Molekulare Mechanismen der Podosomen-Dynamik und Makrophagen-Invasion
Zusammenfassung der Projektergebnisse
In this project, we investigated molecular mechanisms regulating the formation, turnover and localized matrix degradation of macrophage podosomes, which are adhesion and invasion structures of monocytic cells. We found that podosomes are crucially regulated by both microtubule-dependent trafficking and also by actomyosin-dependent contractility. In particular, we found that the kinesin KIF9 is important for matrix degradation at podosomes, by interacting with the vesicle regulator flotillin-2, which enables the trafficking of respective vesicles along microtubules to podosomes. We further found that modification of the microtubules themselves, through acetylation by the acetyltransferase MEC-17/ATAT1 is an important regulatory switch that can be used by cells to modify the trafficking of vesicles that are associated with another kinesin, KIF1C. Enhanced microtubule acetylation led to reduced speed, run length and directionality of KIF1C vesicles and also to reduced matrix degradation at podosomes. Investigating one of the main vesicular cargos transported to podosomes, the metalloproteinase MT1-MMP, we found that, in addition to locally degrading matrix material, MT1-MMP also fulfills an unexpected structural role at podosomes. A motif in the cytoplasmic C-terminus of MT1-MMP leads to anchoring of surface-associated MT1-MMP “islets”, even after dissolation of the podosome structure itself, and also acts as an initiation point for the efficient reformation of podosomes. This showed for the first time that MT1-MMP can fulfill a structural function independently of its proteolytic capacity, thus acting as a “memory device” in macrophages. A major new finding concerned the identification of a novel structure on top of the podosome core, the cap, and its role as a regulatory module for actomyosin contractility. We identified lymphocyte-specific protein 1 (LSP1),as an important component of the cap that enables oscillatory contractility of podosomes that is required for mechanosensing. We could also show that LSP1 is crucial in macrophages to perform an actomyosin symmetry break, i.e. a cellular gradient in contractility, that allows polarized migration. LSP1 and the myosin hyperactivator supervillin are localized to different parts of the macrophage cell, as LSP1 has preferential binding ability for β-actin over α-actin, thus localizing to the β-actin-rich periphery and blocking access of supervillin to this region. These data also gave a detailed molecular explanation for the known LSP1-based defects in immune cell migration in diseases such as rheumatoid arthritis or neutrophil actin dysfunction (NAD), opening potentially new treatment options. A respective press release was picked up by the website of the city of Hamburg: http://www.hamburg.de/bwfg/10402246/immunzellen-lassen-ihre-muskeln-spielen/ A still ongoing line of investigation concerns another novel podosome component, Swiprosin-1, which, like LSP1, localizes to the podosome cap, and is crucial for the regular turnover of podosomes.
Projektbezogene Publikationen (Auswahl)
- (2007). The matrix corroded: podosomes and invadopodia in extracellular matrix degradation. Trends Cell Biol., 17(3): 107-117
Linder, S.
(Siehe online unter https://doi.org/10.1016/j.tcb.2007.01.002) - (2008). Assembly and biological roles of podosomes and invadopodia. Curr. Opin. Cell Biol. 20(2): 235-241
Gimona, M., Buccione, R., Courtneidge, S., Linder, S.
(Siehe online unter https://doi.org/10.1016/j.ceb.2008.01.005) - (2011). Degrading devices: invadosomes in proteolytic cell invasion. Ann. Rev. Mol. Cell Biol. 27: 185-211
Linder, S., Wiesner, C., Himmel, M.
(Siehe online unter https://doi.org/10.1146/annurev-cellbio-092910-154216) - (2011). The kinesin KIF9 and reggie/flotillin proteins regulate extracellular matrix degradation by macrophage podosomes, Mol. Biol. Cell 22, 202-215
Cornfine, S., Himmel, M., Kopp, P., El Azzouzi, K., Wiesner, C., Krüger, M., Rudel, T., Linder, S.
(Siehe online unter https://doi.org/10.1091/mbc.e10-05-0394) - (2012). Proteomic analysis of podosome fractions from macrophages reveals similarities to spreading initiation centres. Eur. J. Cell Biol. 91(11-12), 908-922
Cervero, P., Himmel, M., Krüger, M., Linder, S.
(Siehe online unter https://doi.org/10.1016/j.ejcb.2012.05.005) - (2012). Supervillin couples myosin-dependent contractility to podosomes and enables their turnover, J. Cell Sci. 125, 2300-2314
Bhuwania, R., Cornfine, S., Fang, Z., Krüger, M., Luna, E.J., Linder, S.
(Siehe online unter https://doi.org/10.1242/jcs.100032) - (2013). Podosome reformation in macrophages: assays and analysis. Methods Mol. Biol., Vol 1046 (ed. A. Coutts), 97-121
Cervero, P., Panzer, L., Linder, S.
(Siehe online unter https://doi.org/10.1007/978-1-62703-538-5_6) - (2014). Microtubule acetylation regulates dynamics of KIF1C-powered vesicles and contact of microtubule plus ends with podosomes, Eur. J. Cell Biol. 93, 424-437
Bhuwania, R., Castro-Castro, A., Linder, S.
(Siehe online unter https://doi.org/10.1016/j.ejcb.2014.07.006) - (2014). Podosomes in space: macrophage migration and matrix degradation in 2D and 3D settings. Cell Adh. Migr. 8(3), 179-191
Wiesner, C., Le Cabec, V., El Azzouzi, K., Maridonneau-Parini, I., Linder, S.
(Siehe online unter https://doi.org/10.4161/cam.28116) - (2015). Tools of the trade: podosomes as multipurpose organelles of monocytic cells. Cell. Mol. Life Sci.72(1), 121-135
Linder, S., Wiesner, C.
(Siehe online unter https://doi.org/10.1007/s00018-014-1731-z) - (2016). Metalloproteinase MT1-MMP islets act as memory devices for podosome reemergence, J. Cell Biol. 213, 109-125
El Azzouzi, K., Wiesner, C., Linder, S.
(Siehe online unter https://doi.org/10.1083/jcb.201510043) - (2018). Lymphocyte specific protein-1 regulates mechanosensory oscillation of podosomes and actin isoformbased actomyosin symmetry breaking. Nature Comm. 9, 515
Cervero, P., Wiesner, C., Bouissou, A., Poincloux, R., Linder, S.
(Siehe online unter https://doi.org/10.1038/s41467-018-02904-x)
