translation termination or ribosome recycling.

Place, publisher, year, edition, pages
Oxford University Press, 2019
National Category
Biochemistry and Molecular Biology
Identifiers
urn:nbn:se:umu:diva-164896 (URN)10.1093/nar/gkz600 (DOI)000490576900040 ()31299085 (PubMedID)
Funder
Swedish Research Council, 2017-03783Swedish Research Council, 201504746Swedish Research Council, 2017-04663Ragnar Söderbergs stiftelseThe Kempe Foundations, JCK1627The Kempe Foundations, SMK-1349Magnus Bergvall Foundation, 2017-02098Åke Wiberg Foundation, M14-0207EU, Horizon 2020, 2643Swedish Research Council, 2017-03783
Available from: 2019-11-05 Created: 2019-11-05 Last updated: 2019-12-09Bibliographically approved
Turnbull, K. J., Dzhygyr, I., Lindemose, S., Hauryliuk, V. & Roghanian, M. (2019). Intramolecular Interactions Dominate the Autoregulation of Escherichia coli Stringent Factor RelA. Frontiers in Microbiology, 10, Article ID 1966.
Open this publication in new window or tab >>Intramolecular Interactions Dominate the Autoregulation of Escherichia coli Stringent Factor RelA
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2019 (English)In: Frontiers in Microbiology, ISSN 1664-302X, E-ISSN 1664-302X, Vol. 10, article id 1966Article in journal (Refereed) Published
Abstract [en]

Amino acid starvation in Escherichia coli activates the enzymatic activity of the stringent factor RelA, leading to accumulation of the alarmone nucleotide (p)ppGpp. The alarmone acts as an intercellular messenger to regulate transcription, translation and metabolism to mediate bacterial stress adaptation. The enzymatic activity of RelA is subject to multi-layered allosteric control executed both by ligands - such as "starved" ribosomal complexes, deacylated tRNA and pppGpp - and by individual RelA domains. The auto-regulation of RelA is proposed to act either in cis (inhibition of the enzymatic activity of the N-terminal region, NTD, by regulatory C-terminal region, CTD) or in trans (CTD-mediated dimerization leading to enzyme inhibition). In this report, we probed the regulatory roles of the individual domains of E. coli RelA and our results are not indicative of RelA dimerization being the key regulatory mechanism. First, at growth-permitting levels, ectopic expression of RelA CTD does not interfere with activation of native ReIA, indicating lack of regulation via inhibitory complex formation in the cell. Second, in our biochemical assays, increasing RelA concentration does not decrease the enzyme activity, as would be expected in the case of efficient auto-inhibition via dimerization. Third, while high-level CTD expression efficiently inhibits the growth, the effect is independent of native RelA and is mediated by direct inhibition of protein synthesis, likely via direct interaction with the ribosomal A-site. Finally, deletion of the RRM domain of the CTD region leads to growth inhibition mediated by accumulation of (p)ppGpp, suggesting de-regulation of the synthetic activity in this mutant.

Place, publisher, year, edition, pages
Frontiers Media S.A., 2019
Keywords
(p)ppGpp, RelA, RSH, stringent response, ribosome, translation
National Category
Biochemistry and Molecular Biology Medical Biotechnology
Identifiers
urn:nbn:se:umu:diva-163667 (URN)10.3389/fmicb.2019.01966 (DOI)000482805400001 ()
Available from: 2019-10-21 Created: 2019-10-21 Last updated: 2019-10-21Bibliographically approved
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