Ein Adaptorproteinnetzwerk, das generelle und regulatorische Proteolyse in Bacillus subtilis kontrolliert
Zusammenfassung der Projektergebnisse
This DFG project was a continuation of our previous project on investigation of the role of general and regulatory proteolysis by AAA+ protease complexes, their mechanisms of substrate selection and their adaptor protein network. Furthermore, we wanted to explore how these AAA+ protein complexes are embedded in the cellular protein homeostasis and stress response network of the Gram-positive model organism Bacillus subtilis. By continuing to investigate the heat shock regulator Spx and its various functions we learned to appreciate that the cellular stress response facilitates the down regulation and specific modulation of translation to lower the load of for the protein quality control system together with the control and upregulation of (redox) chaperones to ensure a functional cellular protein homeostasis and protect the stressed cells. We also investigated the role of the sHsp chaperone YocM and how it is embedded in the cellular protein quality control and stress response system. Importantly, we could also utilize a fusion of this sHsp to mCherry as a reporter for subcellular protein aggregates. By applying this new reporter and together with additional experiments we could identify the protein arginine kinase McsB as the ClpC adaptor protein necessary and sufficient for the removal of protein aggregates. Our in vitro and in vivo experiments established that the McsB, ClpC system can function together with the kinase activator McsA and the YwlE phosphatase and are forming a unique disaggregation and refolding AAA+ chaperone system. Interestingly, the protein arginine phosphorylation and de-phosphorylation appeared to play a significant role in this process, since our results suggested that McsB targets the misfolded substrates to ClpC unfolding, and that these unfolded substrates are unable to properly refold, due to their concurrent modification by arginine phosphorylation. However, the specific dephosphorylation by YwlE allowed to facilitate the inherent ability of these unfolded protein species to successfully refold. This new mechanism to support refolding seems to also interfere with the dynamic association of ClpC to ClpP, allowing the protection of refolded substrate protein from degradation by ClpP. The central role of the ClpCP and McsB system in protein homeostasis and stress response was confirmed by the identification of ClpC as a target for antibiotics and supported by the characterization of toxic ClpC variants, whose toxicity depended in B. subtilis on the presence of McsB. Interestingly a phage adaptor-like protein for ClpC was identified and characterized, which allowed the phage to repurpose the ClpCP system for the takeover of the infected B. subtilis cells.
Projektbezogene Publikationen (Auswahl)
- (2016) Role of Hsp100/Clp protease complexes in controlling the regulation of motility in Bacillus subtilis. Front. Microbiol. 7:315
Molière, N., Hoßmann, J., Schäfer, H. & Turgay, K.
(Siehe online unter https://doi.org/10.3389/fmicb.2016.00315) - (2017) Functional Diversity of AAA+ Protease Complexes in Bacillus subtilis, Front. Mol. Biosci. 4:44
Elsholz AKW, Birk MS, Charpentier E, Turgay K.
(Siehe online unter https://doi.org/10.3389/fmolb.2017.00044) - (2017) Regulatory coiled-coil domains promote head-tohead assemblies of AAA+ chaperones essential for tunable activity control. eLife 6:e30120
Carroni, M., Franke, K.B., Maurer, M., Jäger, J., Hantke, I., Gloge, F., Linder, D., Gremer, S., Turgay, K., Bukau, B. & Mogk, A.
(Siehe online unter https://doi.org/10.7554/eLife.30120) - (2017) Structure of the Bacillus subtilis hibernating 100S ribosome reveals the basis for 70S dimerization. EMBO J 36:2061-72
Beckert, B., Abdelshahid, M., Schäfer, H., Steinchen, W., Arenz, S., Berninghausen, O., Beckmann, R., Bange, G., Turgay, K. & Wilson, D.N.
(Siehe online unter https://doi.org/10.15252/embj.201696189) - (2018) Structural changes of TasA in biofilm formation of Bacillus subtilis. Proc. Natl. Acad. Sci. USA 115:3237-42
Diehl, A., Roske, Y., Ball, L., Chowdhury, A., Hiller, M., Molière, N., Kramer, R., Stöppler, D., Worth, C. L., Schlegel, B., Leidert, M., Cremer, N., Erdmann, N., Lopez, D., Stephanowitz, H., Krause, E., van Rossum, B.-J., Schmieder, P., Heinemann, U., Turgay, K., Akbey, Ü. & Oschkinat, H.
(Siehe online unter https://doi.org/10.1073/pnas.1718102115) - (2019) Spx, the central regulator of the heat- and oxidative stress response in B. subtilis, can repress transcription of translation-related genes. Mol. Microbiol. 111:514-33
Schäfer, H., Heinz, A., Sudzinová, P., Voß, M., Hantke, I., Krásný, L. & Turgay, K.
(Siehe online unter https://doi.org/10.1111/mmi.14171) - (2019) Xenogeneic modulation of the ClpCP protease of Bacillus subtilis by a phage-encoded adaptor-like protein. J. Biol. Chem. 294:17501–17511
Mulvenna, N., Hantke, I., Burchell, L, Nicod, S., Bell, D., Turgay, K. & Wigneshweraraj, S.
(Siehe online unter https://doi.org/10.1074/jbc.RA119.010007) - (2019) YocM a small heat shock protein can protect Bacillus subtilis cells during salt stress. Mol. Microbiol. 111:423-40
Hantke, I., Schäfer, H., Janczikowski, A., & Turgay, K.
(Siehe online unter https://doi.org/10.1111/mmi.14164)
