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Low Copy Numbers of DC‐SIGN in Cell Membrane Microdomains: Implications for Structure and Function
Author(s) -
Liu Ping,
Wang Xiang,
Itano Michelle S.,
Neumann Aaron K.,
de Silva Aravinda M.,
Jacobson Ken,
Thompson Nancy L.
Publication year - 2014
Publication title -
traffic
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.677
H-Index - 130
eISSN - 1600-0854
pISSN - 1398-9219
DOI - 10.1111/tra.12138
Subject(s) - lipid microdomain , biology , dc sign , microscopy , fluorescence microscope , biophysics , microbiology and biotechnology , resolution (logic) , membrane , total internal reflection fluorescence microscope , fluorescence , dendritic cell , antigen , biochemistry , physics , optics , immunology , computer science , artificial intelligence
Abstract Presently, there are few estimates of the number of molecules occupying membrane domains. Using a total internal reflection fluorescence microscopy ( TIRFM ) imaging approach, based on comparing the intensities of fluorescently labeled microdomains with those of single fluorophores, we measured the occupancy of DC‐SIGN , a C‐type lectin, in membrane microdomains. DC‐SIGN or its mutants were labeled with primary monoclonal antibodies ( mAbs ) in either dendritic cells ( DCs ) or NIH3T3 cells, or expressed as GFP fusions in NIH3T3 cells. The number of DC‐SIGN molecules per microdomain ranges from only a few to over 20, while microdomain dimensions range from the diffraction limit to > 1 µm. The largest fraction of microdomains, appearing at the diffraction limit, in either immature DCs or 3 T3 cells contains only 4–8 molecules of DC‐SIGN , consistent with our preliminary super‐resolution Blink microscopy estimates. We further show that these small assemblies are sufficient to bind and efficiently internalize a small (∼50 nm) pathogen, dengue virus, leading to infection of host cells.