Abstract
Total internal reflection fluorescence microscopy enables to analyze the localizations and dynamics of cellular events that occur at or near the plasma membrane. Total internal reflection fluorescence microscopy exclusively illuminates molecules in the close vicinity of the glass surface, thereby reducing background fluorescence and enabling observation of the plasma membrane in the glass-attached cells with a high signal-to-noise ratio. Here, we describe the application of total internal reflection fluorescence microscopy to analyze the dynamics of caveolae, which play essential physiological functions, including membrane tension buffering, endocytosis, and signaling at the plasma membrane.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Sako Y, Minoghchi S, Yanagida T (2000) Single-molecule imaging of EGFR signalling on the surface of living cells. Nat Cell Biol 2:168–172
Boucrot E, Howes MT, Kirchhausen T, Parton RG (2011) Redistribution of caveolae during mitosis. J Cell Sci 124:1965–1972
Johnson A, Vert G (2017) Single event resolution of plant plasma membrane protein endocytosis by TIRF microscopy. Front Plant Sci 8:612
Mattheyses AL, Simon SM, Rappoport JZ (2010) Imaging with total internal reflection fluorescence microscopy for the cell biologist. J Cell Sci 123:3621–3628
Rosendale M, Perrais D (2017) Imaging in focus: imaging the dynamics of endocytosis. Int J Biochem Cell Biol 93:41–45
Senju Y, Itoh Y, Takano K et al (2011) Essential role of PACSIN2/syndapin-II in caveolae membrane sculpting. J Cell Sci 124:2032–2040
Senju Y, Suetsugu S (2015) Possible regulation of caveolar endocytosis and flattening by phosphorylation of F-BAR domain protein PACSIN2/Syndapin II. BioArchitecture 5:70–77
Sinha B, Köster D, Ruez R et al (2011) Cells respond to mechanical stress by rapid disassembly of caveolae. Cell 144:402–413
Senju Y, Rosenbaum E, Shah C et al (2015) Phosphorylation of PACSIN2 by protein kinase C triggers the removal of caveolae from the plasma membrane. J Cell Sci 128:2766–2780
Tachikawa M, Morone N, Senju Y et al (2017) Measurement of caveolin-1 densities in the cell membrane for quantification of caveolar deformation after exposure to hypotonic membrane tension. Sci Rep 7:7794
Lamaze C, Tardif N, Dewulf M et al (2017) The caveolae dress code: structure and signaling. Curr Opin Cell Biol 47:117–125
Grassart A, Cheng AT, Hong SH et al (2014) Actin and dynamin2 dynamics and interplay during clathrin-mediated endocytosis. J Cell Biol 205:721–735
Shvets E, Bitsikas V, Howard G et al (2015) Dynamic caveolae exclude bulk membrane proteins and are required for sorting of excess glycosphingolipids. Nat Commun 6:6867
Macia E, Ehrlich M, Massol R et al (2006) Dynasore, a cell-permeable inhibitor of dynamin. Dev Cell 10:839–850
Pédelacq J-D, Cabantous S, Tran T et al (2006) Engineering and characterization of a superfolder green fluorescent protein. Nat Biotechnol 24:79–88
Yoshiyuki Arai. Fast single molecule particle tracking and analysis plugin with Java Native Interface (ImageJ User and Developer Conference). http://imagejconf.tudor.lu/program/poster/yoshiyuki_arai73473173
Smith MB, Karatekin E, Gohlke A et al (2011) Interactive, computer-assisted tracking of speckle trajectories in fluorescence microscopy: application to actin polymerization and membrane fusion. Biophys J 101:1794–1804
Jaqaman K, Loerke D, Mettlen M et al (2008) Robust single-particle tracking in live-cell time-lapse sequences. Nat Methods 5:695–702
Acknowledgments
This work was supported by grants from the Funding Program for Next Generation World Leading Researchers (NEXT program) (grant number: LS031); Japan Society for the Promotion of Science (JSPS) KAKENHI (grant numbers: 26291037, 15H01641, 15H05902). Astellas Foundation for Research on Metabolic Disorders to S.S.
Author information
Authors and Affiliations
Corresponding authors
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2020 Springer Science+Business Media, LLC, part of Springer Nature
About this protocol
Cite this protocol
Senju, Y., Suetsugu, S. (2020). Spatiotemporal Analysis of Caveolae Dynamics Using Total Internal Reflection Fluorescence Microscopy. In: Blouin, C. (eds) Caveolae. Methods in Molecular Biology, vol 2169. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-0732-9_6
Download citation
DOI: https://doi.org/10.1007/978-1-0716-0732-9_6
Published:
Publisher Name: Humana, New York, NY
Print ISBN: 978-1-0716-0731-2
Online ISBN: 978-1-0716-0732-9
eBook Packages: Springer Protocols