Nanoscale organization of the endogenous ASC speck
Ivo M. Glück,
Grusha Primal Mathias,
Sebastian Strauss,
Virgile Rat,
Irene Gialdini,
Thomas Sebastian Ebert,
Che Stafford,
Ganesh Agam,
Suliana Manley,
Veit Hornung,
Ralf Jungmann,
Christian Sieben,
Don C. Lamb
Affiliations
Ivo M. Glück
Department of Chemistry, Ludwig Maximilians-Universität München, Butenandtstraße 5-13, 81377 München, Germany; Center for Nano Science (CENS), Ludwig Maximilians-Universität München, Butenandtstraße 5-13, 81377 München, Germany
Grusha Primal Mathias
Department of Chemistry, Ludwig Maximilians-Universität München, Butenandtstraße 5-13, 81377 München, Germany
Sebastian Strauss
Faculty of Physics and Center for Nanoscience, Ludwig Maximilian University, Munich, Germany; Max Planck Institute of Biochemistry, Martinsried, Germany
Virgile Rat
Department of Chemistry, Ludwig Maximilians-Universität München, Butenandtstraße 5-13, 81377 München, Germany; Center for Nano Science (CENS), Ludwig Maximilians-Universität München, Butenandtstraße 5-13, 81377 München, Germany
Irene Gialdini
Department of Chemistry, Ludwig Maximilians-Universität München, Butenandtstraße 5-13, 81377 München, Germany; Center for Nano Science (CENS), Ludwig Maximilians-Universität München, Butenandtstraße 5-13, 81377 München, Germany
Thomas Sebastian Ebert
Gene Center and Department of Biochemistry, Ludwig-Maximilians-Universität, Munich, Germany
Che Stafford
Gene Center and Department of Biochemistry, Ludwig-Maximilians-Universität, Munich, Germany
Ganesh Agam
Department of Chemistry, Ludwig Maximilians-Universität München, Butenandtstraße 5-13, 81377 München, Germany; Center for Nano Science (CENS), Ludwig Maximilians-Universität München, Butenandtstraße 5-13, 81377 München, Germany
Suliana Manley
Laboratory of Experimental Biophysics, École Polytechnique Fédérale de Lausanne, BSP 427 (Cubotron UNIL), Rte de la Sorge, CH-1015 Lausanne, Switzerland
Veit Hornung
Max Planck Institute of Biochemistry, Martinsried, Germany; Gene Center and Department of Biochemistry, Ludwig-Maximilians-Universität, Munich, Germany
Ralf Jungmann
Faculty of Physics and Center for Nanoscience, Ludwig Maximilian University, Munich, Germany; Max Planck Institute of Biochemistry, Martinsried, Germany
Christian Sieben
Laboratory of Experimental Biophysics, École Polytechnique Fédérale de Lausanne, BSP 427 (Cubotron UNIL), Rte de la Sorge, CH-1015 Lausanne, Switzerland; Corresponding author
Don C. Lamb
Department of Chemistry, Ludwig Maximilians-Universität München, Butenandtstraße 5-13, 81377 München, Germany; Center for Nano Science (CENS), Ludwig Maximilians-Universität München, Butenandtstraße 5-13, 81377 München, Germany; Corresponding author
Summary: The NLRP3 inflammasome is a central component of the innate immune system. Its activation leads to formation of the ASC speck, a supramolecular assembly of the inflammasome adaptor protein ASC. Different models, based on ASC overexpression, have been proposed for the structure of the ASC speck. Using dual-color 3D super-resolution imaging (dSTORM and DNA-PAINT), we visualized the ASC speck structure following NLRP3 inflammasome activation using endogenous ASC expression. A complete structure was only obtainable by labeling with both anti-ASC antibodies and nanobodies. The complex varies in diameter between ∼800 and 1000 nm, and is composed of a dense core with emerging filaments. Dual-color confocal fluorescence microscopy indicated that the ASC speck does not colocalize with the microtubule-organizing center at late time points after Nigericin stimulation. From super-resolution images of whole cells, the ASC specks were sorted into a pseudo-time sequence indicating that they become denser but not larger during formation.
