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Legionella pp 267-276 | Cite as

Methods for Noncanonical Ubiquitination and Deubiquitination Catalyzed by Legionella pneumophila Effector Proteins

  • Jiazhang Qiu
  • Zhao-Qing LuoEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1921)

Abstract

Protein ubiquitination is one of the most prevalent posttranslational modifications; it regulates a wide range of critical cellular processes in eukaryotes. This modification occurs by covalent attachment of the ubiquitin molecule to other proteins via an isopeptide bond in reactions typically catalyzed by sequential actions of three enzymes, including ubiquitin-activating enzyme (E1), ubiquitin-conjugating enzyme (E2), and ubiquitin ligase (E3). Ubiquitination is a reversible process catalyzed by a group of proteins known as deubiquitinase (DUB), which specifically cleaves the isopeptide bond between ubiquitin and modified proteins. Recently, a novel form of ubiquitination catalyzed by the SidE family of effectors from the bacterial pathogen Legionella pneumophila was reported. These proteins ubiquitinate structurally diverse host proteins such as reticulons and ER-associated Rab small GTPases by a two-step mechanism that uses NAD as the energy source for ubiquitin activation prior to being transferred to serine residues in target proteins. This process bypasses the need for E1 and E2 enzymes. Intriguingly, ubiquitination induced by SidEs is regulated by SidJ, another L. pneumophila effector protein which reverses the modification by functioning as an unconventional DUB. Here, we summarize the experimental details of Rab small GTPases (use Rab33b as an example) ubiquitination catalyzed by SidEs (use SdeA as an example) as well as deubiquitination catalyzed by SidJ.

Key words

Dot/Icm mART ADP-ribosylation Phosphodiesterase Rab small GTPase 

Notes

Acknowledgments

We thank members of our laboratory for helpful discussion. Our work is supported by NIH-NIAID grants R21AI105714 and R01AI127465. J.Q. is a recipient of the “Young 1000 Talents Program” scholar supported by the Chinese Central Government.

References

  1. 1.
    Hershko A, Ciechanover A (1998) The ubiquitin system. Annu Rev Biochem 67:425–479.  https://doi.org/10.1146/annurev.biochem.67.1.425CrossRefPubMedGoogle Scholar
  2. 2.
    Mevissen TET, Komander D (2017) Mechanisms of deubiquitinase specificity and regulation. Annu Rev Biochem 86:159–192.  https://doi.org/10.1146/annurev-biochem-061516-044916CrossRefPubMedGoogle Scholar
  3. 3.
    Zhou Y, Zhu YQ (2015) Diversity of bacterial manipulation of the host ubiquitin pathways. Cell Microbiol 17(1):26–34CrossRefGoogle Scholar
  4. 4.
    Newton HJ, Ang DK, van Driel IR, Hartland EL (2010) Molecular pathogenesis of infections caused by Legionella pneumophila. Clin Microbiol Rev 23(2):274–298.  https://doi.org/10.1128/CMR.00052-09CrossRefPubMedPubMedCentralGoogle Scholar
  5. 5.
    Kubori T, Nagai H (2016) The Type IVB secretion system: an enigmatic chimera. Curr Opin Microbiol 29:22–29.  https://doi.org/10.1016/j.mib.2015.10.001CrossRefPubMedGoogle Scholar
  6. 6.
    Qiu J, Luo ZQ (2017) Legionella and Coxiella effectors: strength in diversity and activity. Nat Rev Microbiol 15(10):591–605.  https://doi.org/10.1038/nrmicro.2017.67CrossRefPubMedGoogle Scholar
  7. 7.
    Dorer MS, Kirton D, Bader JS, Isberg RR (2006) RNA interference analysis of Legionella in Drosophila cells: exploitation of early secretory apparatus dynamics. PLoS Pathog 2(4):315–327CrossRefGoogle Scholar
  8. 8.
    Qiu J, Luo ZQ (2017) Hijacking of the Host Ubiquitin Network by Legionella pneumophila. Front Cell Infect Microbiol 7:487.  https://doi.org/10.3389/fcimb.2017.00487CrossRefPubMedPubMedCentralGoogle Scholar
  9. 9.
    Qiu J, Sheedlo MJ, Yu K, Tan Y, Nakayasu ES, Das C et al (2016) Ubiquitination independent of E1 and E2 enzymes by bacterial effectors. Nature 533(7601):120–124.  https://doi.org/10.1038/nature17657CrossRefPubMedPubMedCentralGoogle Scholar
  10. 10.
    Bhogaraju S, Kalayil S, Liu Y, Bonn F, Colby T, Matic I et al (2016) Phosphoribosylation of ubiquitin promotes serine ubiquitination and impairs conventional ubiquitination. Cell 167(6):1636–49 e13.  https://doi.org/10.1016/j.cell.2016.11.019CrossRefPubMedGoogle Scholar
  11. 11.
    Kotewicz KM, Ramabhadran V, Sjoblom N, Vogel JP, Haenssler E, Zhang M et al (2016) A single legionella effector catalyzes a multistep ubiquitination pathway to rearrange tubular endoplasmic reticulum for replication. Cell Host Microbe 21(2):169–181.  https://doi.org/10.1016/j.chom.2016.12.007CrossRefPubMedPubMedCentralGoogle Scholar
  12. 12.
    Qiu JZ, Yu KW, Fei XW, Liu Y, Nakayasu ES, Piehowski PD et al (2017) A unique deubiquitinase that deconjugates phosphoribosyl-linked protein ubiquitination. Cell Res 27(7):865–881CrossRefGoogle Scholar
  13. 13.
    Berger KH, Isberg RR (1993) Two distinct defects in intracellular growth complemented by a single genetic locus in Legionella pneumophila. Mol Microbiol 7(1):7–19CrossRefGoogle Scholar
  14. 14.
    Xu L, Shen X, Bryan A, Banga S, Swanson MS, Luo ZQ (2010) Inhibition of host vacuolar H+-ATPase activity by a Legionella pneumophila effector. PLoS Pathog 6(3):e1000822.  https://doi.org/10.1371/journal.ppat.1000822CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Key Laboratory of Zoonosis, Ministry of Education, College of Veterinary MedicineJilin UniversityChangchunChina
  2. 2.Department of Biological Sciences, Purdue Institute for Inflammation, Immunology and Infectious DiseasePurdue UniversityWest LafayetteUSA

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