Legionella pneumophila inhibits immune signalling via MavC-mediated transglutaminase-induced ubiquitination of UBE2N

Abstract

The bacterial pathogen Legionella pneumophila modulates host immunity using effectors translocated by its Dot/Icm transporter to facilitate its intracellular replication. A number of these effectors employ diverse mechanisms to interfere with protein ubiquitination, a post-translational modification essential for immunity. Here, we have found that L. pneumophila induces monoubiquitination of the E2 enzyme UBE2N by its Dot/Icm substrate MavC(Lpg2147). We demonstrate that MavC is a transglutaminase that catalyses covalent linkage of ubiquitin to Lys92 and Lys94 of UBE2N via Gln40. Similar to canonical transglutaminases, MavC possess deamidase activity that targets ubiquitin at Gln40. We identified Cys74 as the catalytic residue for both ubiquitination and deamidation activities. Furthermore, ubiquitination of UBE2N by MavC abolishes its activity in the formation of K63-type polyubiquitin chains, which dampens NF-κB signalling in the initial phase of bacterial infection. Our results reveal an unprecedented mechanism of modulating host immunity by modifying a key ubiquitination enzyme by ubiquitin transglutamination.

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Fig. 1: L. pneumophila induces a molecular weight shift in UBE2N in a process that requires the Dot/Icm effector MavC.
Fig. 2: MavC induces UBE2N ubiquitination independently of the canonical ubiquitination machinery.
Fig. 3: MavC catalyses ubiquitination of UBE2N by transglutamination.
Fig. 4: Characteristics of MavC as transglutaminase and ubiquitin deamidase.
Fig. 5: Effects of MavC on UBE2N activity and on NF-κB activation during L. pneumophila infection.
Fig. 6: Effects of MavC on NF-κB activation in cells overexpressing various relevant proteins.

Data availability

The data that support the conclusions of this study are included in this published article along with its Supplementary Information files, and are also available from the corresponding author upon request.

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Acknowledgements

The authors thank R. Isberg (Tufts School of Medicine) for bacterial strains, F. Shao (National Institute for Biological Sciences, Beijing) for plasmids, and R. Moore and A. Shukla at the Pacific Northwest National Laboratory (PNNL) for technical support. This work was supported by National Institutes of Health grants R01AI127465(ZQL), R21AI117205 (PJH and ZQL) and GM103493. Part of this work was performed in the W. R. Wiley Environmental Molecular Sciences Laboratory (EMSL), a Department of Energy (DOE) office of Biological and Environmental Research (BER) National User Facility located at the PNNL.

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N.G. and Z.-Q.L. conceived the ideas for this work. N.G. planned and performed experiments. N.G., E.S.N., P.J.H. and Z.-Q.L. interpreted the data. E.S.N. performed mass spectrometric analyses. N.G. and Z.-Q.L. wrote the manuscript and all authors provided editorial input.

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Correspondence to Zhao-Qing Luo.

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Supplementary Figures 1–11.

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Supplementary Table 1

Proteins potentially modified by 3×Ub-AA in mammalian cells.

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Gan, N., Nakayasu, E.S., Hollenbeck, P.J. et al. Legionella pneumophila inhibits immune signalling via MavC-mediated transglutaminase-induced ubiquitination of UBE2N. Nat Microbiol 4, 134–143 (2019). https://doi.org/10.1038/s41564-018-0282-8

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