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Journal Article

Quantitative single-protein imaging reveals molecular complex formation of integrin, talin, and kindlin during cell adhesion

MPS-Authors
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Fischer,  Lisa
Grashoff, Carsten / Molecular Mechanotransduction, Max Planck Institute of Biochemistry, Max Planck Society;

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Klingner,  Christoph
Grashoff, Carsten / Molecular Mechanotransduction, Max Planck Institute of Biochemistry, Max Planck Society;

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Schlichthärle,  Thomas
Jungmann, Ralf / Molecular Imaging and Bionanotechnology, Max Planck Institute of Biochemistry, Max Planck Society;

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Strauss,  Maximilian T.
Jungmann, Ralf / Molecular Imaging and Bionanotechnology, Max Planck Institute of Biochemistry, Max Planck Society;

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Böttcher,  Ralph T.
Fässler, Reinhard / Molecular Medicine, Max Planck Institute of Biochemistry, Max Planck Society;

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Fässler,  Reinhard
Fässler, Reinhard / Molecular Medicine, Max Planck Institute of Biochemistry, Max Planck Society;

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Jungmann,  Ralf
Jungmann, Ralf / Molecular Imaging and Bionanotechnology, Max Planck Institute of Biochemistry, Max Planck Society;

/persons/resource/persons78025

Grashoff,  Carsten
Grashoff, Carsten / Molecular Mechanotransduction, Max Planck Institute of Biochemistry, Max Planck Society;

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Fulltext (public)

s41467-021-21142-2.pdf
(Publisher version), 3MB

Supplementary Material (public)

41467_2021_21142_MOESM1_ESM.pdf
(Supplementary material), 3MB

41467_2021_21142_MOESM2_ESM.pdf
(Supplementary material), 5MB

41467_2021_21142_MOESM3_ESM.pdf
(Supplementary material), 317KB

Citation

Fischer, L., Klingner, C., Schlichthärle, T., Strauss, M. T., Böttcher, R. T., Fässler, R., et al. (2021). Quantitative single-protein imaging reveals molecular complex formation of integrin, talin, and kindlin during cell adhesion. Nature Communications, 12(1): 919. doi:10.1038/s41467-021-21142-2.


Cite as: http://hdl.handle.net/21.11116/0000-0008-2B9D-8
Abstract
Single-molecule localization microscopy (SMLM) enabling the investigation of individual proteins on molecular scales has revolutionized how biological processes are analysed in cells. However, a major limitation of imaging techniques reaching single-protein resolution is the incomplete and often unknown labeling and detection efficiency of the utilized molecular probes. As a result, fundamental processes such as complex formation of distinct molecular species cannot be reliably quantified. Here, we establish a super-resolution microscopy framework, called quantitative single-molecule colocalization analysis (qSMCL), which permits the identification of absolute molecular quantities and thus the investigation of molecular-scale processes inside cells. The method combines multiplexed single-protein resolution imaging, automated cluster detection, in silico data simulation procedures, and widely applicable experimental controls to determine absolute fractions and spatial coordinates of interacting species on a true molecular level, even in highly crowded subcellular structures. The first application of this framework allowed the identification of a long-sought ternary adhesion complex-consisting of talin, kindlin and active beta 1-integrin-that specifically forms in cell-matrix adhesion sites. Together, the experiments demonstrate that qSMCL allows an absolute quantification of multiplexed SMLM data and thus should be useful for investigating molecular mechanisms underlying numerous processes in cells. Single-molecule localisation microscopy is limited by low labeling and detection efficiencies of the molecular probes. Here the authors report a framework to obtain absolute molecular quantities on a true molecular scale; the data reveal a ternary adhesion complex underlying cell-matrix adhesion.