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Elucidating mitotic spindle assembly mechanisms by super-resolution fluorescence microscopy

01.04.2015 – 31.03.2019

Faithful genome transmission in dividing cells relies on correct attachment of all chromosomes to the mitotic spindle. Functional spindles in animal cells require cortical anchorage, maintenance of separate spindle poles, and chromosome attachment to the poles. How microtubules are assembled into such a complex array, wherein distinct microtubule subsets mediate each of these functions, is poorly understood. Here, we

aim to elucidate mechanisms underlying spindle assembly using novel microscopy techniques. A scanned Bessel-beam microscope will be built for ultra-fast 3-D live-cell imaging beyond the diffraction limit. This cutting-edge technology will allow us to study, in unprecedented detail, how microtubules originating from centrosomes, kinetochores and the spindle body are organized into the bundles that attach sister chromatid

to opposing spindle poles. 3-D structured illumination microscopy will be used to visualize spindle morphology beyond the resolution limit of conventional fluorescence microscopes, allowing us to investigate how post-translational tubulin modifications contribute to the functional diversification of spindle microtubules. Overall, this project will yield insights into the mechanisms underlying spindle assembly and faithful chromosome segregation. Furthermore, implementation of a Bessel beam microscope will contribute to develop the Vienna Biocenter Campus into an internationally leading center for bioimaging.

 

Project number

LS17-047

Publications

SiR-Hoechst is a far-red DNA stain for live-cell nanoscopy. Lukinavi?ius G, Blaukopf C, Pershagen E, Schena A, Reymond L, Derivery E, Gonzalez-Gaitan M, D'Este E, Hell SW, Gerlich DW, Johnsson K. Nat Commun. 2015 Oct 1;6:8497. doi: 10.1038/ncomms9497.

Ki-67 acts as a biological surfactant to disperse mitotic chromosomes. Cuylen S, Blaukopf C, Politi AZ, Müller-Reichert T, Neumann B, Poser I, Ellenberg J, Hyman AA, Gerlich DW. Nature. 2016 Jul 14;535(7611):308-12. doi: 10.1038/nature18610.

Profiling DNA damage response following mitotic perturbations. S Pedersen R, Karemore G, Gudjonsson T, Rask MB, Neumann B, Hériché JK, Pepperkok R, Ellenberg J, Gerlich DW, Lukas J, Lukas C. Nat Commun. 2016 Dec 15;7:13887. doi: 10.1038/ncomms13887.

MicroRNA-34/449 controls mitotic spindle orientation during mammalian cortex development. Fededa JP, Esk C, Mierzwa B, Stanyte R, Yuan S, Zheng H, Ebnet K, Yan W, Knoblich JA, Gerlich DW. EMBO J. 2016 Nov 15;35(22):2386-2398. Epub 2016 Oct 5.

Maxim I. Molodtsov, Christine Mieck, Jeroen Dobbelaere, Alexander Dammermann, Stefan Westermann and Alipasha Vaziri A force-induced directional switch of a molecular motor enables parallel microtubule bundle formation Cell 176:2, 539 (2016)

Cytokinesis in vertebrate cells initiates by contraction of an equatorial actomyosin network composed of randomly oriented filaments.

Spira F, Cuylen-Haering S, Mehta S, Samwer M, Reversat A, Verma A, Oldenbourg R, Sixt M, Gerlich DW. Elife. 2017 Nov 6;6. pii: e30867. doi: 10.7554/eLife.30867. 

DNA Cross-Bridging Shapes a Single Nucleus from a Set of Mitotic Chromosomes. Samwer M, Schneider MWG, Hoefler R, Schmalhorst PS, Jude JG, Zuber J, Gerlich DW. Cell. 2017 Aug 24;170(5):956-972.e23. doi: 10.1016/j.cell.2017.07.038.

Dynamic subunit turnover in ESCRT-III assemblies is regulated by Vps4 to mediate membrane remodelling during cytokinesis. Mierzwa BE, Chiaruttini N, Redondo-Morata L, von Filseck JM, König J, Larios J, Poser I, Müller-Reichert T, Scheuring S, Roux A, Gerlich DW. Nat Cell Biol. 2017 Jul;19(7):787-798. doi: 10.1038/ncb3559. Epub 2017 Jun 12.

Vienna Science and Technology Fund (Link)