TRIM Proteine in Autophagie-vermittelter antiviraler Immunabwehr
Virologie
Zusammenfassung der Projektergebnisse
Autophagy and interferon (IFN)-mediated innate immunity are critical antiviral defense mechanisms, and recent evidence highlighted tripartite motif (TRIM) proteins as important regulators of both intricately connected processes. While the role of TRIM proteins in modulating antiviral cytokine responses has been well established, much less is known about their involvement in autophagy in response to different viral pathogens. Using a targeted RNAi screen examining the relevance of selected TRIM proteins in autophagy induced by herpes simplex virus 1 (HSV-1), encephalomyocarditis virus (EMCV), and influenza A virus (IAV), we identified several TRIM proteins that regulated autophagy in a virusspecies specific manner, as well as a few TRIM proteins that were essential for autophagy triggered by all three viruses and rapamycin, among them TRIM23. Surprisingly, TRIM23 was critical for autophagymediated restriction of all viruses tested, which included Respiratory syncytial virus, Adenovirus (Ad) and Sindbis virus (SINV). TRIM23 is unique among the family of TRIM proteins, as it harbors dual emzymatic activity: ubiquitin E3 ligase activity encoded by its RING domain and GTPase activity by its ARF domain. Interestingly, the ARF domain alone is representative of a family of small GTPases. We could establish that autophagy induction activity of TRIM23 was dependent on both its RING E3 ligase and ADP- ribosylation factor (ARF) GTPase activity. Mechanistic studies revealed that highly unconventional K27-linked auto-ubiquitination of the ARF domain is essential for the GTP hydrolysis activity of TRIM23. This GTPase activity drives in turn the activation of TANK-binding kinase 1 (TBK1) by facilitating its dimerization and auto-transphosphorylation. Activated TBK1 induces autophagy then by phosphorylating the selective autophagy receptor p62. Our work identifies the TRIM23-TBK1-p62 axis as a key component of selective virus-induced autophagy and further reveals a novel and unexpected role for K27-linked ubiquitination in GTPase-dependent TBK1 activation. The main goals of the original project were achieved as proposed, as well as the work schedule completed as outlined, resulting in a publication of this project in Nature Microbiology shortly after the end of the fellowship. During my stay in the lab of Prof. Michaela Gack I received confound training in stateof-the-art laboratory techniques, project planning and scientific writing. Furthermore, my first-author project spawned exciting ideas for future research. The intricate connection between autophagy and type-I interferon induction has to be examined focusing on TRIM23, which might connect both pathways by activating the kinase TBK1. TBK1 was found to be critical for both selective autophagy and type-I interferon induction. Furthermore, the upstream activation mechanism of TRIM23 needs to be addressed, to gain fundamental insight on how the autophagic signal is conveyed from viral stimulation down to the induction of autophagic flux and to dissect why TRIM23 selectively works in virus or rapamycin induced autophagy but is dispensable for basal and starvation-induced autophagy. The impact of TRIM23 on immune defenses against viruses in vivo is an intriguing new perspective using the already existing TRIM23 KO mouse model. These mice, however, were never challenged with a virus. Lastly, targeting the two enzymatic functions of TRIM23 may provide the basis for therapeutics against diseases caused by dysregulation of autophagy, such as microbial infections, inflammation, and cancer. Beyond TRIM23, I could show a new possible role of K27-linked ubiquitination in the modification of TRIM23-related small GTPases: ARF proteins. Researching the impact of K27-linked ubiquitin also on the ARF family of proteins will be the subject of future research. In summary, this project was highly successful in terms of reaching its proposed goals, identifying the TRIM protein TRIM23 as being essential for virus-induced autophagy and unravelling its unusual molecular mechanism and further highlighting the role of TBK1 during selective, p62-dependent autophagy. The results of the project were published in Nature Microbiology and will provide the basis for compelling future studies.
Projektbezogene Publikationen (Auswahl)
- Intracellular detection of viral nucleic acids. Curr Opin Microbiol. 2015 Aug;26:1-9
Sparrer KM, Gack MU
(Siehe online unter https://doi.org/10.1016/j.mib.2015.03.001) - TRIM23 mediates virus-induced autophagy via activation of TBK1. Nature Microbiology, 2017 Aug
Sparrer KM, Gableske S, Zurenski M, Parker ZM, Full F, Baumgart GJ, Kato J, Pacheco-Rodriguez G, Liang C, Pornillos O, Moss J, Vaughan M, Gack MU
(Siehe online unter https://doi.org/10.1038/s41564-017-0017-2) - TRIM proteins: New players in virus-induced autophagy. PLoS pathogens. 2018 14 (2), e1006787
Sparrer KM, Gack MU
(Siehe online unter https://doi.org/10.1371/journal.ppat.1006787)
