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Proceedings Paper

Correlated fluorescence-atomic force microscopy studies of the clathrin mediated endocytosis in SKMEL cells
Author(s): Amy Hor; Anh Luu; Lin Kang; Brandon Scott; Elizabeth Bailey; Adam Hoppe; Steve Smith
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Paper Abstract

Clathrin-mediated endocytosis (CME) is one of the central pathways for cargo transport into cells, and plays a major role in the maintenance of cellular functions, such as intercellular signaling, nutrient intake, and turnover of plasma membrane in cells. The clathrin-mediated endocytosis process involves invagination and formation of clathrin-coated vesicles. However, the biophysical mechanisms of vesicle formation are still debated. Currently, there are two models describing membrane bending during the formation of clathrin cages: the first involves the deposition of all clathrin molecules to the plasma membrane, forming a flat lattice prior to membrane bending, whereas in the second model, membrane bending happens simultaneously as the clathrin arrives to the site to form a clathrin-coated cage. We investigate clathrin vesicle formation mechanisms through the utilization of tapping-mode atomic force microscopy for high resolution topographical imaging in neutral buffer solution of unroofed cells exposing the inner membrane, combined with fluorescence imaging to definitively label intracellular constituents with specific fluorophores (actin filaments labeled with green phalloidin and clathrin coated vesicles with the fusion protein Tq2) in SKMEL (Human Melanoma) cells. An extensive statistical survey of many hundreds of CME events, at various stages of progression, are observed via this method, allowing inferences about the dominant mechanisms active in CME in SKMEL cells. Results indicate a mixed model incorporating aspects of both the aforementioned mechanisms for CME.

Paper Details

Date Published: 21 February 2017
PDF: 6 pages
Proc. SPIE 10071, Single Molecule Spectroscopy and Superresolution Imaging X, 100710I (21 February 2017); doi: 10.1117/12.2256683
Show Author Affiliations
Amy Hor, South Dakota School of Mines and Technology (United States)
Anh Luu, South Dakota School of Mines and Technology (United States)
Lin Kang, South Dakota School of Mines and Technology (United States)
Brandon Scott, South Dakota State Univ. (United States)
Elizabeth Bailey, South Dakota State Univ. (United States)
Adam Hoppe, South Dakota State Univ. (United States)
Steve Smith, South Dakota School of Mines and Technology (United States)

Published in SPIE Proceedings Vol. 10071:
Single Molecule Spectroscopy and Superresolution Imaging X
Jörg Enderlein; Ingo Gregor; Zygmunt Karol Gryczynski; Rainer Erdmann; Felix Koberling, Editor(s)

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